Gulden Camci-Unal, University of Massachusetts Lowell
Surya Mallapragada, Iowa State University
Matteo Moretti, IRCCS Insituto Ortopedico Galeazzi
Pamela Yelick, Tufts University
Multifunctional Materials | IOP Publishing
BM04.01: Biomaterials for Regeneration of Tissues I
Monday AM, November 27, 2017
Sheraton, 2nd Floor, Independence West
8:00 AM - BM04.01.01
Conductive Collagen—Mimetic Foams for Cardiac Tissue Regeneration
Worrapong Kit-Anan 1 2 3 , Paresh Parmar 1 2 3 , Muzamir Mahat 1 2 3 , Manuel Mazo 1 2 3 , Anna Regoutz 1 , Astrid Armgarth 1 2 3 , Violet Stoichevska 4 , David Payne 1 , Yong Peng 4 , Jerome Werkmeister 4 , John Ramshaw 4 , Sian Harding 5 , Cesare Terracciano 5 , Molly Stevens 1 2 3 Show Abstract
1 Materials, Imperial College London, London United Kingdom, 2 Bioengineering, Imperial College London, London United Kingdom, 3 Institute of Biomedical Engineering, Imperial College London, London United Kingdom, 4 , CSIRO Manufacturing Flagship, Clayton, Victoria, Australia, 5 National Heart and Lung Institute, Imperial College London, London United Kingdom
Cardiovascular diseases are the top cause of mortality worldwide, accounting for over 30% of total deaths, and the limited supply of donor hearts cannot combat the high demand for transplants . Cardiac tissue engineering to replace the damaged tissue still presents significant challenges due to the complex and dynamic interplay of electrical and biomechanical signals involved in the development and physiology of the myocardium. Conductive polymers are of particular interest for this application since they can reinstate the lost electrical activity, as demonstrated in recent findings from our group . However, they need further optimisation in order to be applicable as a cardiac patch.
Here, we developed a fabrication approach that immobilises the electrically-active “dopant” in a streptococcal collagen-like 2 (Scl2) construct of physiologically-relevant elasticity , generating a hybrid polyaniline (PANI)-Scl2 scaffold. Using X-ray photoelectron spectroscopy, we prove that the strong chelation between phytic acid and Scl2 leads to the formation of conductive constructs with retained electroactivity, low surface resistivity (10-2 S cm-1 range), and oxidised form after a week in physiological media. The PANI-Scl2 and Scl2 showed no cell toxicity when interfaced with electrosensitive human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Gene expression showed a reduction of cardiac ion channels RYR2 and KCNJ2 in PANI-Scl2 (N=3, p=0.013). The MYH6/MYH7 ratio showed a significant increase in Scl2, indicating an increase in maturity level (N=3, p=0.023) while maintaining insignificant difference in the cardiac subtype indicator, MYL2/MYL7 ratio. Sarcomere length and alignment, while not different between the two materials, was close to the value reported for cultured adult human CMs. Calcium transients, a determinant of cardiac functionality, showed significant differences in the duration of the transient (N=3, p=0.042), taking longer for PANI-Scl2 to contract and reinforcing the basis for the maturation effect provided by the PANI-Scl2 construct.
These findings overcome a major hurdle for the application of PANI in bioelectronic devices over extended operational times and demonstrate the feasibility of this material-based strategy to induce a set of mature features in hiPSC-CMs by matching the physical and electrical properties of the scaffold with those of the tissue. Further work regarding the ability to encapsulate cells in hybrid scaffolds paves the way for applying this system in an in vivo context for cardiac regeneration applications.
 Mendis S, et. al. World Health Organisation; 2011.
 Mawad, D., et al. (2016). A conducting polymer with enhanced electronic stability applied in cardiac models. Science Advances, 2(11), pp.e1601007-e1601007.
 Parmar PA, et al. Collagen-mimetic peptide-modifiable hydrogels for articular cartilage regeneration. Biomaterials 2015;54:213-25.
8:15 AM - BM04.01.02
Enhancing the Adhesion Strength of Nanotubular Oxide Layer onto Ti6Al7Nb Substrate for Bone-Tissue Engineering Applications
Batur Ercan 1 2 , Merve Izmir 1 Show Abstract
1 Metallurgical and Materials Engineering, Middle East Technical University, Ankara Turkey, 2 Biomedical Engineering, Middle East Technical University, Ankara Turkey
Titanium and its alloys have been used in orthopedic applications for over 50 years. Although titanium alloys have high specific density, excellent corrosion resistance and optimal mechanical strength, their bioactivity still needs to be improved. To enhance bioactivity of titanium alloy surfaces, nanostructures with different nanotubular morphologies have been grown on titanium alloys via an electrochemical process called anodization. Anodized titanium alloys were found to enhance bone cell adhesion, proliferation and long-term cellular functions, which are important indicator of increased biocompatibility of the alloy surfaces. Though anodized surfaces have enhanced bone cell functions in vitro and osseointegration in vivo, poor adhesion and delamination of the oxide-based nanotubular layer onto the titanium substrate restricted the use of anodized materials in clinical applications.
In our study, Ti6Al7Nb samples were anodized and nanotubular surface morphologies tailored to possess diameters ranging 25nm to140nm were observed using a mixture of 1.4 M H3PO4 and 0.02 wt% HF electrolyte solution. When the adhesion strength of these samples were measured using a scratch test, adhesive forces ranging 0.3 N to 1 N were obtained for different tubular diameters upon scratching a 2-mm-long region. This result confirmed the weakly adherent nature of the nanotubular oxide layer onto the bulk Ti6Al7Nb alloy. To enhance the adhesion strength of the oxide layer, while maintaining optimal implant performance in vivo by using an oxide layer thickness above 600nm, a post-treatment process with cyclohexane was applied to the anodized nanotubular Ti6Al7Nb surfaces having different nanostuctured diameters. When nanotubular layer adhesion of cyclohexane treated anodized Ti6Al7Nb samples having 1 µm thick oxide layer were examined with a scratch test, adhesive forces raging 10-30N was observed, indicating up to 30 folds increase in adhesion stress (an indication of layer cohesion) between the Ti6Al7Nb alloy substrate and grown oxide layer. The results were attributed to hydrogen-assisted cracking mechanism and the cyclohexane post-treatment was proposed to decrease the efficacy of the embrittlement process. Importantly, in vitro tests using bone cells indicated that the applied post treatment did not alter the biocompatibility of the anodized surfaces.
To conclude, post-treatment process using cyclohexane enhanced the layer adhesion between anodized oxide layer with the Ti6Al7Nb substrate up to 30 folds in a wide diameter range between 50 nm to 200 nm, while maintaining biocompatible characteristics of the anodized surface. Thus, cyclohexane post-treatment on anodized surfaces is a potential candidate to enhance bioactivity of Ti6Al7Nb alloys for bone-tissue engineering applications.
8:30 AM - BM04.01.03
Advanced Antibacterial and Bioactive Glass-Ceramic Particles for Tissue Healing and Regeneration
Natalia Pajares 1 , Yadav Wagley 2 , Neal Hammer 1 , Kurt Hankenson 2 , Xanthippi Chatzistavrou 1 Show Abstract
1 , Michigan State University, East Lansing, Michigan, United States, 2 , University of Michigan, Ann Arbor, Michigan, United States
Bone infections by multi-drug resistant bacterial strains are a global threat. Staphylococcus aureus (S. aureus) is the most common bacterium in bone infectious diseases. S. aureus has been characterized by the World Health Organization (WHO) as a high priority bacterium, and multidrug-resistant strains are prevalent in both the hospital and community settings. Unfortunately, the efficacy of traditional antibiotics is dramatically reduced against these resistant strains. There is an urgent need for developing alternative antibacterial approaches that could promote new bone formation while combating bacteria in osteomyelitis without the development of resistance. We have developed novel Ag-doped bioactive glass (Ag-BG) particles with antibacterial and bioactive properties using the SiO2-CaO-P2O5-Al2O3-Na2O-K2O-Ag2O system. The aim of our research was to determine whether Ag-BG particles loaded with vancomycin (VAN/Ag-BG) can enhance cell proliferation and differentiation of osteoblasts in vitro and moreover can combat S. aureus utilizing synergism between silver ions and the drug. Ag-BG particles were fabricated by the sol-gel (solution-gelation) method and characterized microstructurally and morphologically using the Fourier Transform Infrared Spectroscopy (FTIR), X-Ray Diffraction analysis (XRD) and Scanning Electron Microscopy (SEM). The cell-material interaction was studied using human marrow-derived mesenchymal progenitor cells. Cell proliferation and differentiation to osteoblast was studied for different concentrations of Ag-BG. The antibacterial activity of Ag-BG alone, vancomycin alone and combination of both VAN/Ag-BG was observed against S. aureus. Different concentrations of Ag-BG were used. The Minimum Bactericidal and Inhibitory Concentrations (MBC and MIC respectively) were measured. Our studies showed that the new Ag-BG particles can significantly enhance cell proliferation and differentiation to osteoblasts compared to control (cells cultured alone). Enhanced cell differentiation was observed for all co-cultures of cells with different concentrations of Ag-BG in both cultures with osteogenic and traditional growth medium. Finally, we found bacteria inhibition for all concentrations of Ag-BG, while the MBC was 7mg of Ag-BG. Interestingly, vancomycin did not inhibit bacteria growth when the co-culture was in Phosphate Buffered Saline (PBS). However, the combined delivery of VAN/Ag-BG significantly increased S. aureus inhibition compared to Ag-BG alone in PBS. In conclusion, the novel Ag-BG particles and moreover the combined VAN/Ag-BG present advanced antibacterial properties and enhanced biological behavior. This work is a foundation for further studies that can potentially advance treatment options in orthopedic infections.
8:45 AM - *BM04.01.04
Regenerative Engineering—The Convergence Quest
Cato Laurencin 1 2 3 , Naveen Nagiah 1 Show Abstract
1 , University of Connecticut Health Center, Farmington, Connecticut, United States, 2 Department of Chemical and Biomolecular Engineering, University of Connecticut, Storrs, Connecticut, United States, 3 Department of Materials Science and Engineering, University of Connecticut, Storrs, Connecticut, United States
We define Regenerative Engineering as a Convergence of Advanced Materials Science, Stem Cell Science, Physics, Developmental Biology, and Clinical Translation. We believe that an “un-siloed’ technology approach will be important in the future to realize grand challenges such as limb and organ regeneration. We also believe that biomaterials will play a key role in achieving overall translational goals. Through convergence of a number of technologies, with advanced materials science playing an important role, we believe the prospect of engaging future grand challenges is possible.
Regenerative Engineering as a field is particularly suited for solving clinical problems that are relevant today. The paradigms utilized can be applied to the regeneration of tissue in the shoulder where tendon and muscle currently have low levels of regenerative capability, and the consequences, especially in alternative surgical solutions for massive tendon and muscle loss at the shoulder have demonstrated significant morbidity. Polymer, polymer-cell, and polymer biological factor, and polymer-physical systems can be utilized to propose a range of solutions to shoulder tissue regeneration. The approaches, possibilities, limitations and future strategies, allow for a variety of clinical solutions in musculoskeletal disease treatment.
9:45 AM - *BM04.01.05
Pre-Clinical Development of EpiBone's Tissue Engineered Bone Graft for Cranial and Maxillofacial Reconstruction
Sarindr Bhumiratana 1 Show Abstract
1 , EpiBone, Inc, Brooklyn, New York, United States
In the United States alone, there are over 1 million surgical procedures per year requiring a bone graft. Of which, craniomaxillofacial (CMF) reconstruction accounts for almost 10% of all procedures. CMF reconstruction is a significant challenge due to the complex bone geometries and the need to restore both the aesthetics and function. EpiBone’s mission is to advance skeletal reconstruction through personalized 3D design and living cells. This is enabled by core proprietary technologies such as fabrication methods and bioreactor systems. The fabrication methods enable graft customization, and the bioreactor systems are critical for culturing clinically-sized tissue engineered constructs. EpiBone’s approach follows the concept of “personalization”: bioreactors and scaffolds are shaped according to 3D reconstructions of patient-specific clinical images to create tissue engineered constructs that replicate the actual geometry of the patients’ defects. We previously demonstrated that living tissue engineered bone graft is safe and effective for promoting bone regeneration in a rat calvarial defect model and a pig ramus condyle unit (RCU) defect model. To advance the technology toward clinical trial, we have developed manufacturing process, built our GMP manufacturing facility, and conducted a pivotal GLP study. This talk will cover the overview pre-clinical development of our first EpiBone-CMF graft, a living tissue engineered bone graft for cranial and maxillofacial reconstruction.
10:15 AM - BM04.01.06
Tuning Stem Cell Fate by Nanostructure Mediated Physical Signals for Tissue Engineering and Regeneration
Hong Liu 1 2 Show Abstract
1 State Key Laboratory of Crystal Materials, Shandong University, Jinan, Shandong, China, 2 Institute for Advanced Interdisciplinary Research (IAIR), University of Jinan, Jinan, Shandong, China
Besides the biological growth factors, small organic molecules, and chemical ions, physical signals is the other category of very important factors to tune/regulate the fate of stem cells. Recent years, more attention has been paid on the differentiation of stem cells on the physical signal, including, localized electric or magnetic field, surface topology of biomaterials, photo irradiation, and even pressure and strain from the materials. With progress of research in this field, some cues of connection between physical signal and bio pathway for differentiation have been discovered. However, more phenomena have still not been understood. Because the physical signals possess controllability and can be localized in a specific area, they are benefit to be used in tissue engineering for tissue regeneration. Therefore, finding new physical approaches for regulation fate of stem cells is a great challenge for alive biomaterials design and applications.
Recent year, some novel phenomena about the effect of physical signal on stem cell differential has been noticed. For example, nano-network morphology of HAp film can induce differentiation of bone marrow mesenchymal stem cells (MSCs) to vascular endothelial cells, surface charges on LiNbO3 wafer can regulate MSCs differentiate to osteogenic cells, and a pressure from biomaterials can differentiate MSCs to neural cells.
In this talk, we will report the above works, and try to explain the reasons for physical signal induced differentiation from both physical mechanism and bio pathways. We believe that the regulation effect of physical signal will attract more attention, and will have great impact for design and application of biomaterials, especially for tissue engineering scaffold, and will bring great progress in tissue regeneration medicine.
*H. Liu, tel: +86-88362807; firstname.lastname@example.org
 J. Li, H. Liu, et. al Nanoscale, 2016, 8, 7416-7422.
 W. Guo, H. Liu, e. al., ACS Nano, 10, 5086-5095
 J. Qiu, H. Liu, et. al., Small, 2016, 10, 1770-1778
 X. Mu, H. Liu, et. al. Nanoscale, 2016, 8, 13186-13191
 X. Mu, H. Liu, et. al. Nanoscale, 2017 (accepted)
10:30 AM - BM04.01.07
Piezo- and Magnetoelectric Polymers as Biomaterials for Novel Tissue Engineering Strategies
Clarisse Ribeiro 1 , DM Correia 1 , Sylvie Ribeiro 1 , Senentxu Lanceros-Mendez 1 2 3 Show Abstract
1 , University of Minho, Braga Portugal, 2 , BC Materials, Bilbao Spain, 3 , Ikerbasque, Bilbao Spain
Tissue engineering and regenerative medicine are increasingly taking advantage of active materials, allowing to provide specific clues to the cells. In particular, the use of electroactive polymers that deliver an electrical signal to the cells upon mechanical solicitation, open new scientific and technological opportunities, as they in fact mimic signals and effects present in living tissues, allowing the development of suitable microenvironments for tissue regeneration. Piezoelectric polymers have already shown strong potential for novel tissue engineering strategies, once they can account for the existence of piezoelectricity within some specific tissues, indicating their requirement also during tissue regeneration. Further, they can modulate the electrical signals existing in tissue development and function. Still, in some cases, the patient is immobilized, and as a result the natural mechanical stimulus does not occur. Such limitation points to the development of new materials able to remotely mechanical and/or electrically stimulate tissues from outside of the human body and/or during in-vitro cell culture to explore specific differentiation paths. Magnetoelectric composite materials provide such an innovative tool, allowing the use of an external magnetic field to remotely control tissue stimulation, without the need of patient movement. Those composites are composed of magnetostrictive and piezoelectric materials.
Thus, a novel overall strategy for bone and muscle tissue engineering was developed based on the fact that these cells type are subjected to mechano-electrical stimuli in their in vivo microenvironment and that piezo- and magnetoelectric polymers, used as scaffolds, are suitable for delivering those cues. The processing and functional characterizations of piezoelectric and magnetoelectric polymers based on poly(vinylindene fluoride) and poly-L-lactic acid in a variety of shapes, from microspheres to electrospun matts and three differential scaffolds, will be shown as well as their performance in the development of novel bone and muscle tissue engineering. Further, the development of specific bioreactors for in-vitro tests will be presented.
The authors thank the Portuguese Fundação para a Ciência e Tecnologia (FCT) for financial support under Strategic Funding UID/FIS/04650/2013 and project PTDC/EEI-SII/5582/2014, including FEDER funds, UE. The authors also thank the FCT for financial support under grants SFRH/BPD/90870/2012 (CR), SFRH/BPD/121526/2016 (DMC). Financial support from the Spanish Ministry of Economy and Competitiveness (MINECO) through the project MAT2016-76039-C4-3-R (AEI/FEDER, UE) (including the FEDER financial support) and from the Basque Government Industry Department under the ELKARTEK Program is also acknowledged.
10:45 AM - BM04.01.08
Drug Delivered Poly(ethylene glycol) Diacrylate (PEGDA) Hydrogels and Their Mechanical Characterization Tests for Tissue Engineering Applications
Kerolos Hanna 1 , Ozgul Yasar-Inceoglu 2 , Ozlem Yasar 1 Show Abstract
1 Mechanical Engineering Technology, City University of New York, Brooklyn, New York, United States, 2 Mechanical Engineering, California State University, Chico, California, United States
Tissue Engineering has been studied to develop tissues as an alternative approach to the organ regeneration. Successful artificial tissue growth in regenerative medicine depends on the precise scaffold fabrication as well as the cell-cell and cell-scaffold interaction. Scaffolds are extracellular matrices that guide cells to grow in 3D to regenerate the tissues. Cell-seeded scaffolds must be implanted to the damaged tissues to do the tissue regeneration. Scaffolds’ mechanical properties and porosities are the two main scaffold fabrication parameters as the scaffolds must be able to hold the pressure due to the surrounding tissues after the implantation process. In this research, scaffolds were fabricated by photolithography and Poly(ethylene glycol) Diacrylate (PEGDA) which is a biocompatible and biodegradable material was used as a fabrication material. In order to compare the compressive properties of PEGDA only with the compressive properties of drug delivered PEGDA, firstly, PEGDA only solutions were prepared. Then, PEGDA was mixed with Meloxicam 15 mg, Hydrochlorothiazide 12.5 mg, Cyclobenzaprine 10 mg and Spironolactone-hctz 25-25 mg respectively and they were placed under the UV light for about 15 minutes to solidify the cylindrical shaped hydrogels. 5 samples from each group were fabricated under the same conditions. Laboratory temperature, photoinitiator concentration and UV light intensity was kept constant during the fabrication process. After the fabrication was completed, Instron 3369 universal mechanical testing machine with the 5 mm/min compression rate was used to do the compression tests to compare the drug effects on PEGDA hydrogels. Our results indicate that average ultimate strength of PEGDA only samples was 3.820 MPa. Also, due to the fact that Meloxicam 15 mg and PEGDA mixture did not solidify under the UV light at all, compression test could not be performed for PEGDA- Meloxicam 15 mg mixture. However, Hydrochlorothiazide 12.5 mg, Cyclobenzaprine 10 mg and Spironolactone-hctz 25-25 mg dissolved within the PEGDA completely and our compression results show that average ultimate strengths were 3.372 MPa, 1.602 MPa, 1.09 MPa respectively. This preliminary research showcases that compressive properties of the PEGDA-based photopolymerized scaffolds can be altered with the control of the drug type and drug concentration.
11:00 AM - BM04.01.09
Engineering Highly Conductive and Biocompatible Hydrogels for Cardiac Tissue Regeneration
Ehsan Shirzaei Sani 1 , Roberto Portillo Lara 1 2 , Brian Walker 1 , Nasim Annabi 1 3 4 Show Abstract
1 , Northeastern University, Boston, Massachusetts, United States, 2 Centro de Biotecnología FEMSA, Tecnológico de Monterrey, Monterrey, NL, Mexico, 3 Biomaterials Innovation Research Center, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts, United States, 4 Harvard-MIT Division of Health Sciences and Technology, Massachusetts Institute of Technology, Boston, Massachusetts, United States
Heart failure following myocardial infarction (MI) is the number one cause of death in the US. Due to the inability of the adult heart to self-regenerate, the irreversible loss of functional myocardium after MI often leads to heart failure and death. In recent years, injectable hydrogel therapy has emerged as an attractive strategy to deliver cellular- and pharmacological-based therapies for the regeneration of the damaged heart. Due to the insulating nature of polymeric hydrogels and the responsiveness of cardiac tissues to electrical stimuli, electroconductive hydrogels (ECHs) could be used to restore electrophysiological coupling to the affected area and thus, enhance therapeutic efficacy. However, conventional ECHs are often limited by the cytotoxicity, and poor solubility, processability, and biodegradability of their components. Here, we describe a novel method to engineer photocrosslinkable ECHs through the functionalization of different polymers with a conductive choline-based bio-ionic liquid (Bio-IL). Bio-IL conjugated hydrogels exhibited a wide range of highly tunable mechanical and conductive properties, as well as remarkable biocompatibility both in vitro and in vivo. In particular, hydrogels made from Bio-IL functionalized gelatin methacryloyl (GelMA) were shown to support the growth and function of primary cardiomyocytes in both two- and three-dimensional cultures in vitro. GelMA/Bio-IL hydrogels were also shown to restore the propagation of electrical stimuli across severed skeletal muscle tissues ex vivo. Subcutaneous implantation in a murine animal model demonstrated that GelMA/Bio-IL hydrogels could be efficiently biodegraded without eliciting any significant immunogenic response in vivo. Taken together, these results demonstrate the potential of GelMA/Bio-IL ECHs to be used for regenerative cardiac tissue engineering applications. Moreover, Bio-IL functionalization constitutes a versatile and efficient approach for the engineering of ECHs for a wide range of biomedical applications.
11:15 AM - BM04.01.10
Theranostics-Embedded Multifunctional Tissue Engineering Scaffolds for Cancer Detection and Treatment
Lin Guo 1 , Min Wang 1 Show Abstract
1 Department of Mechanical Engineering, The University of Hong Kong, Hong Kong Hong Kong
Electrospun nanofibrous scaffolds are popularly used in tissue engineering owing to their distinctive advantages. For post-surgery cancer patients, these scaffolds can be employed to regenerate tissue at the original tumor site after tumor resection. The high recurrence rate of cancer threatens lives of post-surgery patients, and hence effective detection and treatment of recurrent cancer become vitally important. In recent years, gold nanoparticle (AuNP)-based theranostics have attracted great attention for cancer detection and treatment due to their unique properties including surface enhanced Raman scattering (SERS) effect. The strongly amplified SERS signals can be used for high-sensitivity cancer detection. Additionally, AuNP-based theranostics can convert light into heat, providing photothermal therapy for cancers. In this study, multifunctional tissue engineering scaffolds incorporated with AuNP-based theranostics were fabricated, aiming to achieve both tissue regeneration and detection and treatment of recurrent cancer. Folic acid-conjugated chitosan (CS-FA) was firstly made and the folic acid ligand in CS-FA would provide cancer cell targeting ability. CS-FA-capped AuNPs (Au@CS-FA) were then synthesized with highly branched AuNP core and cross-linked CS-FA shell. A Raman reporter, Rhodamine 6G (R6G), was incorporated in Au@CS-FA for generating SERS signals. Concurrent electrospinning and co-axial electrospray were used to fabricate multifunctional scaffolds. Au@CS-FA theranostics were encapsulated in core-shell structured PLGA50/50 microspheres embedded in PLGA75/25 scaffolds. TEM and SEM results revealed that theranostics were well encapsulated in microspheres and the microspheres with uniform size were randomly distributed in scaffolds. In vitro immersion tests showed that controlled release of theranostics could be achieved when PLGA shell of microspheres gradually degraded. SERS measurements were conducted and high SERS signals by theranostics before encapsulation and after release were observed, indicating cancer detection ability of theranostics. In vitro biological experiments were conducted to investigate cancer cell targeting and photothermal treatment of released theranostics. The experiments used HeLa cells, which exhibited high-level folate-receptor expression, with MCF-7 cells as the control. It was shown that theranostics bounded to HeLa cells rather than MCF-7 cells. For HeLa cells treated with theranostics-incorporated scaffolds, after irradiation by a NIR laser, Live/Dead viability tests were conducted. High death rates of HeLa cells due to heat generated by theranostics were observed. In contrast, for HeLa cells irradiated under the same conditions but treated with PLGA scaffolds without theranostics, high degree of cell survival was observed. These results indicated that the released theranostics were effective photothermal agent for cancer treatment.
BM04.02: Biomaterials for Regeneration of Tissues II
Monday PM, November 27, 2017
Sheraton, 2nd Floor, Independence West
1:30 PM - BM04.02.01
3D Tissue Engineering by Tuning Porous Carbon and Hydrogel Scaffolds
Mohammadreza Taale 1 , Fabian Schütt 1 , Yogendra Mishra 1 , Rainer Adelung 1 , Christine Selhuber-Unkel 1 Show Abstract
1 , Kiel University, Kiel Germany
The microscopic three-dimensional geometry of a biomaterial is a key element in providing spatial organization for cell growth and appropriate nutritional conditions. Scaffolds containing fully interconnected pore structures fulfill the geometrical requirements for tissue engineering by mimicking the microstructure of natural extra cellular matrix.
To generate such scaffolds, micron-sized ZnO tetrapods can serve as a sacrificial template and have been employed for a range of porous material types. Several approaches are possible, as the ZnO can be coated with various types of materials and is later hydrolyzed or converted to graphite. Therefore, by using different types of coating materials, various biochemical signals may be introduced through the porous scaffold to provide appropriate conditions for 3D cell culture applications. On the other hand, physical and mechanical properties of fabricated microstructures can be widely tuned via combination of different fabrication processes such as nanoparticle infiltration and chemical vapor deposition (CVD) for ZnO to graphite conversion .The flexibility in tuning mechanical properties, electrical conductivity, biochemical activity, geometry, porosity and size of the scaffolds provide excellent opportunities to develop artificial tissues. We verified the biocompatibility of the scaffolds by measuring the protein absorption (BCA assay) and toxicity assay (MTT). The feasibility of fabricated scaffolds for cell growth have been confirmed through cell proliferation assays (WST-1) and microscopic observation of cultured cells on the scaffolds.
Another approach to provide suitable 3D scaffolds for cell growth are porous hydrogels, based on inverted ZnO tetrapod networks. A hydrogel with interconnected microchannels has successfully been employed and adopted in order to accommodate cell migration and growth inside the hydrogel. Both stiffness and structural shape of the microchannel containing hydrogel can be tuned.
In conclusion, structures generated by our approach of using ZnO tetrapods as sacrificial materials for either generating micro-fibers or micropores provide excellent materials for investigating cell behavior in 3D matrices and studying 3D cellular mechanotransduction.
Reference:  C. Lamprecht, M. Taale, et al. (2016), ACS Applied Materials & Interfaces, 8:14980-14985.
1:45 PM - BM04.02.02
Freeze-Cast, Biomimetic Multifunctional Core-Shell Device for Peripheral Nerve Repair
Kaiyang Yin 1 , Prajan Divakar 1 , Jennifer Hong 2 , Karen Moodie 1 , Joseph Rosen 1 , Michael Matthew 1 , Ulrike Wegst 1 Show Abstract
1 , Dartmouth College, Hanover, New Hampshire, United States, 2 , Dartmouth–Hitchcock Medical Center, Lebanon, New Hampshire, United States
An estimated 2 million (5%) of the annual 40.2 million trauma-related emergency room admissions in the US require treatment for peripheral nerve injuries. These injuries occur in upper and lower extremities as well as in facial nerves, and frequently result in permanent disabilities such as sensory deficit, paralysis, and painful neuropathies. Although peripheral nerves possess the inherent ability to regenerate, nerve transections require surgical intervention to optimize functional recovery. While small gaps (≤5 mm) may be directly coapted, larger gaps (>5 mm) require interposition grafting with nerve auto- or allografts or conduits to provide tension-free mechanical support, axon guidance, and protection from fibrous tissue ingrowth. Conduits are devices that avoid the substantial disadvantages of autografts such as a second surgical site, donor site morbidity, limited availability, and high treatment cost, however, they still have limited regenerative capacity of neurons. As a result, autografts remain the gold standard.
Presented will be the design, manufacture, and in vivo assessment of a novel multifunctional freeze-cast core-shell device for peripheral nerve repair with highly-aligned, continuous porosity that emulates key structural and mechanical functions of the natural tissue. The in vivo efficacies of the core-shell assembly, the shell alone, and as control the inverted nerve autograft were evaluated in a Lewis rat sciatic nerve model at time points of 4, 8 and 12 weeks after implantation. Walking track analysis was performed to assess nerve function every two weeks after the 4th week; biocompatibility and regenerative efficacy of different scaffold materials and designs were analyzed through the histological sections at each time point. The core-shell design was found to have the most promising regenerative characteristics, promoting through a combination of structural, mechanical and chemical cues the desired neovascularization, axonal ingrowth and alignment.
2:00 PM - BM04.02.03
Paper-Based Cell Culture Platforms for Personalized Medicine and Regenerative Engineering
Gulden Camci-Unal 1 Show Abstract
1 , University of Massachusetts Lowell, Lowell, Massachusetts, United States
Traditional tissue engineering models use sophisticated instrumentation or costly set-ups for fabrication of 3D scaffolds, require extensive optimization procedures, and do not provide physiologically relevant size structures without mass transport limitations. Therefore, it is challenging to fabricate biocompatible scaffolds for personalized medicine. To tackle these hurdles, we developed paper-based cell culture platforms for a range of applications that involve the use of different types of cells (e.g. stem cells, primary cells, cells from patient biopsies, immune cells, fibroblasts, osteoblasts, epithelial cells, tumor cells, bacteria, fungi, plant cells). Paper-based cell culture platforms are flexible, tunable, simple, low-cost, and amenable to high-throughput sample preparation and analysis. Herein, we used paper-based scaffolds as matrices to support cells and/or hydrogels in 3D. After the desired cell culture period, we monitored and characterized the behavior of cells through standard analytical assays such as cytotoxicity, metabolic activity, proliferation, DNA content, protein content, apoptosis, immunostaining, high-resolution imaging, or mechanical tests.
We fabricated paper-based scaffolds for different applications in personalized medicine and regenerative engineering. For example, we investigated migration of primary human tumor cells that were isolated from patient biopsies in a multilayered paper-based cell culture platform. This approach can be adapted to screen different doses of chemotherapeutics or radiation in a patient-specific fashion. We also used paper scaffolds to induce template-guided biomineralization in origami-inspired structures. This method can be used to fabricate constructs for patients who have irregular size and shape bone defects. In addition, we generated wax-printed patterns in paper scaffolds that are for high-throughput sample preparation and analysis of osteoblast cultures. We have shown that we can form and control gradients of oxygen, nutrients, and other biological molecules in paper-based cell culture platforms. We also used these systems to culture bacteria, fungi, and plant cells and developed in vitro disease models. Our results demonstrated that the paper scaffolds enable patterning from micron to cm scale, adapt modular configurations, and can provide physiologically relevant tissue models. Paper scaffolds can also be used for origami-inspired tissue engineering.
We developed paper-based cell culture platforms to obtain multicellular and compartmentalized tissue-mimetics for clinical applications. To overcome the major limitations of the traditional tissue models, we adapted a layer-by layer strategy to assemble tissue-like structures from low-cost and biocompatible paper-based materials. This approach offers unique opportunities from understanding fundamental biology to developing disease models for personalized medicine, and assembling different tissues for organ-on-paper configurations.
2:15 PM - BM04.02.04
A Strain-Stiffening Polymer with Advanced Properties for Tissue Engineering—Let Nature Help Us
Michael Timmermann 1 , Christine Selhuber-Unkel 1 Show Abstract
1 , University of Kiel - Institute for Materials Science - Biocompatible Nanomaterials, Kiel Germany
When designing materials for tissue engineering, a typical strategy is to mimic the in vivo environment of cells. A drastically underestimated property is the non-linear mechanical behavior of body tissues. In fibrin and collagen type I gels, the elastic modulus increases by orders of magnitude in response to applied strain [1,2]. Studies have shown that NIH 3T3 fibroblasts and hMCSs react to this strain stiffening not only with advanced spreading but also with an increased communication distance, which might be a key to pattern formation during tissue development .
Materials that mimic this behavior for tissue engineering are nowadays mostly limited to soft hydrogels. Instead, we use a cell-inspired approach to make the strain-stiffening effect applicable to elastic materials regardless of their initial stiffness. This could open the field for a whole variety of artificially grown body tissues.
Cells react to external forces by reversibly cross-linking cytoskeletal fibers and therefore increasing their stiffness. The material we are developing mimics this behavior. Our material has a specially designed microstructure so that deformation leads to a reversible interconnection of internal surfaces and, due to higher friction between these surfaces, to a stiffening of the material as a whole. This increased friction arises either from the material's tack or can be increased by reversible biological binding-systems. FEM analysis has been used to optimize the sample shape to achieve a maximum in strain-stiffening. In addition, our material is biocompatible and has the potential to be fabricated with high-throughput methods.
A successful implementation of this approach could revolutionize the field of strain-stiffening materials and might find applications not only in biomaterials for tissue engineering, but also in classical engineering as special macroscopic shock absorbing structures.
 Storm C et al. (2005) Nonlinear elasticity in biological gels. Nature 435: 191–194.
 Janmey PA et al. (2007) Negative normal stress in semiflexible biopolymer gels. Nat Mater 6: 48–51.
 Winer JP et al. (2009) Non-Linear Elasticity of Extracellular Matrices Enables Contractile Cells to Communicate Local Position and Orientation. PLoS ONE 4(7): e6382.
2:30 PM - *BM04.02.05
Molecularly and Cellularly Imprinted, Intelligent Scaffolds for Tissue Engineering
Nicholas Peppas 1 , John Clegg 1 , Marissa Wechsler 1 Show Abstract
1 , The University of Texas at Austin, Austin, Texas, United States
The development of molecularly imprinted polymers using biocompatible production methods enables the possibility to further exploit this technology for biomedical applications. Tissue engineering (TE) approaches use the knowledge of the wound healing process to design scaffolds capable of modulating cell behavior and promote tissue regeneration. Biomacromolecules bear great interest for TE, together with the established recognition of the extracellular matrix, as an important source of signals to cells, both promoting cell-cell and cell-matrix interactions during the healing process. This review focuses on exploring the potential of protein molecular imprinting to create bioactive scaffolds with molecular recognition for TE applications based on the most recent approaches in the field of molecular imprinting of macromolecules. Considerations regarding essential components of molecular imprinting technology will be addressed for TE purposes. Molecular imprinting of biocompatible hydrogels, namely based on natural polymers, is also reviewed here. Hydrogel scaffolds with molecular memory show great promise for regenerative therapies. The first molecular imprinting studies analyzing cell adhesion report promising results with potential applications for cell culture systems, or biomaterials for implantation with the capability for cell recruitment by selectively adsorbing desired molecules.
3:30 PM - *BM04.02.06
Guiding Tissue Regeneration through Biofabrication—The Role of Biomaterials Chemistry and Multiscale Structural Properties
Lorenzo Moroni 1 Show Abstract
1 , Maastricht University, Maastricht Netherlands
A key factor in scaffold-based tissue and organ regeneration relies on controlling (stem) cell-material interactions to obtain the same original functionality. Different approaches include delivery of biological factors and surface topography modifications. Although both strategies have proved to augment cell activity on biomaterials, they are still characterized by limited control in space and time, which hampers the proper regeneration of complex tissues. Here, we present a few examples where tbiofabrication technology platforms allowed the generation of a new library of 3D scaffolds with tailored biological, physical, and chemical cues at different scales.
By engineering their topological properties, these porous biomaterials influence the activity of seeded cells, thereby initiating the regeneration of the targeted tissues. Future efforts should aim at further improving our understanding of scaffold topological properties to achieve a fine control on cell fate at multiple scales. This will enable the regeneration of complex tissues including vasculature and ineural networks, which will result in enhanced in vivo integration with surrounding tissues. By doing so, the gap from tissue to organ regeneration will be reduced, bringing regenerative medicine technologies closer to the clinics.
4:00 PM - BM04.02.07
Progress in the Development of Bioengineered Teeth and Jaw Bone
Pamela Yelick 1 , Weibo Zhang 1 , Elizabeth Smith 1 Show Abstract
1 , Tufts University, Boston, Massachusetts, United States
Currently preferred methods to repair craniofacial jaw bone and tooth defects require multiple surgical procedures to first acquire autologous bone (i.e. iliac crest, rib, tibia), and then to repair the jaw bone defect, followed by an extended healing time. Once, and if, sufficient jaw bone has been regenerated, additional surgical procedures are required to first place synthetic titanium implants, and finally to place posts and crowns after the dental implant has securely ankylosed with the jaw bone (~ 3 months). This approach, although sufficient for many individuals, is not ideal in that it creates donor site morbidity, requires an extended period of time (~1 year with no complications), and does not regenerate tissues exhibiting important properties of natural teeth, including proprioception, vitality, and compatible physical properties. To improve upon these current gold standard clinical approaches, we propose Tissue Engineering strategies to create bioengineered jaw bone and teeth that better reflect natural tissue properties, in a more timely and effective manner. Our methods include the use of a variety of scaffold materials and designs, and post-natal dental stem cells harvested from extracted wisdom teeth. Furthermore, we propose the fabrication of composite jaw bone+tooth constructs, to coordinately create bioengineered tissues that are physically integrated in a manner that more faithfully reflects the features and functions of natural jaw bone and teeth.
4:15 PM - BM04.02.08
Biomimetic Tooth Repair—Amelogenin-Derived Peptides Guide Remineralization of Human Enamel and Dentin
Deniz Yucesoy 1 , Hanson Fong 1 , Sanaz Saadat 2 , Sami Dogan 3 , Mehmet Sarikaya 1 4 2 Show Abstract
1 GEMSEC & Materials Science and Engineering, University of Washington, Seattle, Washington, United States, 2 Department of Oral Health Sciences, University of Washington, Seattle, Washington, United States, 3 Restorative Dentistry, University of Washington, Seattle, Washington, United States, 4 Chemical Engineering, University of Washington, Seattle, Washington, United States
Caries is a global health issue that affects a large percentage of the population despite the widespread use of fluoride and other preventive treatments. Early stage carious lesions are associated with a variety of clinical conditions, including white spot lesions, incipient caries, and hypersensitivity. If left untreated, caries can lead to tooth loss or require complex restorative procedures. The aim of this study is to develop a peptide-based biomimetic remineralization treatment to restore the demineralized dental hard tissues. For this purpose, we have identified a set of remineralization-directing peptides derived from amelogenin (the major protein in enamel development). This is achieved through a design algorithm which includes biocombinatorial selection using peptide libraries, similarity analysis using bioinformatics tools, molecular dynamics simulations, and iterative binding and mineralization assays. The effect of Amelogenin Derived Peptides (ADPs) in controlling remineralization of dental hard tissues was tested in vitro on artificially demineralized enamel, dentin and cementum tissues of extracted human molars. The remineralization was performed under simulated physiological conditions by delivering the peptide and ionic calcium and phosphate, either in aqueous solutions or in a gel formulation. Following the treatment, the structure and morphology of the remineralized tissues were characterized in detail using scanning electron microscopy and energy dispersive X-ray spectroscopy. Local mechanical properties, including elastic modulus and hardness, were determined using nanoindentation tests. In both aqueous solution-based and gel formulation-based treatment modalities, the peptide treated test groups resulted in the formation of continuous hydroxyapatite mineral layer on both enamel and dentin, at a rate of approximately 5-10 µm/hr. No detectable mineral layer was observed in the absence of peptide. More significantly, cross-sectional SEM imaging revealed that the mineralized layer formed a transition region by completely integrating with the underlying dentin, similar to dentin-enamel junction (DEJ). The resulting mineral layers had mechanical properties that are comparable to that of dentin. In conclusion, the ADP-guided remineralization is a new approach in reversing demineralization of enamel through biomimetic remineralization. The treatment developed has the potential for clinical implementation in a variety of formulations as well as for developing over-the-counter products in novel dental health care. The work was supported by WA-State Life Sciences Discovery Funds, UW-School of Dentistry Spencer Funds, and Amazon-UW/CoMotion Catalyst Program.
4:30 PM - BM04.02.09
Spatiotemporal Control of Morphogen Delivery to Induce Multi-Differentiation of Stem Cells Using Fluidic Channels in Hydrogels
Brian O'Grady 1 , Daniel Balikov 1 , Ethan Lippmann 1 , Leon Bellan 1 Show Abstract
1 , Vanderbilt, Nashville, Tennessee, United States
Stem cell differentiation is a complex and tightly regulated process that relies on multiple parameters, including intercellular interaction, extracellular matrix stiffness, and gradients of soluble morphogenic cues. Established protocols to direct differentiation are generally geared towards 2D culture, and those that do make use of 3D scaffolds allow control over either the length of time cells are exposed to morphogens (temporal control) or the space within which morphogens are delivered (spatial control), but not both. To fill this gap, we have developed a platform consisting of undifferentiated human mesenchymal stem cells (hMSCs) embedded in a large-scale hydrogel scaffold (on order of centimeters) containing two parallel fluidic channels for localized delivery of soluble compounds. Using a custom-built pump and perfusion system, we are able to direct hMSCs to differentiate into multiple cell types within a single hydrogel using appropriate morphogen delivery via the channel system. After 45 days of perfusion with osteogenic media in one channel and chondrogenic media in the other, the resulting cell-laden hydrogels were characterized using immunofluorescence and western blot analysis for genetic expression of osteogenic and chondrogenic markers, as well as extracellular matrix production from osteocytes and chondrocytes. Immunofluorescence showed the presence of osteocalcin, a bone protein, in the hydrogel regions near the osteogenic media channel, and increased collagen IV production in the hydrogel regions near the chondrogenic media channel. Additionally, cells in hydrogel regions receiving more osteogenic media showed increased expression of Runx2, an osteogenesis-specific transcription factor, and cells receiving more chondrogenic media showed increased expression of Sox9, a critical regulator of chondrocyte differentiation. Our collective data show a spatially defined gradient of controlled differentiation of hMSCs into osteocytes and chondrocytes in the space between the two channels. This cartilage- and bone-containing hydrogel may be useful for in vitro screening of drug candidates, meniscal injury repair, and other regenerative medicine applications. Additionally, the platform we have developed serves as a new tool for stem cell researchers studying fundamental stem cell biological mechanisms, as it offers a new level of control over the presentation of critical differentiation cues in a 3D culture system. By varying the morphogen type, concentration, and flow rate, as well as the channel geometry in the gel, a wide variety of cellular phenotypes and tissue organization may be achieved.
BM04.03: Poster Session I: Biomaterials for Regenerative Medicine
Monday PM, November 27, 2017
Hynes, Level 1, Hall B
8:00 PM - BM04.03.01
Material Flow Cytometry for Rapid Screening of Biomaterial-Specific Stem Cell Behavior
Kirsten Parratt 1 , Jenny Jeong 2 , Peng Qiu 3 , Krishnendu Roy 3 Show Abstract
1 School of Materials Science and Engineering, Georgia Institute of Technology, Atlanta, Georgia, United States, 2 School of Electrical and Computer Engineering, Georgia Institute of Technology, Atlanta, Georgia, United States, 3 Department of Biomedical Engineering, Georgia Institute of Technology, Atlanta, Georgia, United States
High replicate number is critical for statistical power and quantifying the variability of biological systems, but experimentally difficult to achieve. Flow cytometry is a powerful tool to collect high replicate, high throughput data, however, it has not been applied to biomaterial-encapsulated cells and has untapped potential to generate high powered data on biomaterials systems. Our group has developed 3D Material Cytometry (3DMaC), which uses flow cytometry to analyze cell-containing microhydrogel barcodes based on shape and size, and an analysis platform in IDEAS software to sort microgels with high accuracies (82-99%). Based on the multiplexing potential of these barcodes, we hypothesize that our system could screen >600 different materials while maintaining high replicate number, increasing space efficiency over microspotting methods and allowing greater experimental flexibility.
Here, two 3DMaC studies evaluate material properties related to the long-term suitability of implanted biomaterials. (1) How do material properties, specifically the inclusion of osteogenic peptides, influence differentiation and (2) how do material properties mitigate cellular stresses, both as a function of cell concentration. Flow cytometry is unique in its ability to measure individual cells and calculate population statistics. As cell concentration is important in biomaterials studies, 3DMaC can control for this variable to isolate the effect of material properties.
Microgels consisted of poly(ethylene glycol) diacrylate (MW3400) and human mesenchymal stem cells (hMSCs) which are currently studied as long-term implanted cells and as short-term cytokine “factories”. The fabrication process (adapted from the Khademhosseini group) gives controlled cell concentrations and high viability (~78%). The first question was investigated with microgels containing peptides previously shown to direct osteogenesis, osteogenic growth peptide (OGP) and RGD. Osteogenesis is measured by (1) fluorescently-tagged antibodies specific for osteogenic markers (alkaline phosphatase) and hMSC markers (CD90, 105), or (2) Alizarin Red staining for mineralized extracellular matrix. For the second goal, reactive oxygen species (ROS) activity was measured using dihydrorhodamine 123 dye. Two different material combinations (30% PEGDA +/- 0.66 µM RGD) were used to encapsulate cells which were stained immediately after crosslinking. 3DMaC showed that the crosslinking process produces statistically indistinguishable levels of stress in these biomaterials. Future studies will incorporate more material combinations and test cellular stresses from inflammatory microenvironments.
Acknowledgements: NSF Biomanufacturing IGERT, NDSEG Fellowship Program, NSF CCF (# 1552784), and NIH (# CA163481).
 A. Dolatshahi-Pirouz et al, Scientific Reports, 2014, 4, 3896.
 S. L. Vega et al, Annals of Biomedical Engineering, 2016, 44, 1921-1930.
 J. Yeh et al, Biomaterials, 2006, 27, 5391-5398.
8:00 PM - BM04.03.03
ZrCuFeAlAg Metallic Glass with Potential for Biomedical Applications
Lin Liu 1 Show Abstract
1 , Huazhong University of Science and Technology, Wuhan China
The mechanical properties and biocompatibility of Zr60.14Cu22.31Fe4.85Al9.7Ag3 metallic glass (MG) were investigated in detail to evaluate its potential as a bio-material. The Zr-based MG was found to have a low Young's moduli of 82± 1.9 GPa, a high strength of 1720± 28 MPa, and a high fracture toughness of 94±19 MPa√m as well as good fatigue strength over 400 MPa. The corrosion behavior of the alloy was investigated in simulate body fluid (SBF) by electrochemical measurements. It was found that the corrosion resistance of the Zr-based MG is even better than that of pure Zr and Ti6Al4V，XPS analysis revealed that the passive film enriched in aluminum and zirconium oxide can account for the good corrosion resistance of the MG. On the other hand, metal-ion release of the MG in SBF were determined with inductively coupled plasma mass spectrometry (ICP-MS) after being immersed in SBF at 37°C for 30 days, showing a metal-ion release in PPb ( ng/ml ) level. The in vitro test via cell culture indicates that the MG exhibits a cytotoxicity of Grade 0-1, which is as good as Ti6Al4V alloy. Cell adhesion morphological analysis shows the cell flattened and spread well on the surfaces of the MG，exhibiting good biocompatibility. The combination of good mechanical properties and biocompatibility demonstrates that the Zr-based MG can be a good candidate as a new type of load bearing biomedical material.
8:00 PM - BM04.03.04
Design, Manufacture and In Vivo Testing of a Tissue Scaffold for Permanent Female Sterilization by Tubal Occlusion
Prajan Divakar 1 , Isabella Caruso 1 , Karen Moodie 1 , P. Jack Hoopes 1 , Ulrike Wegst 1 Show Abstract
1 , Dartmouth College, Hanover, New Hampshire, United States
Current FDA-approved permanent female sterilization procedures occlude the fallopian tubes by means of invasive surgical intervention or the use of non-biodegradable implants that pose risks and complications, such as device migration, fracture, and tubal perforation. As a safer alternative, we have developed a new tissue scaffold-based device for placement in a non-surgical, two-step procedure. The fallopian tubes are first mechanically de-epithelialized and a tissue scaffold is subsequently placed into each tube. Advantages of this method include the use of a fully bioresorbable polymer and the translational potential, also globally, because of lower costs, lower risks, and procedural ease as no general anesthesia is required. The new tissue scaffold-based devices were made by freeze-casting, a process that allows for the custom-design of structural, mechanical, and chemical cues through material composition, processing parameters, and functionalization. The performance of the device and procedure was tested and optimized in an iterative process through in vivo studies using a rat uterine horn model. The scaffold response and tissue-biomaterial interactions were characterized on histological sections after 6, 12, 18 and 30 days in situ. Overall, the study resulted in the successful fabrication of resilient (easy-to-handle) devices with an anisotropic scaffold architecture that encourages rapid biointegration through a remarkable degree of angiogenesis, directed cell infiltration, and native collagen deposition. Successful tubal occlusion was achieved in vivo in 30 days, revealing the great promise of the new tissue scaffold-based device for non-surgical permanent female sterilization.
8:00 PM - BM04.03.05
Rheological Characterisation of Agarose and Poloxamer 407 (P407) Based Hydrogels
Nehir Kandemir 1 , Yuqing Xia 1 , Pengfei Duan 1 2 , Jinju Chen 1 Show Abstract
1 School of Mechanical and Systems Engineering, Newcastle University, Newcastle upon Tyne United Kingdom, 2 School of Chemical Engineering and Advanced Materials, Newcastle University, Newcastle upon Tyne United Kingdom
Poloxamer 407 (P407) is a block copolymer which has thermo-reversible characteristics and a distinctive sol-gel transition temperature. Agarose is a polysaccharide which has biocompatible characteristics and easily tuneable mechanical properties. Both of these are appealing materials in pharmaceutical and drug delivery applications where characterising their mechanical properties is important as they would affect injectability and release of drugs encapsulated in hydrogel carriers at different temperatures. This study adopted frequency sweep and temperature sweep rotational rheological tests to investigate the rheological characteristics of various concentrations of agarose and P407 mixtures. The frequency sweep tests have revealed that the addition of Poloxamer 407 has a significant effect on the elastic modulus and the phase angle of hydrogels at room temperature suggesting that the structure of the different concentration gels varies. Also, gels with a high P407 concentration have lower elastic modulus especially at lower temperatures. The overall transition temperature for all tested gel concentrations is governed by agarose where the phase change starts at 30oC and the gels completely collapse at the melting temperature of the agarose. However for the higher concentration P407 gels, the phase angle is relatively higher at low temperatures indicating a more viscous structure due to P407 being at a liquid state at 4oC. These findings will contribute to the characterisation of temperature dependent hydrogel mixtures.
8:00 PM - BM04.03.06
Wetspun GelMA/PEDOT:PSS Conductive Fibers for Excitable Tissue Regeneration
Andrew Spencer 1 , Ryan Koppes 1 , Nasim Annabi 1 2 3 Show Abstract
1 , Northeastern University, Boston, Massachusetts, United States, 2 , Harvard-MIT Division of Health Sciences and Technology, Cambridge, Massachusetts, United States, 3 , Wyss Institute for Biologically Inspired Engineering, Boston, Massachusetts, United States
Cell-laden constructs that mimic the structural and mechanical properties of native tissues show great promise for musculoskeletal tissue repair following traumatic injury or disease. To maintain cell viability during materials processing while yielding necessary scaffold architecture, rapid fabrication is required. Recent developments in electrospinning, wet-spinning, and three dimensional (3D) bioprinting have enabled improved cell viability, construct scaling, and architecture resolution. Here we report a method to rapidly produce cell-laden conductive hydrogel fibers, which mimic the native structure and conductivity of excitable fibrous tissues, such as muscle and neural tissue. The conductivity of the fibers aids in cellular communication and promotes regeneration of the engineered tissues. To fabricate these fibers, a composite precursor solution composed of 7% (w/v) gelatin methacrylamide (GelMA) from porcine skin and 0.3% of a conductive polymer complex, poly(3,4-ethylenedioxythiophene):polystyrene sulfonate (PEDOT:PSS), was injected into a bath of aqueous calcium chloride at 4°C. Bivalent calcium ions crosslinked the anionic polystyrene sulfonate in PEDOT:PSS and the cold temperature solidified porcine GelMA, rapidly generating solid fibers. To further increase the mechanical properties and stability of the fibers, a secondary crosslinking step with visible light was employed to crosslink the GelMA and create a semi-interpenetrating network. As expected, the impedance of hydrogels containing 0.3% PEDOT:PSS was significantly lower than plain GelMA hydrogels (261 vs. 449 kOhm) at 1 Hz frequency. Towards the development of mature muscle fibers, the viability of C2C12 cells encapsulated in these fibers was >90% up to day 7. This method of conductive fiber production has the potential for a range of tissue engineering applications, including fibrous tissue regeneration, 3D bioprinting and rapid manufacturing of dual-stage crosslinked conductive fibers.
8:00 PM - BM04.03.07
An Experimental and Theoretical Approach to Understand Mineralization and Role of Organic Materials
Weitian Zhao 1 , Jacques Lemaitre 1 , Paul Bowen 1 Show Abstract
1 , Ecole Polytechnique Federale de Lausanne, Lausanne Switzerland
Understanding biomineralization processes carries an important medical significance, while at the same time serving as a guidance for functional biomaterials design. As one of its applications, the in vivo deposited calcium phosphate (hydroxyapatite) layer on implant materials such as bone implant can help to avoid the typical host response, the encapsulation by fibrous tissues, and achieve a direct bonding with the bone. In fact, the prediction of in vivo results for implantation of material surfaces is of utmost interest in the biomaterials research. Therefore, it is highly desired to find a suitable correlation between easy-to-perform in vitro experimental methods and in vivo outcomes. An in vitro method using simulated body fluids (SBFs) to simulate the process of hydroxyapatite (HA) formation was already proposed and widely used in current academic research. However, the current SBF composition lacks critical components of human blood plasma, such as proteins, which are well known to play an important role in the mineralization process as well as material-cell interactions.
In this project, we investigated the effect of organic materials on HA nucleation from both experimental and computational approach. We selected TiO2 rutile as substrate and several amino acids as a model system to link our experimental efforts to atomistic simulations (molecular dynamics and meta-dynamics). The results show that alanine and serine, although exhibiting similar degree of complexation with free calcium ions, showed different inhibitory effects on HA growth on rutile substrate. Serine showed a stronger inhibition effect compared to alanine, which might be explained by different adsorption strength on rutile surface observed from atomistic simulation using meta-dynamics. On the other hand, the use of arginine not only influence the HA growth but also completely changed its typical morphology from globules composed of nano-flakes to uniform layer of nanoparticles covering the rutile surface. This suggest a change in growth kinetic and also preferential adsorption of organics to certain facets of HA crystal. A concentration-dependent effect of amino acids were also found with 50 mM (alanine or serine) exhibiting a much stronger inhibitory effect compared to a concentration of 10 mM.
The role of organic species in the nucleation and growth of calcium phosphate crystals were obvious from our results. More in-depth atomistic simulation capturing the adsorption/desorption event of amino acids onto rutile surface and its interaction with Ca and P ions present in aqueous solution are currently in progress and will be reported at the conference. Our results set up a preliminary platform to try to understand fundamentally the organic modulators on mineral deposition and could be useful in the field of biomineralization with an application towards functional material’s design for biomedical applications.
8:00 PM - BM04.03.08
An Electroresponsive Dressing for On-Demand Drug Delivery
Gita Kiaee 1 , Pooria Mostafalu 2 , Sameer Sonkusale 1 Show Abstract
1 , Tufts University, Medford, Massachusetts, United States, 2 , Harvard Medical School, Boston, Massachusetts, United States
The pH-responsive wound dressing patch with the ability to release the drug in response to wound healing status is proposed. Such a dressing would improve wound healing outcomes in patients with chronic wounds such as diabetic foot ulcers. The wound patch is made using PEGDA/Laponite hydrogel embedded with chitosan nanoparticles (CHP) containing drugs. The wound patch is designed to release the drug in response to pH. In acidic pH (2 and 5.5) the drug release rate is expected to be low however, at basic pH, the release is expected to be high. We tested the wound dressing patch using for the release of FITC (as a drug model) in different pH conditions. While the very low release was seen for acidic pH, complete release within an hour for pH above 7.4 was seen in experimental results. The high degradation rate of the hydrogel in basic pH is the main reason for the complete release of drug at high pH. pH change was induced using electric fields providing a pH-mediated electrical triggering of drug release. Applying DC bias between the patch and reference electrodes results in cathodic or anodic behavior at the wound dressing patch altering local pH. The biocompatibility of the proposed system was assessed on fibroblast cells and showed an excellent response. Exposing cells to electrical field (2.5 V and 4V) for 20 minutes did not show to have any toxic effect on cells as the wound closure speed remain approximately constant in different groups. The biocompatibility of PEGDA/LAP/CHP wound dressing was also confirmed in proliferation assay for 7 days. In summary, the proposed pH-mediated electroresponsive on-demand drug delivery has great potential to speed up wound healing in patients with chronic wounds.
8:00 PM - BM04.03.09
Novel Photocuring-Assisted 3D Plotting Using Photocurable Ceramic Slurry for Complex-Shaped Ceramic Architectures with High Shape Retention
Wooyoul Maeng 1 Show Abstract
1 , Korea University, Seoul, SE, Korea (the Republic of)
We herein propose a novel type of additive manufacturing (AM) techniques, denoted as “photocuring-assisted 3D plotting”, which can rapidly solidify extruded green filaments comprised of ceramic powders and photocurable monomers in situ using UV light. In particular, the use of a mixture of 90 wt% diruethane dimethacrylate (UDMA) and 10 wt% triethylene glycol dimethacrylate (TEGDMA) monomers enabled the favorable extrusion of the ceramic slurry through a fine nozzle with high green strength after photocuring. This innovative approach allowed for the construction of ceramic architectures with high shape retention. As an example, a free-standing helical structure with a circular cross-section was successfully produced even without the use of any supporting materials by continuously plotting an extruded filament in accordance with a predetermined 3D model. In addition, the unique in situ photocuring process during extrusion allowed the green filaments to be strongly bonded together, while maintaining their circular cross-section, and thus a porous ceramic scaffold with a tightly controlled porous structure could be produced.
8:00 PM - BM04.03.10
Interaction of Stem Cells with Metal Oxide Thin Films
Wilfrid Prellier 1 Show Abstract
1 , CRISMAT Laboratory, Caen France
tem cells are extensively studied due to their unique properties and great potential in biomedical applications . Thanks to a variety of biological responses, metal-oxide coatings are widely used for interacting with stem cells [2, 3]. A very limited number of oxides have however been studied, and mechanism of the cell-material interaction is still not completely clear.
Here, oxides thin films, that are well-known for their electronic properties, are deposited on substrates using the pulsed laser deposition (PLD) technique. Bone marrow mesenchymal stem cells were cultured on the surfaces of thin films and their adhesion, proliferation and differentiation were evaluated as a function of the growth conditions, thickness of the films, surface roughness and morphology. These results will be of a great clinical significance, in particular, in the field of regenerative medicine.
M.K. thanks support from Normandie Université for her PhD fellowship. Partial support from project Emergence Normandie, and the FEDER are also acknowledged.
8:00 PM - BM04.03.11
Tensile Strain-Controlled Drug Release from Micro-Cracked Membrane
Minseo Kim 1 , Hyungkook Jeon 1 , Seong Kyung Hong 1 , Seong Jin Cho 2 , Geunbae Lim 1 Show Abstract
1 , POSTECH, Pohang Korea (the Republic of), 2 , Chungnam National University, Daejeon Korea (the Republic of)
The mechanical stimuli-responsive systems such as self-healing polymers, strain sensors, and mechanically induced drug delivery systems have received high interest from many researchers. Especially, mechanically induced drug delivery system has recently been placed at the center of attention. This is because the mechanical stimulus has close relation with the human body, such as compression in cartilage, tension in bone, tendon and muscle, and external force applied on skin. Mechanical stimulus is ubiquitously achieved in the human body and can be externally applied with ease. Despite these advantages, only a few researches of mechanical controlled drug delivery system have been carried out.
Here, we have fabricated micro-cracked membranes to control drug release depending on tensile strain. Our micro-cracked membranes were fabricated through a very simple and low-cost process. The micro-cracks were formed on the polyurethane(PU) membrane using the difference of modulus between the non-elastic Titanium(Ti) metal layer and the elastic PU layer. When the external tensile stress was applied to the membrane, Ti metal layer formed micro-crack structures and metal cracks were opened up and widened depending on the tensile strain. The drugs were diffused through the Ti metal cracks and the amount of released drugs was proportional to the magnitude of the tensile strain. As the metal cracks grow wide, the surface of the drug loaded layer is more exposed to the buffer solution.
Our micro-cracked membrane was able to precisely control drug release by controlling the tensile strain, for the exposed surface area of the drug loaded layer was linear to the degree of tensile strain. This tensile strain-controlled release system can be integrated into other flexible devices due to its membrane shape and can be applied to implantable medical devices such as stents. We believe that this strain-controlled release system will enhance the performances of existing mechanically induced drug delivery systems.
8:00 PM - BM04.03.12
Low-Temperature Synthesis of Beta-Tricalcium Phosphate Materials as Bone Graft Substitutes
Wei Cui 1 , Lei Cao 1 , Xing Zhang 1 2 Show Abstract
1 , Institute of Metal Research, Chinese Academy of Sciences, Shenyang China, 2 , University of Science and Technology of China, Hefei, Anhui, China
Beta-tricalcium phosphate (β-TCP) has been wildly used for biomedical applications due to good biocompatibility and biodegradability. Traditionally, β-TCP particles have been prepared by solid-state reactions or calcination at high temperature. In this study, β-TCP scaffolds have been synthesized from natural echinoderm calcium carbonate skeletons (mainly magnesian calcite) by hydrothermal reactions at relatively low temperature, which can be used for bone substitution. Our results revealed that Mg2+ ions played an important role in hydrothermal reactions of magnesian calcite to β-TCP. The β-TCP scaffolds by hydrothermal conversion of sea urchin spines retained the original mesostructures and showed mechanical strength comparable to human trabecular bone. New bone grew into the open-cell structures of β-TCP scaffolds, which were mostly filled after implantation in rabbit femoral defects for three months forming tight interface between the newly formed bone and the scaffolds. The thin struts can be easily degraded in vivo, which further facilitate the ingrowth of new tissue. Thus, β-TCP scaffolds, converted from echinoderm skeletons, while retaining the hierarchical mesostructures, are ideal scaffolds for bone regeneration.
Inspired by nature, biomimetic Mg-calcite products were fabricated through crystallization of amorphous (Ca,Mg)CO3 precursors, which can be further hydrothermally converted to nano-size β-TCP at low temperatures. Moreover, we developed a novel method to synthesize nano-size β-TCP at low temperature using metastable amorphous calcium phosphate precursors. A number of bioactive nano-size β-TCP materials with Mg2+ and/or Sr2+ ion substitution have been prepared following this method. Therefore, a variety of β-TCP materials have been synthesized at relatively low temperature, which can be used as biodegradable bone graft substitutes.
8:00 PM - BM04.03.13
3D Printed Bionic Nose with Electronic Olfactory Epithelium
Yasamin Aliashrafi Jodat 1 , Sudeep Joshi 1 , Kiavash Kiaee 1 , Manu Mannoor 1 Show Abstract
1 , Stevens Institute of Technology, Jersey City, New Jersey, United States
Multi-dimensional integration of electromechanical components with biological entities could enable creation of bionic organs with tremendous impact in regenerative medicine, prosthetics, and human-machine interfaces. A considerable challenge in nasal surgeries such as rhinectomy is the reconstruction of the nose anatomy, given the unique geometries of nasal cartilage structures which necessitate higher degrees of precision in implant fabrication. Current methods rely on transposition of autologous mucosal flaps, cartilage grating and coverage using skin flaps. However, this approach does not include retention of odorant perception.
As a continously improving technology, 3D bioprinting possesses tremendous potential to be utilized as a non-invasive and highly precise approach to repair nasal injuries, and the integration of such technology with biosensors can open new doors to restoring olfaction. Present research reports on 3D printed nostrils consisting of graphene bio-sensors that have been bio-transferred onto the nose construct creating a viable Olfactory Neural Prosthetics. Functionalized with a variety of protein-ligand binding forms, including peptide-bacteria, antibody-virus, and peptide-chemicals, the developed bionic nose can be easily “tuned” to “sniff out” a variety of targets, such as odorants, specific disease indicative biomarkers, explosives and toxics which can’t be detected by the native human nose.
As a demonstrative design, we have chosen to detect three targets of interest: an explosive chemical; 2,4,6-trinitrotoluene(TNT), airborne virus; influenza virus, and a respiratory tract pathogen; Staphylococcus aureus bacteria encompassing a varied spectrum of targets. To detect each target, CVD grown graphene films transferred on a polydimethylsiloxane (PDMS) substrate were functionalized with the corresponding peptides or antibodies and were exposed to the ligand in a custom-made chamber with controllable flow rate. Moreover, to construct the biological structure of the nose, a viable density of chondrocyte cells isolated from the articular cartilage of one month old calves were cultured and seeded with an alginate hydrogel matrix to construct the bio-ink for 3D bio-printing. Finally, a viable 3D bio-printed nose construct was successfully achieved, ready to be integrated with the functionalized graphene bio-sensor. Using a similar approach, the developed bionic nose could promote an accurate and non-invasive strategy for early diagnosis of an impending disease condition, such as an asthma attack, via detection of the appropriate bio-markers. In the long run, in vitro culture of printed hybrid architecture would empower the growth of “cyborg organs” exhibiting enhanced functionalities over human biology.
8:00 PM - BM04.03.14
Engineered Small Diameter Blood Vessels for Cardiovascular Applications
Ebrahim Mostafavi 1 , Nasim Annabi 1 2 3 Show Abstract
1 Department of Chemical Engineering, Northeastern University, Boston, Massachusetts, United States, 2 Biomaterials Innovation Center, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts, United States, 3 Harvard-MIT Division of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States
Cardiovascular disease is one of the most leading causes of mortality in the USA. The limited availability of implantable blood vessels with appropriate physical and biological properties has led to the fabrication of prosthetic vascular conduits. During the last decades various approaches and biomaterials have been applied to solve the problems associated with small diameter blood vessels, particularly the biomechanical mismatch with the host vessel. Quantification of the mechanical properties of native blood vessels including stiffness, elongation, and ultimate strength are required for designing biomimetic synthetic blood vessels.
In this study, we engineered highly elastic blood vessel made of gelatin methacryloyl (GelMA) and poly(e-caprolactone) (PCL) by using electrospinning technique combined with a mandrel with a diameter of 2 mm. The engineered tube was then endothelialized in vitro by using human umbilical vein endothelial cells (HUVECs) and perfused by culture media to form a functional blood vessel. The electrospun composite scaffolds were engineered by dissolving various concentrations of GelMA in the range of 5 to 15 %(w/v) and PCL ranging from 0 to 5 %(w/v) in hexafluoro-2-propanol (HFIP) and electrospinning of resulting solutions. The engineered fibrous composites were then photocrosslinked by using UV light at the intensity of 6.9 mW/cm2 for 10 min to form stable GelMA/PCL fibrous scaffolds. The mechanical properties of the engineered composite scaffolds were optimized by using different ratios of GelMA/PCL and compared with small diameter pig blood vessels. Both native blood vessels and GelMA/PCL scaffolds were stretched to failure in the longitudinal direction. We showed that the stiffness and elongation of the GelMA/PCL composites significantly increased as compared to pure GelMA scaffolds. For instance, at 10%(w/v) GelMA, by increasing the concentration of PCL from 0 to 5 %(w/v), the stiffness of the scaffolds increased from 1.2 to 6.5 MPa; and the elongation increased from 10 to 220 %. The porosity, degradability, and swellability of the engineered electrospun tubes could be also tuned by altering the ratio of GelMA/PCL ratio. Based on our results, of the 6 various compositions, 10%GelMA/5%PCL composite provided a well-balance of mechanical property, swelling capability, and degradation rate which are suitable for blood vessel formation. Furthermore, we cultured HUVECS on this composite tubes and showed that cells could spread and proliferate on both the surface and inside the tubes. The engineered GelMA/PCL tube has potential to be used as synthetic blood vessel for cardiovascular tissue engineering applications.
8:00 PM - BM04.03.15
Engineering Sprayable and Antimicrobial Naturally-Derived Hydrogel Adhesives
Ehsan Shirzaei Sani 1 , Roberto Portillo Lara 1 2 , Devyesh Rana 3 , Nasim Anabi 1 4 5 Show Abstract
1 Chemical Engineering, Northeastern University, Boston, Massachusetts, United States, 2 Centro de Biotecnología FEMSA, Tecnológico de Monterrey, Monterrey, NL, Mexico, 3 Biomedical Engineering, Northeastern University, Boston, Massachusetts, United States, 4 Biomaterials Innovation Research Center, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts, United States, 5 Harvard-MIT Division of Health Sciences and Technology, Massachusetts Institute of Technology, Boston, Massachusetts, United States
Chronic wounds are estimated to affect 6.5 million patients in the U.S annually. These types of wounds do not always heal efficiently and are at risk of bacterial infection. Hydrogel-based adhesives have emerged as alternatives for sutureless wound closure, since they can be tuned to mimic the composition and properties of the native tissue. However, they often exhibit poor mechanical properties and adhesion to wet tissues, and they lack antibacterial activity. In this study, we developed a novel elastic and sprayable hydrogel with antimicrobial properties through photopolymerization via visible light. The engineered hydrogel adhesive is comprised of methacryloyl-substituted tropoelastin (MeTro) and gelatin methacryloyl (GelMA) polymers, which are conjugated with an antimicrobial peptide (AMP Tet213). The hydrogel was shown to mimic the mechanical properties and to adhere strongly to the native tissue, to form an antibacterial and regenerative barrier. The tensile modulus of the hydrogel was found to be tunable in the range of 5 – 25 kPa, based on varying MeTro/GelMA ratios and final polymer concentrations. In particular, the hydrogel formulation consisting of 70/30 MeTro/GelMA with a 15% (w/v) final polymer concentration and 0.01% (w/v) AMP exhibited optimal mechanical and antimicrobial properties. The engineered hydrogel was effective at preventing the growth of both Gram+ (MRSA – CFU decreased 2.5 fold) and Gram- (E.coli – CFU decreased 6.5 fold) bacteria in vitro. Our results also showed that the hydrogels could support the growth and proliferation of 3T3 fibroblasts in both two-dimensional and three dimensional cultures in vitro. In addition, subcutaneous implantation in a murine host showed that the hydrogels could be efficiently biodegraded in vivo, without eliciting any significant inflammatory response. Taken together, our results demonstrated that MeTro/GelMA-AMP hydrogels can be used for sutureless wound closure strategies, while also preventing infection and promoting healing of chronic wounds.
8:00 PM - BM04.03.16
Engineering Highly Adhesive Composite Hydrogels for Surgical Applications
Ehsan Shirzaei Sani 1 , Bahram Saleh 1 , Benjamin Houang 1 2 , Nasim Annabi 1 3 4 Show Abstract
1 , Northeastern University, Boston, Massachusetts, United States, 2 , Grenoble Institute of Technology Phelma, Grenoble France, 3 Biomaterials Innovation Research Center, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts, United States, 4 Harvard-MIT Division of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States
Conventional sealants used for wound closure or sealing tissue defects have remarkable drawbacks including insufficient elasticity and mechanical strength, poor adhesion to wet tissues, cytotoxic degradation products, and low performance in biological environments. To overcome these challenges, in this study we aimed to engineer a photocrosslinkable surgical sealant with tunable mechanical and adhesion properties by using a naturally derived photocrosslinkable biopolymer, gelatin methacryloyl (GelMA), and a synthetic polymer, polyethylene glycol diacrylate (PEGDA). To enhance the adhesion properties of the composite hydrogels, we also conjugated dopamine (DOPA) to GelMA prepolymers. The prepolymer solutions containing various concentrations of GelMA and PEGDA were then photopolyemrized by using visible light to form highly elastic and adhesive sealants. Our results exhibited that physical properties and adhesion strength of GelMA-DOPA/PEGDA bioadhesives could be tuned by changing the degree of methacryloyl substitution in GelMA, degree of DOPA conjugation, PEGDA molecular weight, total polymer concentration, the ratio of GelMA-DOPA to PEDGA, and light exposure time. In situ photopolymerization of GelMA-DOPA/PEGDA can facilitate easy delivery to the defect site, and allow for curing of the bioadhesive exactly according to the required geometry of the tissue to be sealed. Following American Society for Testing and Materials (ASTM) standard tests, the engineered bioadhesive hydrogels exhibited adhesive properties (i.e. wound closure strength, shear resistance and burst pressure) superior to commercially available adhesives such as Evicel®, Progel™ and Coseal™. In vitro cell studies showed that engineered hydrogels were cytocompatible and supported cellular growth and proliferation. Overall, the engineered composite hydrogel has the potential to be used as a wet tissue adhesive for sealing different tissues such has blood vessel, lung, skin, heart, and cornea repair due to its tunable physical and adhesive properties, biodegradability, high biocompatibility as well as low cost.
8:00 PM - BM04.03.18
Conformational and Binding Properties of Fibronectin Fragments by Electron Spin Resonance
William Lindemann 1 , Julia Ortony 1 Show Abstract
1 Materials Science and Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States
One of the fundamental goals of modern biomaterials is to produce a physical environment suitable for cell-habitation and regeneration. The best strategy to accomplish this is the introduction of protein binding-epitopes into functional materials. These short peptides imitate the function of larger proteins, and play an essential role in cell survival and regrowth. By every metric, these short peptides are orders-of-magnitude less potent than larger protein fragments. We hypothesized that this difference can, in part, be explained by the conformational differences between short peptides and their parent proteins. As a case study, we systematically varied the length of fibronectin-mimetic (RGD-containing) peptides, measured their ability to bind to integrins found in cell membranes, and quantified their conformational and dynamic behavior using electron spin resonance (ESR) experiments. In this presentation, I describe the relationship between conformational behavior and binding in these peptides.
8:00 PM - BM04.03.19
Thermal Stability and Antioxidant Activity of SOD Conjugated with Nanocrystalline Cerium Oxide
Bradley Skelton 1 , Dmitry Gil 1 , Vladimir Ivanov 2 , Vladimir Reukov 1 , Brendan Ward 1 , Misha Bredikhin 1 Show Abstract
1 , Clemson University , Clemson, South Carolina, United States, 2 , Kurnakov Institute of General and Inorganic Chemistry, Moscow Russian Federation
Congestive heart disease affects around 5.7 million adults in the United States and roughly half of people who develop it die within five years of diagnosis. Congestive heart failure also costs the nation an estimated $30.7 billion each year. The most common cause of heart failure is loss of myocardial function, resulting from a myocardial infarction. In the United States alone, approximately one million people per year suffer from a myocardial infarction. After a patient suffers a myocardial infarction, his/her heart has an imbalance between oxygen supply and demand called ischemia and blood flow must be restored immediately to reduce tissue damage. Unfortunately, restoring blood flow to the ischemic myocardium, named reperfusion, can also induce injury. This phenomenon known as myocardial ischemia reperfusion injury accounts for up to 50% of the final size of the myocardial infarction3. Oxidative stress, in the form of reactive oxygen species (ROS), is thought to play a large role in this damage. In an effort to reduce the impact of this oxidative stress, superoxide dismutase (SOD), which is naturally found in the body, is being used. SOD is an enzyme that catalyzes the transition of superoxide radicals into either oxygen or hydrogen peroxide. The hydrogen peroxide produced is a known inhibitor of the SOD and results in SOD failure when used for increased duration. A possible solution for this is conjugating it with nanocrystalline cerium dioxide (nanoceria). Cerium dioxide is known for its antioxidant properties caused by the mixed valence states of Ce3+ and Ce4+. These oxygen vacancies allow nanoceria to be an effective ROS scavenger. Thermal stability of the conjugates was also studied at 4°C, 37°C, and 50°C for 1 day, 3 days, and 5 days, while samples placed in 90°C were stored for 2 hours being tested every 20 minutes. An SOD assay using Xanthine Oxidase/Xanthine and WST-1 dye was conducted at each time point to measure the amount of free radicals present in the sample. Experiments have shown an increase in free radicals as the incubation period increases in both 37°C and 50°C, whilst the 90°C have shown considerable effect of nanoceria on thermal stability of SOD.
8:00 PM - BM04.03.20
Artificial Axons for Neural Stem and Progenitor Cell Research
Daniela Espinosa-Hoyos 1 , Anna Jagielska 1 , Huifeng Du 1 , Nicholas Fang 1 , Krystyn Van Vliet 1 Show Abstract
1 , Massachusetts Institute of Technology, Cambridge, Massachusetts, United States
Oligodendrocyte progenitor cells (OPCs) are a class of multipotent stem-like cells that, when differentiated properly, engage and enclose neuronal axons with a myelin sheath. Poor remyelination, due to hindered OPC migration, axon engagement, or differentiation, is associated with poor nervous system function in diseases such as multiple sclerosis. Understanding causes and potential treatments of disorders characterized by incomplete myelin regeneration is particularly challenging due to a lack of materials and devices that replicate key aspects of the OPC-neuron interactions. Emerging research including our own suggests that mechanosensitivity of the oligodendrocyte lineage, and physical and mechanical characteristics of axons that extend from neuron cell bodies, may impact key features of myelination. These features of OPC response that appear to depend on axon stiffness and geometry include onset of oligodendrocyte differentiation, thickness and length of the myelin segments, and the structure of nodes of Ranvier. Here we discuss the development of engineered, three dimensional arrays of copolymer fibers that serve as mimetics of neuronal axons, using a combination of materials engineering and high resolution 3D microfabrication, which enable study of OPC engagement and subsequent myelination in vitro. Using conventional microscopy techniques and high-throughput analysis methods, we show cell-material interactions in these artificial axons are maintained. These materials and approaches facilitate fundamental studies of regeneration in the complex environment of the central nervous system.
8:00 PM - BM04.03.21
Nanocomposite Small Diameter Vascular Graft Stimulated by Ultrasound Waves
Leonardo Ricotti 1 , Ilaria Di Cioccio 1 , Alice Salgarella 1 , Andrea Cafarelli 1 , Paola Losi 2 , Maria Barsotti 2 , Ilenia Foffa 2 , Paolo Dario 1 , Arianna Menciassi 1 , Giorgio Soldani 2 Show Abstract
1 , The BioRobotics Institute, Scuola Superiore Sant'Anna, Pontedera Italy, 2 Institute of Clinical Physiology, National Research Council (CNR), Massa Italy
The development of small caliber vascular grafts represents an important clinical challenge. Existing solutions are still limited by a sub-optimal biomaterial response and a rather poor re- endothelialization, once the prosthesis is implanted in vivo.
We developed a novel small diameter (< 6 mm) elastomeric vascular graft made of poly(ether)urethane–polydimethylsiloxane doped with barium titanate nanoparticles (BTNPs, diameter: 300 nm). The graft was fabricated through a spray, phase-inversion technique that led to two different layers: (1) a high porosity internal layer about 100 μm thick and a low porosity external layer about 400 μm thick. The internal layer was provided with BTNPs (0.2% w/v) by adding the nanoparticles, previously functionalized with glycol chitosan, in the solution before spraying it. Energy Dispersive X-ray (EDX) analyses revealed a uniform dispersion of such nanoparticles in the inner graft layer.
BTNPs show a rather high piezoelectric coefficient. Previous evidences from our group showed that piezoelectric nanoparticles, when stimulated with mechanical waves (such as ultrasound) can trigger beneficial effects (in terms of enhanced proliferation and differentiation) on a wide range of cells, by exploiting the direct piezoelectric effect (generation of local electrical charges due to external mechanical inputs). Our hypothesis is that a small diameter vascular graft based on a highly piezoelectric internal layer and periodically stimulated by outer ultrasound waves can significantly enhance the recruitment and differentiation of circulating endothelial progenitor cells (EPCs) and promote a proper endothelialization of the graft.
The nanodoped graft and the non-doped control exhibited similar mechanical properties, with a radial elastic modulus of about 500 kPa and a longitudinal elastic modulus of about 450 kPa. The piezoelectric coefficient of the nanodoped materials was evaluated through piezoelectric atomic force microscopy.
Piezoelectric prostheses and non-piezoelectric controls were tested in vitro. All samples were provided with a uniform fibrin coating, before cell seeding. Then, EPCs extracted from the blood of human volunteers were seeded on the grafts. EPCs successfully adhered to both the fibrin-coated elastomeric material and to the nanodoped counterpart, showing in both cases a good cell viability. Ultrasound stimulation experiments are ongoing: all samples will be exposed periodically (once per day) to a highly controlled ultrasound dose (500 mW/cm2 for 300 s, at 1 MHz frequency with a pulse repetition frequency of 1 kHz and a duty cycle of 20%), for a total of 3 days. Experiments will be performed using a custom ultrasound stimulation set-up that guarantees no reflections/attenuations of the mechanical wave.
8:00 PM - BM04.03.22
PEG Cell Culture Platform with In Situ Tunable Mechanical Properties to Study the (Ir)reversibility of the MSC Fate
Anouk Killaars 1 2 , Cierra Walker 1 2 , Tobin Brown 3 2 Show Abstract
1 Materials Science and Engineering, University of Colorado, Boulder, Boulder, Colorado, United States, 2 , BioFrontiers Institute, Boulder, Colorado, United States, 3 Department of Chemical and Biological Engineering, University of Colorado Boulder, Boulder, Colorado, United States
The extracellular matrix (ECM) is highly dynamic; cells constantly remodel the ECM in time and space, especially during development and disease progression. These changes of ECM mechanics can reversibly or irreversibly change MSC’s fate by signaling Yes-associated protein (YAP), an early marker of mechanotransduction, to the nucleus to regulate osteogenic mRNA expression, possibly by changing epigenetic modifications (histone acetylation (AcH3K9)). Precise temporal control of the cellular substrate mechanics is required to elucidate the molecular signaling pathway involved in translating dynamic substrate stiffness to (ir)reversibility of the MSC fate in in vitro cell culture. Therefore, we synthesized poly(ethylene glycol) (PEG) hydrogels that can be in situ degraded, as well as re-stiffened, via a radical addition-fragmentation chain transfer (AFCT) process. Hydrogels were formed with a symmetric allyl sulfide crosslinker flanked with maleimide functionalities (44 mM) crosslinked to octafunctionalized PEG thiol (40 mM, Mn=20k) via cationic polymerization. For photodegradation, the hydrogels were placed in a solution containing 0.05 wt% of photoinitiator lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) and varying concentrations of oxidized glutathione (12.4–1 mM), a cytocompatible biomolecule containing a free thiol moiety. After 10 min of swelling, the gels were softened via AFCT by irradiation with 365 nm light (10 mW/cm2) for 1 min. The degradation was tracked by measuring the storage and loss moduli over time. The storage modulus of the non-degraded hydrogel was 12 kPa and the degraded hydrogels ranged from 10 to 1 kPa. Furthermore, MSC phenotype was studied on non-degraded (G’ 12kPa) and various degraded gels (G’ 10-1 kPa) and showed a stiffness depended YAP nuclear localization and AcH3K9 increase. Gels degraded with 8 mM glutathione (G’ 2 kPa) showed low YAP activation, low AcH3K9 and ideal phenotypic morphology. Additionally, MSCs were seeded onto non-degraded hydrogels and after various time spans (1-7 days), in situ degraded to a soft substrate under conditions that proved cytocompatible (0.05 wt% LAP, 8 mM glutathione). YAP activation was reversible or irreversible depending on the time span on the stiff substrate. Future experiments will include re-stiffening the degraded hydrogels by swelling in various concentrations of octafunctionalized PEG thiol (Mn=10k) with 0.05 wt% of LAP and irradiating with 365 nm light (10 mW/cm2). The re-stiffening will be tracked by measuring the storage and loss moduli over time and the (ir)reversibility of YAP activation will be studied by in situ re-stiffening of the gel at various time points. The role of epigenetic modifications (AcH3K9) will be studied in both softening and re-stiffening conditions. This hydrogel chemistry recapitulates the dynamic nature of the ECM and provides a novel cell culture platform to study molecular mechanisms involved in (ir)reversibility of the MSC fate.
8:00 PM - BM04.03.23
Enhancing the Biocompatibility of Microfluidics-Assisted Microgel Fabrication with Channel Geometry
Suntae Kim 1 , Jonghyun Oh 2 , Chaenyung Cha 1 Show Abstract
1 , Ulsan National Institute of Science and Technology, Ulsan Korea (the Republic of), 2 , Chonbuk National University, Jeonju Korea (the Republic of)
Microfluidic flow-focusing devices (FFD) are widely used to generate monodisperse droplets and microgels with controllable size, shape and composition for various biomedical applications. However, highly inconsistent and often low viability of cells encapsulated within the microgels prepared via microfluidic FFD has been a major concern, and yet this aspect has not been systematically explored. In this study, we demonstrate that the biocompatibility of microfluidic FFD to fabricate cell-laden microgels can be significantly enhanced by controlling the channel geometry. When a single emulsion (“single”) microfluidic FFD is used to fabricate cell-laden microgels, there is a significant decrease and batch-to-batch variability in the cell viability, regardless of their size and composition. It is determined that during droplet generation, some of the cells are exposed to the oil phase which had a cytotoxic effect. Therefore, a microfluidic device with a sequential (‘double’) flow-focusing channels is employed instead, in which a secondary aqueous phase containing cells enters the primary aqueous phase, so the exposure of cells to the oil phase are minimized by directing them to the center of droplets. This microfluidic channel geometry significantly enhances the biocompatibility of cell-laden microgels, while maintaining the benefits of a typical microfluidic process. This study therefore provides a simple and yet highly effective strategy to improve the biocompatibility of microfluidic fabrication of cell-laden microgels.
Gulden Camci-Unal, University of Massachusetts Lowell
Surya Mallapragada, Iowa State University
Matteo Moretti, IRCCS Insituto Ortopedico Galeazzi
Pamela Yelick, Tufts University
Multifunctional Materials | IOP Publishing
BM04.04: Hydrogel-Based Biomaterials
Tuesday AM, November 28, 2017
Sheraton, 2nd Floor, Independence West
8:00 AM - BM04.04.01
Engineering an Immunomodulating Hydrogel Adhesive for Diabetic Wound Healing
Bahram Saleh 1 , Harkiranpreet Kaur Dhaliwal 1 , Ehsan Shirzaei Sani 1 , Roberto Portillo Lara 1 2 , Mansoor Amiji 1 , Nasim Annabi 1 3 4 Show Abstract
1 , Northeastern University, Boston, Massachusetts, United States, 2 Centro de Biotecnología FEMSA, Tecnológico de Monterrey, Monterrey Mexico, 3 Biomaterials Innovation Research Center, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts, United States, 4 Harvard-MIT Division of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States
Diabetes mellitus is an important health problem that affects millions of people worldwide including 29.1 million people in the United States. Impaired vascularization, over-inflammation, and bacterial infection are critical challenges in healing of diabetic wounds. Despite recent advances, the existing wound dressings cannot overcome these challenges due to inappropriate mechanical properties and degradation rate, inability to control inflammation, and promote effective wound healing. Therefore, there is an unmet clinical need to develop innovative therapeutic strategies to overcome these challenges and improve clinical outcomes.
In this project, we have combined CD44 targeting hyaluronic acid (HA)-based nanoparticles (NPs) which can stably encapsulate and protect nucleic acid constructs such as plasmid DNA, microRNA, and siRNA  with highly adhesive hydrogels based on visible light crosslinked gelatin methacryloyl (GelMA)  to engineer an immunomodulating wound dressing for treatment of diabetic wounds. First, we have developed GelMA hydrogel adhesive with mechanical and degradation properties suitable for skin regeneration (elastic modulus of 100-200 kPa, extensibility more than 40% and degradation rate about 3-4 weeks). Macrophages are important immune cells that can promote pro-inflammatory (M1) conditions by local secretion of cytokines, chemokines, and proteolytic enzymes. Inversely, macrophages can support tissue repair by producing anti-inflammatory (M2) cytokines and growth factors that can regulate wound healing. Consequently, for inflammation control, we incorporated HA-based NPs (average particle size: 115.0 ± 10.2 nm) within the GelMA hydrogel adhesive to deliver immunomodulatory miRNA to the wound healing sites. Over 90% cumulative miRNA release was obtained after 48 hours in vitro. Gene expression analysis of lipopolysaccharides (LPS)-induced J77A4 macrophages showed that the expression of the M1 marker iNOS2 decreased 5-fold, while the expression of the M2 marker Arg1 increased 6-fold in miR-222 treated cells.
Our results demonstrate that the engineered hydrogel can be used as a multifunctional sprayable wound patch with the ability to control inflammation and induce tissue healing. In addition, due to its strong adhesion to skin (adhesive strength around 30 kPa) and rapid closure of the wound upon exposing to light, the engineered adhesives will minimize the need for suturing and prevent bacterial infection.
Acknowledgments: The authors acknowledge the support from the FY17 TIER 1 Interdisciplinary Research Seed Grants from Northeastern University, and the startup fund provided by the Department of Chemical Engineering, College of Engineering at Northeastern University.
References:  S. Ganesh, A. K. Iyer, D. V. Morrissey, M. M. Amiji, Biomaterials, 2013, 34, 3489-3502.  N. Annabi, D. Rana, E. Shirzaei Sani, R. Portillo-Lara, J. L. Gifford, M. M. Fares, S. M. Mithieux, A. S. Weiss, Biomaterials, 2017, 1-15.
8:15 AM - BM04.04.02
Tunable 3D Hydrogel Scaffolds to Assess Fibroblast Contractility During the Wound Healing Response
Andrea Gonzalez Rodriguez 1 2 , Megan Schroeder 1 2 , Kristi Anseth 1 2 Show Abstract
1 Department of Chemical and Biological Engineering, University of Colorado Boulder, Boulder, Colorado, United States, 2 , The Biofrontiers Institute, University of Colorado Boulder, Boulder, Colorado, United States
Wound healing is characterized by three different stages: inflammation, proliferation and maturation. While the first steps consist of the recruitment of fibroblasts, pro-inflammatory cytokines and growth factors; the later stages focus on increased contractility and extracellular matrix (ECM) deposition. Cell contractility development is crucial for a proper wound healing response, preventing fibrosis. An in vitro platform that allows the study of cell contraction and wound healing progression could elucidate a better understanding of the causes that lead to fibrosis. Due to the polarizing nature of two-dimensional (2D) cell culture platforms, a three-dimensional (3D) substrate is necessary to properly gauge cell-matrix interactions and contractility. To address this need, we use poly(ethylene glycol)(PEG) based hydrogels capable of being remodeled by encapsulated cells.
To synthesize these hydrogels, 8-arm 40kDa PEG was functionalized with a norbornene moiety, and subsequently reacted with a di-cysteine containing peptide following thiol-ene, photoclick reaction. To allow cell remodeling a matrix metalloproteinase (MMP)-degradable sequence (KCGPQG↓IWGQCK) was included in the di-cysteine crosslinker. Thiol-ene chemistry was selected due to its rapid formation, biorthogonality, optical properties and the ability to readily introduce bioactive moieties. To permit cell contractility and spreading, we introduced a cysteine containing adhesive peptide, CRGDS. Primary valve fibroblasts were encapsulated in hydrogels with a starting modulus of 1kPa and showed high viability (>90%) after ten days in culture. When presented with exogenously delivered cytokines, such as TGF-β and FGF, the fibroblast response and activation to myofibroblasts was assessed. TGF-β treatment increased fibroblast activation, contractility, and allowed for increased cell ECM remodeling, resulting in macroscopic gel contraction and matrix deposition. In contrast, FGF led to proliferation and inhibited myofibroblast activation and gel contraction. Results will also show how the platform allows for collection of encapsulated cells after targeted degradation, which enables further molecular characterization of gene expression (e.g., aSMA, CTGF, Col1a1). Finally, the tunability of the hydrogel provides a platform for probing fibroblast response on substrates with different moduli within a biologically relevant range (1-30 kPa), representing different states of fibrosis.
8:30 AM - BM04.04.03
Silver on Oxidized Cellulose/Alginate Hydrogel for Antibacterial Effect on Wound Site
Ji Un Shin 1 , Young Ju Son 1 , Wei Mao 1 , Myun Koo Kang 1 , Sol Lee 1 , Hyuk Sang Yoo 1 , Sun Young Lee 1 , Jae Gyoung Gwon 1 Show Abstract
1 , Kangwon National University, Chuncheon, SE, Korea (the Republic of)
Antibacterial effect of wound healing matrix is a basic function of the dressing. Using many kinds of natural polymers such as silk fibroin, alginate, chitosan, PVA, and cellulose, hydrogel and nonwoven nanofiber were developed for would dressing. These devices mimic extracellular matrix structure, thereby support wound site regeneration by fibrous pattern. Also, nanoporous electrospun nanofiber mesh physically interfere bacteria invasion to wound site. Chitosan, ciprofloxacin, silver nanoparticle, and honey are well known materials for antibacterial activity that used for scaffold material and incorporated in the scaffold. In this study, we prepared silver-adsorbed cellulose and subsequently embedded in alginate hydrogels to fabricate wound dressing materials with sustained release of silver ion for prolonged antibacterial property.
Oxidized cellulose (OCNF) is prepared to expose aldehyde group on the surface of cellulose nanofibrils. Sodium periodate was incubated with cellulose nanofibrils for 1 and 4h at 45°C. The ring opening reaction of pentose of cellulose by sodium periodate produce two aldehyde groups. And bovine serum albumin (BSA) (0.1, 1, and 5 mg/ml) was added to aldehyde exposed cellulose fibril suspension in 0.1M acetate buffer being BSA coated cellulose nanofibril (BSA-OCNF). After that, silver nitrate dissolved distilled water added to BSA-OCNF suspension, forms Ag-BSA-OCNF by electrostatic interaction, and the Ag+ on cellulose fibril is reduced to Ag by formaldehyde treatment. Released out Ag from BSA-OCNF subsequently absorb on BSA-OCNF being Ag@OCNF. Aldehyde, BSA, and silver ion conjugation efficiency are quantified with elemental analysis, BCA assay and inductively coupled plasma atomic emission spectroscopy (ICP-OES), respectively. Finally, a hydrogel was formed by mixing Ag@OCNF and a sodium alginic acid (SA) solution with various concentration, and crosslinked with calcium chloride (10%, w/v). The concentration of Ag@OCNF is optimized to uniform disperse of the particle in the hydrogel. The hydrogel is expected to show an antibacterial effect by silver nanoparticle and moist condition on wound site that accelerate wound healing speed without inflammation.
8:45 AM - BM04.04.04
Hybrid Hydrogels for Medical Applications
Luisa De Cola 1 Show Abstract
1 , University of Strasbourg, Strasbourg Cedex France
An important goal of tissue engineering is to replace extracellular matrix (ECM) with artificial scaffolds that can be synthesized in mild and biocompatible conditions, introduced to the specific site of interest and used to feed the cells for their growth and proliferation. Despite the substantial progress that has been made in biomaterials synthesis and functionalization, the challenge of mimicking the ECM with implants that are able to reduce immunoresponse is still unmet.
Recent findings have shown that mesenchymal stem cells (MSC) infiltrating into the implant have effects on the scaffold integration by improving the healing process.
Towards this aim, herein we report a novel biocompatible hydrogel with the ability to release an MSC migration-inducing factor, the chemo-attractant Stromal cell-Derived Factor-1α (SDF-1α), for the recruitment of stem cells.
In particular, we synthetized polyamidoamines-based nanocomposite hydrogels, cross-linked with mesoporous silica nanoparticles that can function as nanocontainers for the release of SDF-1α from their pores, to address the cells. The hydrogel proved to provide optimal structural support for MSC infiltration and proliferation and chemotaxis of MSC in vitro was investigated with a dual hydrogel system. Subcutaneous implantation of SDF-1α-releasing hydrogel in mice resulted in a modulation of the inflammatory and fibrotic reaction in vivo, suggesting an improvement of the tissue response towards the implant.
Encouraged by these results the hybrid hydrogels have also been injected in pigs with the final aim to study the gelation kinetic in vivo to perform endoscopic submucose dissection.
9:00 AM - *BM04.04.05
Nano- and Microfabricated Hydrogels for Regenerative Engineering
Ali Khademhosseini 1 Show Abstract
1 , Harvard University, Cambridge, Massachusetts, United States
Engineered materials that integrate advances in polymer chemistry, nanotechnology, and biological sciences have the potential to create powerful medical therapies. Our group aims to engineer tissue regenerative therapies using water-containing polymer networks, called hydrogels, that can regulate cell behavior. Specifically, we have developed photocrosslinkable hybrid hydrogels that combine natural biomolecules with nanoparticles to regulate the chemical, biological, mechanical and electrical properties of gels. These functional scaffolds induce the differentiation of stem cells to desired cell types and direct the formation of vascularized heart or bone tissues. Since tissue function is highly dependent on architecture, we have also used microfabrication methods, such as microfluidics, photolithography, bioprinting, and molding, to regulate the architecture of these materials. We have employed these strategies to generate miniaturized tissues. To create tissue complexity, we have also developed directed assembly techniques to compile small tissue modules into larger constructs. It is anticipated that such approaches will lead to the development of next-generation regenerative therapeutics and biomedical devices.
10:00 AM - *BM04.04.06
Engineering Hydrogels for Musculoskeletal Tissue Repair
Jason Burdick 1 Show Abstract
1 , University of Pennsylvania, Philadelphia, Pennsylvania, United States
Hydrogels represent a class of biomaterials that have great promise for the repair of tissues, particularly due to our ability to engineer their biophysical and biochemical properties. Hydrogels can provide instructive signals through material properties alone (e.g., mechanics, degradation, structure) or through the delivery of therapeutics that can influence tissue morphogenesis and repair. Importantly, hydrogel design should reflect both the clinical context and the natural healing cascades of the damaged tissue. Here, I will give examples of the design of hydrogels based on hyaluronic acid (HA) for the repair of musculoskeletal tissues that have limited natural repair processes.
Towards application in cartilage repair, we have developed hydrogels that introduce numerous biochemical signals to mediate stem cell chondrogenesis. These include binding to receptors (e.g., CD44) through the use of HA backbones or the introduction of peptides (e.g., HAVDI) that mimic n-cadherin interactions found during development. We have utilized engineered screening platforms to probe the influence of these various chemical signals on stem cell fate, as well as developed 3D printing technology to translate the signals into scaffold environments. Towards meniscus repair, multi-polymer fibrous hydrogels that permit control over scaffold porosity and therapeutic release via the engineering of specific fiber populations have been developed. For example, multi-polymer fibers were designed with fibers that selectively release collagenase to provide an environment permissive to cell recruitment, PDGF to actually recruit cells, and poly(ε-caprolactone) (PCL) for stability. When investigated in tissue repair, each fiber population was important to the success of repair tissues. Further engineering of the fibers is used to alter cell interactions and the overall material interface with the biological environment. These studies highlight the importance of hydrogel properties on cartilage repair.
10:30 AM - BM04.04.07
Chitosan/Chondroitin Sulfate Hydrogels Prepared in [Hmim][HSO4] Ionic Liquid
Catia Nunes 1 , Kessily Rufatto 1 , Elizangela de Almeida 1 , Michael da Silva 1 , Alessandro Martins 2 , Fernanda Rosa 1 , Edvani Muniz 1 2 , Adley Rubira 3 Show Abstract
1 , State University of Maringa, Maringa Brazil, 2 PPGCEM, Federal University of Technology - Parana, Londrina Brazil, 3 Dep de Química, Universidade Estadual de Maringá, Maringá Brazil
ILs appear to be a novel and promising possibility for the chemical industries as solvent with abilities to solubilize polysaccharides of high molecular weights1. [Hmim][HSO4] ionic liquid (IL) and bio-renewable sources as chitosan (CHT) and chondroitin sulfate (CS) were used to yield hydrogel-based materials (CHT/CS). The use of IL to solubilize both polysaccharides was considered an innovative way based on “green chemistry” principle, aiming the production of CHT/CS blended systems. CHT/CS hydrogels (at w/w ratios 50/50 and 30/70) were prepared in [Hmim][HSO4] based on method reported by Silva et al.2 The samples were characterized through infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), zeta potential (ZP), scanning electronic microscopy (SEM), X-ray scattering (WAXS) and proton magnetic resonance (1H NMR). Also, the swelling behaviors of hydrogels were assessed at aqueous solutions of different pHs and their cytotoxicity activities against VERO and HT29 cells were evaluated as well. CS release curves were devised in SIF and SGF, using the CHT/CS70 PEC as a carrier matrix. The FTIR results showed an alteration on carbonyl band in 1645-1537 cm-1 range due to interaction of COO- groups on CS with protonated amines on chitosan. Two events related to thermal degradation were observed by TGA. DSC curve showed only an endothermic event peak @ 140 ºC. The CHT/CS 30/70 hydrogel presented ZP of -13.0 mV due to the excess of CS (polyanion), whilst the CHT/CS 50/50 presented ZP of +22.7 mV. SEM images exhibited homogeneous macrostructures by rough surface and when the amount of CS was increased (CHT/CS at 30/70 ratio), smoother surface was formed. All WAX profiles indicated that hydrogels possessing amorphous nature. The signals of H atoms attributed to IL groups do not appear in all CHT/CS 1H NMR spectra of CHT/CS hydrogels indicating the complete IL removal from the PECs after the complex formation process. The cell viability outcomes indicated that hydrogels gleaned from IL were devoid of toxicity. The hydrogels showed excellent biocompatibility towards VERO and HT29 cells (CC50 > 1000 µg mL-1). CHT/CS hydrogels were elicited with high stability and swelling capacities at any studied pH range, being these performances were confirmed mainly by WAXS and SEM techniques, as well as through CS release tests. These findings open perspectives for the developing of new strategies to prepare hydrogels based on biopolymers using green chemistry methodologies. Preparing hydrogels in IL environment is quite new and seems be a opened window for the field.
Work supported by Fundação Araucária, CNPq and CAPES
1 C. Lacroix, E. Sultan, E. Fleury and A. Charlot, Polymer Chemistry 3:538-546 (2012).
2 S. S. Silva, T. C. Santos, M. T. Cerqueira, A. P. Marques, L. L. Reys, T. H. Silva, S. G. Caridade, J. F. Mano and R. L. Reis, Green Chemistry 14:1463-1470 (2012).
10:45 AM - BM04.04.08
Fabrication and Cellular Investigation of Biomimetic Collagen-Gelatin Nanopillar Films
Pinar Alpaslan 1 , Sevde Altuntas 1 , Fatih Buyukserin 1 Show Abstract
1 , TOBB University of Economics and Technology, Ankara Turkey
One of the common problems in implant dentistry is the lack of sufficient amount of bone tissue formation around the implant, which is called restricted osteointegration. So,bone and teeth interaction should be examined and used to modify implant material. Investigation of this interaction brings out the periodontal ligament (PDL) allows the cementum formation and also provides bone regeneration. When regional deformation occurs, PDL manipulates its cells to repair damages on bone and cementum. In addition, it has the cells that are responsible for the secretion of various collagen types.Some parts of PDL include Sharpey fibers which act as shock absorbers between bone and cementum, and consist of structures linking teeth and bone. These fibers have extremely rich and regular structure of collagen. Gelatin is acquired through a controlled denature of the fibrous insoluble protein collection. Many studies also focus on gelatin matrix - cell interaction because of its higher stability in aqueous solution.Our aim in this study is to examine bone cell behaviors on arrays of collagen-gelatin mixed nanorods which are bioinspired from Sharpey fiber structures and compare them to flat collagen-gelatin films. Such nanorod arrays were fabricated using anodic aluminum oxide (AAO) molds. These nanoporous substrates are ideal molds for many applications owing to their modifiable chemistry, ultrahigh surface areas, and controllable porosity and pore dimensions that extend perpendicular to membrane surface. In this study, we fabricated nanoporous AAO molds by the two-step anodization of ultrapure Al foils and decreased their surface energy by using silane chemistry. Different concentrations of collagen-gelatin solution were used to obtain collagen-gelatin films that replicate the topography of the AAO molds. We have also optimized the PEGDGE concentration in the collagen-gelatin solution, which is a biological cross-linker used to obtain stable collagen-gelatin nanostructures. The morphological characterization of these nanorod arrays were examined by using scanning electron microscopy (SEM) and atomic force microscopy (AFM). Their stability tests in water and cell medium were conducted by utilizing SEM imaging and swelling tests. Also, cell viability and toxicity tests on SAOS-2 cells were completed. Improved cell adhesion and viability were observed on the nanorod collagen-gelatin films compared to flat collagen-gelatin films and they were not toxic to the cells. On the other hand, we have studied on chitosan-gelatin nanopillar films simultaneously. Finally, as an ongoing part of this study, mineralization and ALP activity of SAOS-2 cells on nanostructured surfaces will be completed under aseptic cell culture conditions and compared to flat collagen-gelatin and chitosan-gelatin films. Moreover, results of collagen-gelatin films will be compared with chitosan-gelatin films.
The project is partially supported by Turkish Academy of Science (TUBA).
11:00 AM - BM04.04.09
Mechanical Properties of life TMG Cross Linked Gels and Their Effect on Embedded Cells as an Approach for Cell Delivery in Regenerative Medicine
Juyi Li 1 , Clement Marmorat 1 , Marcia Simon 1 , Adriana Pinkas-Sarafovai 1 , Miriam Rafailovich 1 Show Abstract
1 , Stony Brook University, Stony Brook, New York, United States
Embedding cells in a hydrogel matrix can provide a controlled environment to influence their growth and migration characteristics. Gelatin based hydrogels crosslinked by microbial transglutaminase (mTG) provide a proteinous network, both chemically and structurally, similar to the one the cells are exposed to in vivo in the extra cellular matrix. Variation in level of crosslinking will dictates resulting elastic properties and further affect the cellular behavior. In this study we assessed the dynamic rheological properties of hydrogels crosslinked with different concentrations of mTG, and evaluated the corresponding cellular response of Dental Pulp Stem Cells (DPSC). Rheology was first used to quantify cross-linking activity. Gels solvated in deionized water exhibited a stable value for their elastic moduli whereas gels formed in media (αMEM±10%FBS) showed increase in elastic moduli values overtime. DPSC growth, morphology, proliferative capability, and motility in 3D cultures were greatly affected by the gels mechanical strength. qRT-PCR expression pattern analysis of osteogenic differentiation related genes has shown that hard mTG hydrogels support osteogenic-like differentiation of encapsulated cells in the gel. This work shows that in addition to its ability to be used as a cell delivery vehicle, the mTG crosslinked gelatin hydrogels support biomineralization for embedded DPSCs.
This work was supported by the NSF inspire program #1344267.
 Yung, C.W., Wu, L.Q., Tullman, J.A., Payne, G.F., Bentley, W.E. and Barbari, T.A. (2007), Transglutaminase crosslinked gelatin as a tissue engineering scaffold. J. Biomed. Mater. Res., 83A: 1039–1046. doi: 10.1002/jbm.a.31431
11:15 AM - BM04.04.10
3D Encapsulated Stem Cell Proliferation and Differentiation in PEG—GelMA Composite Hydrogels of Varying Mechanical Stiffness
Susan Pomilla 1 , Jason Nichol 1 Show Abstract
1 , Endicott College, Beverly, Massachusetts, United States
In order to better realize the immense potential of stem cells as a clinical therapeutic tool for tissue regeneration and repair, researchers must have a clearer understanding of stem cell behavior in response to typical in vivo stimuli. One major stimulus is mechanical stiffness, as this alone has been shown previously to direct stem cell differentiation and phenotype in multiple in vitro studies. However, a few major shortcomings of many of these studies are the use of 2D surfaces rather than more physiological 3D environments, as well as 3D encapsulation in materials where stiffness and concentration cannot be independently altered. Biomimetic materials should mimic the extracellular matrix (ECM) and the complex architecture of native tissues to be successful in vitro models that give valid cues as to how these cells and tissues would perform in vivo. The most popular biomimetic scaffold material for these purposes are hydrogels and we have chosen to work with 2 well characterized hydrogels gelatin methacrylate (GelMA) and poly(ethylene glycol) dimethacrylate 1000 (PEG). The degree of methacrylation of GelMA can be controlled to vary the crosslinking density of the resultant hydrogel, allowing for variation of the mechanical properties independent of concentration. Furthermore, the authors and others have demonstrated that GelMA is a versatile and suitable hydrogel for culture of many cell types with positive cellular attachment and natural degradation properties, but with limited elastic modulus range. Conversely, PEG1000 alone has good hydration and ultimate elastic modulus properties but poor cell binding and viability, which can be greatly improved by the addition of GelMA. For this study gelMA of varying degrees of methacrylation (low, medium, and high) and PEG were homogenously combined to create a 30% w/v hydrogel (15% w/v gelMA and 15% w/v PEG), while 15% (w/v) gelMA hydrogels of all 3 formulations were also used as controls to study human mesenchymal stem cell (hMSCs) elongation, morphology, proliferation and gene expression using the TruSeq RNA stem cell expression panel over 14 days of static culture. It was hypothesized that the varying elastic moduli of the hydrogels in both the composite and gelMA hydrogels would result in morphological and expression differences. The data showed that hMSCs elongation occurred early (day 1-4) in both the medium and high stiffness hydrogels with continued growth at day 14 but limited elongation of the hMSCs in the low stiffness hydrogels over 14 days. The morphological differences of the hMSCs between the low, medium, and high hydrogels could indicate the beginning of elastic modulus regulated differentiation. More thorough analysis of cell morphology and RNA expression profiles are still ongoing, but appear to validate the use of this model system both to study stem cell differentiation, as well as to provide an open transcriptome reference data set for other studies.
11:30 AM - BM04.04.11
Biocompatible Hydroxylated Boron Nitride Nanosheets/Polyvinyl Alcohol Interpenetrating Hydrogels with Enhanced Mechanical and Thermal Responses
Hongling Li 1 , Lin Jing 1 , Roland Yingjie Tay 1 , Siu Hon Tsang 1 , Edwin Hang Tong Teo 1 , Alfred Tok 1 Show Abstract
1 , Nanyang Technological University, Singapore Singapore
Polyvinyl alcohol (PVA) hydrogels with tissue-like viscoelasticity, excellent biocompatibility and high hydrophilicity have been considered as promising cartilage replacement materials. However, lack of sufficient mechanical properties is a critical barrier to their usage as load-bearing cartilage substitutes. Herein, we report novel hydroxylated boron nitride nanosheets (OH-BNNS)/PVA interpenetrating hydrogels by cyclically freezing/thawing the aqueous mixture of PVA and highly hydrophilic OH-BNNS. Encouragingly, the resulting OH-BNNS/PVA hydrogels exhibit controllable reinforcements in both mechanical and thermal responses by simply varying the OH-BNNS contents. Impressive 45%, 43% and 63% increases in compressive, tensile strengths and Young’s modulus, respectively, can be obtained even with only 0.12 wt% (OH-BNNS:PVA) OH-BNNS addition. Meanwhile, exciting improvements in the thermal diffusivity (15%) and conductivity (5%) can also be successfully achieved. These enhancements are attributed to the synergetic effect of intrinsic superior properties of the as-prepared OH-BNNS and strong hydrogen bonding interactions between the OH-BNNS and PVA chains. In addition, excellent cytocompatibility of the composite hydrogels was verified by cell proliferation and live/dead viability assays. These biocompatible OH-BNNS/PVA hydrogels are promising in addressing the mechanical failure and locally overheating issues as cartilage substitutes and may also have broad utility for biomedical applications such as drug delivery, tissue engineering, biosensors and actuators.
11:45 AM - BM04.04.12
Affinity and in vitro biocompatibility of a PCL based biodegradable polymer to myoblast, bacterial growth and blood cells
Nitu Bhaskar 1 , Bikramjit Basu 1 Show Abstract
1 , Indian Institute of Science, Bangalore, KA, India
Biodegradable polymeric devices have significant potential in various fields of biomedical engineering. Designing a suitable polymer that meet crucial requirement for drug delivery application is one of the main challenges. A significant experimental efforts have been made to prepare a novel crosslinked biodegradable polymer, incorporating salicylic acid (SA) and polycaprolactone (PCL) into a biodegradable backbone, which could be used for drug delivery application with enhanced degradation rate and improved biocompatibility as compared to its synthetic PCL counterpart. Lactic acid was used as a spacer to conjugate PCL and SA. The as prepared polymer was chemically characterized by fourier transform infrared (FT-IR) and nuclear magnetic resonance (1HNMR) spectroscopy. The thermal degradation studies (TGA) of the synthesized polymer showed that it is thermally quite stable thereby extending its use for a wide range of applications. Contact-angle measurement indicated that synthesized SA-PCL exhibited favourable wettability with 53° water-in-air contact angle. In vitro hydrolytic degradation studies for the prepared sample in presence and absence of Pseudomonas lipase in PBS (pH 7.4, 37°C) showed enhanced rate of degradation which was further increased in presence of lipase. Further, optimization of this formulation with in vitro cytocompatibility, hemocompatibility and antibacterial studies were performed. In vitro culture cell study for the time period of 5 days using C2C12 myoblast cell line indicated an enhanced proliferation rate of cells on the surface of the polymer. Minimal hemolysis and platelet activation indicated good hemocompatibility. In addition, the synthesized polymer effectively showed an inhibitory effect on the growth of Escherichia coli and Streptococcus aureus over 24 h of culture. These results suggest the potential of using synthesized SA-PCL polymer for biomedical applications especially in the areas of controlled drug delivery systems.
BM04.05: Polymeric Biomaterials for Regenerative Engineering I
Tuesday PM, November 28, 2017
Sheraton, 2nd Floor, Independence West
1:30 PM - BM04.05.01
Light Responsive Hierarchically Structured Liquid Crystal Polymer Networks for Harnessing Cell Adhesion and Migration
Gulistan Kocer 1 , Jeroen ter Schiphorst 2 , Matthew Hendrikx 2 , Hailu Kassa 3 , Philippe Leclere 3 , Albertus Schenning 2 , Pascal Jonkheijm 1 Show Abstract
1 , University of Twente, Enschede Netherlands, 2 , Technische Universiteit Eindhoven, Eindhoven Netherlands, 3 , University of Mons, Mons Belgium
Extracellular matrix (ECM) is equipped with multiple chemical, mechanical and physical cell-instructive cues which are dynamically (with high spatial and temporal precision) regulated to direct cell and tissue behavior. Recapitulating these dynamic changes using stimuli-responsive materials has been very attractive to generate biomaterials which can closely mimic the natural microenvironment for regenerative medicine applications. It is known that size of the topographical features (from nano to micrometer scale) and their spatial arrangement as one of the physical matrix cues critically influence cell adhesion, migration, differentiation as well as tissue organization. Therefore, it is highly interesting to generate new biomaterials which can offer dynamic control over the presentation of topographical cues to cells to guide them towards the desired fate. In this study, azobenzene based light responsive liquid crystal polymer networks (LCNs) were used for their adaptive and programmable nature to achieve, for the first time, hybrid biointerfaces presenting (light-responsive) topographical cues at micrometer scale and changes in nanoscale roughness at the same time to control cell migration as one of the critical processes in tissue remodeling, wound healing as well as biomaterial colonization by cells upon implantation. Optical profilometry measurements confirmed that upon light stimulation, micrometer scale pillar like topographical features with various heights could be formed on the LCN surfaces. Furthermore, atomic force microscopy (AFM) measurements revealed that illumination causes an increase in surface nanoroughness without changing the stiffness. Live cell imaging experiments using NIH3T3 fibroblasts as a model cell type showed that cell speed and migration patterns were strongly dependent on the height of the (light-responsive) micrometer scale topographies and changes in surface nanoroughness. Moreover, upon in situ temporal changes (i.e. illumination of surfaces in presence of cells) in surface nanoroughness, cell migration patterns could be switched dramatically pointing out the ability to dynamically control cell behavior on these surfaces preserving high cell viability during illumination. Finally, the possibility to form photo-switchable topographies was shown which is interesting for future studies where topographical cues could be presented in a reversible fashion to cells.
1:45 PM - BM04.05.02
Ion Transport Study of Conducting Polymer Nanotubes for Biomedical Applications
Martin Antensteiner 1 , Mohammad Javad Eslamian 1 , Mohammad Reza Abidian 1 Show Abstract
1 , University of Houston, Houston, Texas, United States
Biomedical actuators capable of mimicking the elegant motion of biological muscle are highly desirable for biomedical applications such as microsurgical instruments and microcontainers. Conducting polymers (CP) are promising materials for bioactuators owing to their ability to reversibly change volume through mass transport during the redox process and their excellent biocompatibility. Poly(pyrrole) is among the most studied CPs for this application because of its excellent mechanical stability and high strain tolerance while Poly(3,4-ethylenedioxythiophene) (PEDOT) has superior chemical stability and electrical conductivity. Although most studies have focused on the actuation properties of PPy, less has been done for PEDOT. To the best of our knowledge, no work has compared the actuation behavior of PPy and PEDOT with large dopant ions such as polystyrene sulfonate (PSS). Further, less attention has been given to the impact of ion motion on the surface roughness of CP films, which has been shown to influence ion exchange behavior and cycling stability. In this work, we compared the ion transport behavior of PPy and PEDOT nanotubes doped with PSS. For fabrication of CP nanotubes, first poly (L-lactic acid) (PLLA) template nanofibers (200-500nm) were electrospun onto standard gold quartz-crystals. Then, CPs were galvanostatically deposited with 0.5mA/cm2 current density on the electrodes and around the PLLA fibers. Finally, PLLA fibers were dissolved to create hollow CP nanotubes. The electrical properties of CP nanotubes were studied using impedance spectroscopy and cyclic voltammetry (voltage between -0.8 and 0.4 V with scan rates 0.05, 0.1 and 0.2V/s). Ion mass transports as a function of scan rates will be quantified using an electrochemical quartz crystal microbalance (EQCM). In addition, the degradation (cycling stability) of CP nanotubes will be tested over 1000 cycles. The surface roughness of CP nanotubes will be assessed using materials confocal microscopy. The study will pave the way to investigate the application of conducting polymers for drug delivery, tissue regeneration and bioactuator applications.
2:00 PM - BM04.05.03
Two-Dimensional Scaffold Fabricated with Micro-Patterned Diamond-Like Carbon (DLC) and 2-Methacryloyloxyethyl Phosphorylcholine (MPC) Polymer for the Control of Cell Adhesion
Kenta Bito 1 , Terumitsu Hasebe 2 1 , Shunto Maegawa 1 , Keisuke Miura 1 , Tomohiro Matsumoto 2 1 , Tetsuya Suzuki 1 , Atsushi Hotta 1 Show Abstract
1 Department of Mechanical Engineering, Keio University, Yokohama, Kohoku-ku, Japan, 2 Department of Radiology, Tokai University Hachioji Hospital, Tokai University School of Medicine, Tokyo, Hachioji, Japan
Controlling the cell adhesion is essential for many biomedical and tissue engineering applications. One of the biomaterials for the medical device coating, the 2-methacryloyloxyethyl phosphorylcholine polymer (MPC polymer) is attracting a lot of attention and has been clinically used, because MPC dramatically suppresses the protein adsorption and the following cell adhesion and cell activation. In this study, we proposed a new system to control the cell behavior, which consisted of an MPC polymer coated with a micro-patterned diamond-like carbon (DLC) film. DLC is known as a coating material that improves the biocompatibility of base materials, while DLC also possesses the cell-adhesion property. It was considered that the micro-patterned DLC on the surface of an MPC polymer (DLC/MPC) would work as a scaffold to decide the behavior of endothelial cells. In this work, the solution of 0.5 wt% poly[MPC-co-butyl methacrylate (BMA)] in absolute ethanol was coated on two substrates using a spin coater at a speed of 2000 rpm. Two types of metal mesh grids, whose pore sizes were 40 × 40 µm2 and 60 × 60 µm2, were prepared and put on the substrates. DLC was synthesized on the substrates by the radio-frequency plasma-enhanced chemical vapor deposition (RF-PECVD) in C2H2 precursor gas. MPC polymers were coated with two differently micro-patterned DLC (the densely-patterned and the sparsely-patterned DLC/MPC polymer). The cell growth on the substrates was evaluated using human umbilical vein endothelial cells (HUVEC) incubated on the DLC/MPC polymer substrates. HUVEC were suspended at the density of 2 × 104 cells/mL onto each substrate, which were allowed to grow within a humidified incubator up to 4 days. The growth of the cells on each sample surface was estimated by the fluorescent microscopy for 24 hours of culturing. The microscopic images clearly showed that the adhesion of the HUVEC was only observed on the micro-patterned DLC blocks on each substrate after an hour. Moreover, the HUVEC spread on the DLC blocks after 6 hours of culturing, which could extend over the MPC region after 12 hours of culturing by constructing bridges from one DLC block to another. After 24 hours, the amount of adherent HUVEC and the cell-cell connections were both observed to increase on the densely-patterned and the sparsely-patterned surfaces. Though the morphology of the HUVEC on the sparsely-patterned DLC blocks was found to be affected by the shape of the DLC blocks even when the HUVEC spread, HUVEC were found not to be affected by the densely-patterned DLC blocks. These results demonstrated that the DLC/MPC polymer could have the ability to control the cell proliferation and the cell growth of HUVEC by changing the pattern of the DLC blocks.
2:15 PM - BM04.05.04
Bio-Inspired Design—Fabrication of Silk Fibers and Scaffolds
Yaopeng Zhang 1 Show Abstract
1 State Key Laboratory for Modification of Chemical Fibers and Polymer Materials, College of Materials Science and Engineering, Donghua University, Shanghai China
Animal silks produced by insects and spiders exhibit high strength and toughness and are of great interests for various medical applications. Inspired by the spinning process, spinning apparatus of silkworms and structure of silkworm silk, artificial silks tougher than natural silkworm silk were spun from regenerated silk fibroin (RSF) aqueous solutions in air by using a dry spinning process. Recombinant spider silk protein aqueous solution was also adopted to spin tough artificial spider silk via a bio-inspired microfluidic chip. Moreover, nanofillers such as carbon nanotubes, TiO2 nanoparticles, graphene oxide nanosheets were applied to further reinforce the artificial silks and understand the structural evolution of the hybrid fibers. A green, sustainable ,and promising route was also developed to produce reinforced and ultraviolet resistant silkworm silk by feeding silkworms food containing titanium dioxide nanoparticles. Synchrotron radiation wide-angle X-ray diffraction (WAXD) and small-angle X-ray scattering (SAXS) were adopted to investigate the aggregation mechanism of natural silk fibroin in silkworm gland and RSF in spinning process. Various RSF scaffolds with different structures, improved mechanical properties and bioactive functions were prepared and applied for urethral reconstruction, nerve regeneration and liver function recovery.
2:30 PM - *BM04.05.05
Size-Tailored Gold Nanoparticles—Novel Therapeutic Agents as Electroactuators to Guide Stem Cell Differentiation
Bikramjit Basu 1 Show Abstract
1 , IISc, Bangalore, Bangalore, KA, India
The versatile synthesis of gold nanoparticles (GNPs) with a precise control over their size and surface chemistry offers interesting prospects for their use in biomedical applications. This talk will demonstrate how tailoring the size and surface chemistry of GNPs can be exploited to elicit desired cell response in terms of directing stem cell differentiation and bacteriotoxic effects against pathogenic staphylococcal strains, implicated in prosthetic infection. The first half of the presentation is centered on utilizing citrate-capped GNPs with a median size of 20 nm as intracellular actuators coupled with alternating layer by layer assembly of electroconducting polyaniline and citrate-capped GNPs as extracellular actuators, in response to external electric field (EF) stimulation. Direct current EF stimulation with 100 mV/cm promoted neural-like morphology and the genotypic as well as phenotypic expression of neural markers (nestin, tubulin and neurofilament) in human mesenchymal stem cells (hMSCs). On the other hand, pulsed EF stimulation with 100 mV/cm, 1Hz frequency similar to the heart rate led to the tube-like cell morphology and enhanced mRNA expression of cardiomyogenic markers (GATA-4, ACTN2, TnT2) as well as phenotypic expression of desmin and actinin in hMSCs. Such selective differentiation of hMSCs was triggered by cellular events like increase in reactive oxygen species production, elevated intracellular calcium levels and cell cycle arrest at G0/G1 phase.
3:30 PM - *BM04.05.06
Engineered Tissue Substitutes for the Replacement of Corneal Structures
Stephanie Proulx 1 Show Abstract
1 , Laval University, Quebec City, Quebec, Canada
The cornea is one of the most transplanted tissues, accounting for more than 50 000 grafts each year in North America. As the population ages, the number of patients needing corneas for corneal dysfunction is expected to rise. In addition, the increasing severity of the eye banking exclusion criteria for donor tissue will reduce accessibility to corneal transplant and increase waiting times. Recent progress in tissue engineering has made it possible to consider new solutions to the problems of corneal tissue shortage. Autologous cells are more suitable for permanent tissue replacement since they are not rejected by the patient’s immune system. The approach used in our laboratory is to use the cell’s regenerative capacity for the engineering of an autologous tissue. We have used this approach to engineer all three layers of the cornea: the corneal epithelium, the corneal stroma and the corneal endothelium. Corneal epithelial stem cells can safely be harvested, cultured and transplanted back to the patient in order to reform the corneal surface. Stromal keratocytes can form thick sheets of extracellular matrix in vitro that can be used for stromal replacement. These stromal substitutes are transparent and were found to be biocompatible and functional in preclinical experiments. Tissue-engineered corneal endothelia are also functional in vivo. We have also engineered corneal endothelia using dystrophic endothelial cells. Potential applications for these models are numerous, and include in vitro pathophysiological and pharmacological studies, as well as clinical applications to treat corneal diseases.
4:00 PM - BM04.05.07
A Biodegradable Hybrid Inorganic Nanoscaffold for Advanced Stem Cell Therapy
Letao Yang 1 , Ki-Bum Lee 1 Show Abstract
1 Chemistry, Rutgers, The State University of New Jersey, Piscataway, New Jersey, United States
Identifying a reliable therapeutic method to treat central nervous system (CNS) diseases and injuries [e.g. parkinson disease and spinal cord injury (SCI)] has been a major challenge due to the complex and dynamic cellular microenvironment during disease progression. To this end, stem cell-based tissue engineering has emerged as a highly promising route of treatment. However, crucial barriers such as low survival rate, as well as incomplete differentiation and maturation of transplanted cells remain to be hurdles to achieve the full therapeutic potential of stem cell-based therapies. Therefore, to narrow the gap between current scaffold-based stem cell therapy and neuro-regenerative medicine, the development of an innovative method is dire.
Through mimicking the natural extracellular microenvironment and addressing the challenges raised above, we have developed an innovative biodegradable hybrid inorganic (BHI) nanoscaffold and integrated the development of 3D-novel nanomaterials with precision material simulation into the advanced stem cell therapy for the treatment of CNS injuries (e.g. SCI). Remarkably, our BHI nanoscaffold was self-assembled from atomic-thin 2D transition-metal-oxides and natural ECM proteins through a non-covalent crosslinking mechanism. More specifically, this innovative and biocompatible scaffold has: i) unconventional redox-mediated tunable biodegradation; ii) upregulated ECM-protein binding for creating biomimicry microenvironment; iii) highly efficient drug loading with a novel sustainable drug delivery mechanism; iv) innovative magnetic resonance imaging-based drug release monitoring; and v) DFT simulation-based drug screening, which give us distinctive advantages over conventional scaffolds for stem cell-based tissue engineering. With these unique properties, our bioinspired BHI nanoscaffold significantly improved stem cell survival, selectively guided stem cell differentiation into functional neurons, and promoted the neurite outgrowth of stem cell derived neurons. Most importantly, by incorporating human induced pluripotent stem cell-derived neural stem cells (hiPSC-NSCs), we achieved enhanced functional recovery in vivo in a murine SCI model by combining therapeutic effects of our nanoscaffold and spatiotemporal controlled delivery of computationally-screened neurogenic and anti-inflammatory drugs.
In conclusion, addressing the critical challenges in treating CNS diseases, we developed an innovative BHI nanoscaffold and demonstrated its ability to control stem cell neuronal differentiation and to promote neuronal behaviors in vitro and in vivo. Considering its excellent therapeutic potential for enhanced stem cell transplantation and sustainable drug delivery, our BHI nanoscaffold would provide a unique route to overcome the crucial barriers that limit stem cell-based tissue engineering and regenerative medicine.
4:15 PM - BM04.05.08
The Impact of Poly(4-vinphylpridine) Electrospun Fiber Scaffolds on Dental Pulp Stem Cells Proliferation and Differentiation
Linxi Zhang 1 , Chung-Chueh Chang 2 , Miriam Rafailovich 1 , Marcia Simon 3 Show Abstract
1 Materials Science and Engineering, Stony Brook University, Stony Brook, New York, United States, 2 Advanced Energy Center, Stony Brook University, Stony Brook, New York, United States, 3 School of Dental Medicine, Stony Brook University, STONY BROOK, New York, United States
P4VP has been shown to be a promising polymer substitute for tissue culture plastic (M. Apostol, Polymer Journal 43(8):723-732, June2011). P4VP has similar rheological properties to polystyrene and hence can be processed as transparent thin films, fibers, or molded structures. Previous studies have mostly focused on 2-D structured films which hardly represent the real conditions of scaffolds in vivo environment. Here we fabricated P4VP 3-D fiber scaffolds by electrospinning technique in both nano and micro fiber size to mimic tissue’s natural extracellular matrix. Dental pulp stem cells are cultured on the P4VP fiber scaffolds and thin films and incubated for 28 days to study the impact of substrate morphology to the DPSCs. Our research shows that the cell proliferation is regulated on the fiber substrate. Hydroxyl apatite deposition templated on coiled protein-rich structure on the fibers is revealed by SEM, EDAX and Raman. Real-time PCR analysis and osteocalcin protein (OCN) staining suggest that P4VP fiber scaffolds can promote the differentiation of DPSCs in the absence of chemical cue, such as dexamethasone. In conclusion, DPSCs are sensitive to the substrate morphology change from 2-D thin film to 3-D fiber scaffold. The electrospun P4VP fibrous scaffolds support DPSCs differentiation.
BM04.06: Poster Session II: Biomaterials for Regenerative Medicine
Tuesday PM, November 28, 2017
Hynes, Level 1, Hall B
8:00 PM - BM04.06.01
Hydroxyapatite Nanocrystal Deposited Titanium Dioxide Nanotubes Loaded with Antibiotics for Combining Biocompatibility and Antibacterial Properties
Xuefei Zhang 1 , Yuan Zhang 1 , Matthew Yates 1 Show Abstract
1 , University of Rochester, Rochester, New York, United States
The number of patients that need internal fixation devices or orthopedic implants all over the world has increased rapidly over the past few decades. The success of these implants depends on two factors: the bone-implant integration, and the prevention of bacterial infection. Hydroxyapatite (HA, Ca10(PO4)6(OH)2) is a type of calcium phosphate compounds widely used in orthopedic applications, such as bone repair surgery and load-bearing implants, due to the similarity of synthetic HA to the natural mineral component of calcified tissue. The HA is often applied as a thin coating to enhance bioactivity of load-bearing metal implant surfaces. Unfortunately, the biocompatibility of HA also allows for the proliferation of bacterial cells on the HA surface. Bacterial infection is one of the most serious complications of implant surgeries and can lead to severe physiological damage and, consequently, the need for additional costly surgical procedures. Therefore, an HA-coated implant which can not only enhance the osseointegration but also act as a vehicle for local delivery of antibiotics to prevent infections is desired. In the present work, HA-coated titanium dioxide nanotubes loaded with antibiotics were developed to address the challenge. Titanium dioxide nanotubes were first fabricated on titanium plates using anodization techniques. Then HA nanocrystals were synthesized on the titanium dioxide nanotubes by electrochemical deposition, followed by loading of a model antibiotics. The antibiotics release kinetics from the coating was investigated. The antibacterial activity of the coating was also demonstrated by bacterial tests. The antibiotics-loaded titanium dioxide nanotubes with HA nanocrystals offer a desirable surface feature combining biocompatibility and antibacterial properties.
8:00 PM - BM04.06.02
Fabrication of Photo-Responsible Cell Culture Vessels Using Titanium Oxide Films
Masato Ueda 1 , Chika Fujita 1 , Masahiko Ikeda 1 Show Abstract
1 , Kansai University, Suita Japan
Several techniques have been employed to attach/detach cells to/from a substrate. Cells cultured on a substrate are generally detached from the substrate into a sheet by the destruction of protein between the cells and the substrate using enzymes such as trypsin. However, the enzymes also damage the adhesion molecules among the cells.
Anatase-type TiO2 is an n-type semiconductor with an energy band gap of Eg=3.2 eV, which displays a relatively high photocatalytic activity under light irradiation at wavelengths of λ<380 nm. When an n-type semiconductor is immersed in an aqueous solution, an up-hill potential gradient is produced towards the surface in the conduction and the valence bands. Under ultraviolet (UV) irradiation, electrons and holes are formed in the conduction and the valence band, respectively. These photogenerated charges are then spatially separated by the potential gradient. This phenomenon can be regarded as photocurrent, electromotive force, or redox reaction.
The purpose of this work was to fabricate photo-responsive cell culture vessels using TiO2 and to investigate adhesion/proliferation behaviors of cells on them. Following three types of vessels were fabricated: (a) TiO2 film/Ti, (b) TiO2 film/SiO2 and (c) Ti/TiO2 film/Hanks’ solution/SiO2. In the vessels of (b) and (c), light was irradiated from back-side of the vessels.
TiO2 films were prepared on commercial purity Ti plates or SiO2 plates by the combined chemical-hydrothermal treatment or a general sol-gel method using titanium tetraisopropoxide, respectively. Primary osteoblasts were seeded on the vessels and then incubated at 37 °C. During the incubation, the light irradiation was performed intermittently or continuously.
Basically the number of cells monotonically increased with incubation periods on both Ti and TiO2 under darkness. The intermittent light irradiation promoted the adhesion of cells on the surface in the vessel of (a). The formation of Ti-OH groups on the TiO2 seems to be facilitated by the UV irradiation. In contrast, the cells decreased under continuous light irradiation. Direct UV irradiation is known to damage to the cells. The decrease was significant in the TiO2, not in the Ti. The cells might be additionally received some sort of stimulus from the surface of TiO2. In the vessel of (b), the cells are not exposed to UV since it is completely absorbed by the TiO2 layer. However, the cell adhesion was suppressed by light irradiation; it might be due to generated photocurrent or hydroxyl radicals on the surface of TiO2. On the other hand, the cell adhesion was also suppressed by continuous light irradiation in the vessel of (c). The cells adhered under dark could be detached by light irradiation in the vessels of (b) and (c), though the efficiency was not so high. These results imply that the adhesion/proliferation/detachment behaviors of cells can be controlled by the photocatalytic reaction of TiO2 and the irradiation patterns.
8:00 PM - BM04.06.03
Compressed Collagen Blended with Corneal-Derived Material for Developing Mechanically Improved and Biomimetic Tissue-Engineered Corneal Stroma
Hyeonjun Hong 1 , Hyeon Ji kim 1 , Man-Il Huh 2 , Hong-Kyun Kim 2 , Dong-Woo Cho 1 , Dong Sung Kim 1 Show Abstract
1 Department of Mechanical Engineering, Pohang University of Science and Technology (POSTECH), Pohang Korea (the Republic of), 2 Department of Ophthalmology, Kyungpook National University School of Medicine, Daegu Korea (the Republic of)
In corneal tissue-engineering, collagen compression process has received much attentions to develop tissue engineered corneal stroma, enabling to fabricate mechanically enhanced collagen construct with facile incorporation of corneal cells inside it. Though, the compressed collagen can provide suitable biological environment for the keratocyte attachment and growth, the single component of corneal extracellular matrix (ECM) cannot satisfy natural cornea-like environment for keratocyte functions. In recent studies, corneal-derived decellularized extracellular matrix (co-dECM) has been widely utilized to develop a tissue-engineered corneal stroma. The co-dECM composed of various corneal ECM components including, type I, IV collagen and glycosaminoglycans (GAGs) and thus can provide biomimetic environment for promoting keratocyte functions. Moreover, it was noticed that the co-dECM showed significantly higher optical transmission as compared to the conventional collagen gel. However, the co-dECM gel has lower mechanical properties than conventional type I collagen gel, which restrict the utilization of co-dECM gel in development of tissue-engineered corneal stroma. In this regard, this study developed a mechanically improved and more biomimetic corneal stroma through compression/dehydration of collagen gel blended with the co-dECM. By blending the co-dECM with typical type I collagen, we could obtain the proper conditions of hydrogel for compression/dehydration process which leads to achieve the mechanically improved biomimetic collagen construct. The physical, optical and mechanical characteristics of compressed collagen blended co-dECM sheet was also evaluated with changing co-dECM blending ratio. Finally, the in vitro cell culture revealed that the compressed collagen blended with co-dECm significantly promoted the keratocytes functions.
8:00 PM - BM04.06.04
Mechanical Regulation of Neurite Outgrowth of Hippocampal Cells by Controlling Substrate Stiffness
Aya Tanaka 1 , Yuki Fujii 2 , Nahoko Kasai 1 , Takaharu Okajima 2 , Hiroshi Nakashima 1 Show Abstract
1 , NTT Basic Research Laboratories, Kanagawa Japan, 2 , Hokkaido University, Hokkaido Japan
The mechanosensitivity of neurons in the central nervous system (CNS) is an interesting issue as regards understanding neuronal development and designing compliant materials as neural interfaces between neurons and external devices for treating CNS injury and disease. Although neurite initiation from a cell body is known to be the first step towards forming a functional nervous network during development or regeneration, our knowledge is still limited about how the mechanical properties of the extracellular microenvironment affect neuritogenesis. Here, we investigated the actin cytoskeletal structures of neurons, which are a key factor in neuritogenesis, on substrates with a stiffness-controlled hydrogel to reveal the relationship between substrate stiffness and neuritogenesis.
The cell samples were prepared from Wistar rat hippocampi (embryo day 18), and plated on hydrogel substrates at an initial cell density of 4.0×103 cells/cm2. The hydrogel substrates were prepared on methacrylated coverslips using the photo-initiating polymerization of hydrogel precursor solutions containing acrylamide, N,N’-methylenebisacrylamide (Bis) with a photoinitiator. After the gelation, the hydrogel surface was modified with poly-D-lysine via a heterocrosslinker. The phalloidin labeled actin cytoskeleton of neurons on the hydrogel substrates was observed with confocal microscopy. The elastic modulus of the hydrogel substrates was measured with an atomic force microscope.
We confirmed that the Young’s modulus of the hydrogel substrates was in the 1.5×102 to 3.5×103 Pa range depending on the Bis concentration. We observed that neuritogenesis was significantly suppressed at an elastic modulus of ca. 1.0 kPa, which is stiffer than that of in vivo brain tissue. The confocal microscopy observations showed that the organization of F-actin was closely associated with substrate stiffness. Circumferential F-actin meshworks and arcs were formed at the edge of the cell body as the substrate stiffness exceeded the in vivo brain stiffness. The suppression of F-actin cytoskeleton formation improved neuritogenesis.
The results indicate that the neuronal actin cytoskeletal organization, which depends on the surrounding microenvironment, plays a decisive role in regulating neuritogenesis. It implies that the mechanical properties of the microenvironment can regulate such neuronal behaviors as neurite outgrowth and functional maturation during nervous network formation. We also find that the neurite initiation is suppressed at the elastic modulus, whose stiffness exceeds that of the in vivo brain. Our findings provide insights that help us to understand the mechanosensitivity of neurons and to design the mechanical compliance of materials for a neural interface.
8:00 PM - BM04.06.05
Combing Phosphodiesterase Inhibitor and Connective Tissue Stimulating Drugs in Nanofibrous Scaffold to Augment Bone Fracture Healing
Islam Khalil 1 2 , Isra Ali 2 , Ibrahim El-Shebiny 2 Show Abstract
1 Department of Pharmaceutics and Industrial Pharmacy, Misr University for Science and Technology, 6th of October City,, Giza, Egypt, 2 Nanomaterials Lab, Center of Material Science, Zewail City of Science and Technology, 6th of October City,, Giza, Egypt
According to several attempts to accelerate both wound healing and bone healing, it was observed that both healing process have several common healing features as inflammation, angiogenesis, epithetlization, collagen deposition and remodeling, Topical application of drugs shows promising bone healing activity e.g. phenytoin and sildenafil. However, no available product in the market loaded with any these drugs. Therefore, we propose to fabricate a highly porous electrospun nanofibrous scafold loaded with the two drugs with different healing mechanisms for bone healing to augment bone fracture healing.
The scaffold would be fabricated in the form of a two-layer sandwich structure, where each layer loaded with different drug. The first layer is composed of Phenytoin-loaded PLLA based nanofibers that could deliver the required dose in a well-controlled and sustained manner. The second layer is composed of Sildenafil-loaded PLLA based nanofibers that could deliver the required dose in a well-controlled and sustained manner. The two layers were prepared successfully through electrospinning technique after tailoring different parameters to produce highly porous scaffold in order to facilitate oxygen and nutrient diffusion for cell. Scanning electron microscope revealed that diameter of both PLLA based nanofibers layers were around ranged between 180 ± 20 nm to 290 ± 25 nm. Physicochemical characterization showed that PLLA based nanofibers showed swelling maximum after 6. In addition, biodegedability test showed that the nanofibers were capable of losing more than 25% of their weights in 14 days. Finally in vitro drug release profile showed that the nanofibers could deliver Phenytoin and sildenafil in a well-controlled manner along more than 20 days. Hence, the fabricated drug-loaded implantable scaffold could be a promising solution in complicated bone fracture/defects through targeting different phases in healing process.
8:00 PM - BM04.06.06
FDM Printed Scaffold Topographies and Their Influence of Cell Behavior
Kuan-Che Feng 1 , Adriana Pinkas-Sarafovai 1 , Yuval Shmueli 1 , Chung-Chueh Chang 1 , Marcia Simon 1 , Miriam Rafailovich 1 Show Abstract
1 , Stony Brook University, Stony Brook, New York, United States
Fused deposition modeling (FDM) is a rapidly growing method for device fabrication. The technique is inexpensive and the product, such as bone inserts, dental devices, can be printed directly from CT scans or impressions, and hence specifically tailored for the individual. Furthermore, computer modeling of the thermal distribution within the FDM filament shows that extruded filament temperature is the function of the nozzle temperature, nozzle geometry and extrusion rate. This difference can cause distinctive topography on the surface, and when in contact with a tissue, they could have a profound influence on cell behavior.
In this study we produced FDM printed scaffolds under different settings (nozzle temperature and extrusion rate) and characterized their surface topography by Scanning Electron Microscopy (SEM) and Atomic Force Microscopy (AFM). Changing FDM printing parameters produce surfaces with mixed roughness known as sharkskin, with different periodicity and amplitude.
The impact of topography on cell behavior (mobility, proliferation, differentiation) was measured using dental pulp stem cells. For these cells were cultured on substrates of identical dimensions produced with different printing temperatures and extrusion rates. Biomineralization without dexamethasone was observed in all cultures. qRT-PCR of marker genes (ALP, OCN, DSPP, BSP) demonstrated differences in the cell response to the surfaces
8:00 PM - BM04.06.07
Bactericidal Nanofibrous Polymeric Mats for Wound Dressing Applications
Adnan Memic 1 2 3 , Kasturi Navare 2 , Musab Aldhahri 1 , Ali Tamayol 3 4 , Ali Khademhosseini 3 4 , Sidi Bencherif 2 5 6 Show Abstract
1 Center for Nanotechnology, King Abdulzziz University, Jeddah Saudi Arabia, 2 Department of Chemical Engineering, Northeastern University, Boston, Massachusetts, United States, 3 Biomaterials Innovation Research Center, Brigham and Women's Hospital, Harvard Medical School, Cambridge, Massachusetts, United States, 4 , Harvard-MIT Division of Health Sciences and Technology, MIT, Cambridge, Massachusetts, United States, 5 School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts, United States, 6 Sorbonne University, Biomechanics and Bioengineering (BMBI), University of Technology of Compiègne, Compiègne France
In the local treatment of both chronic and acute wounds it is crucial to prevent infections, control the removal of exudates and create a moist environment to allow for skin healing. To address these challenges it is necessary to develop the next generation of bioactive dressings. These dressings should be able to play an active role in wound protection and healing. Electrospun nanofibrous poly(glycerol
sebacate)-poly(ε-caprolactone) (PGS-PCL) substrates have been extensively used for various medical applications due to their tunable biophysical properties. Electrospun sheets are mechanically flexible and they exhibit a high degree of porosity due to their unique microarchitecture, allowing fluid absorption. In this study we have engineered biodegradable, elastic, and antimicrobial mats generated from a PGS-PCL polymer mixture by electro spinning.
Radicals-generating nanoparticles (NPs) are physically entrapped within the nanofibers by mixing the NPs with the PGS-PCL prepolymer solution prior to electrospinning. The hybrid NPs containing PGS-PCL mats were designed to control the release of bactericidal hydroxyl radicals for several days for the inhibition of various types of robust bacteria, including multi drug resistant Escherichia coli, methicillin resistant Staphylococcus aureus, and Pseudomonas aeruginosa. Our preliminary data suggest that these antimicrobial wound dressing substrates are suitable for absorbing light to moderate drainage while releasing radicals at a controlled level for broad spectrum antimicrobial action via a radical-dependent mechanism, without harming mammalian cells.
Acknowledgement: This project was funded by the National Plan for Science, Technology and Innovation (MAARIFAH) - King Abdulaziz City for Science and Technology (KACST), Kingdom of Saudi Arabia, award number 12-MED3096-3. The authors also acknowledge with thanks Science and Technology Unit, King Abdulaziz University for their technical support.
8:00 PM - BM04.06.08
Substrate Mechanics Regulates the Uptake of Nanoparticles in Stem Cell and Its Differentiation
Ya-Chen Chuang 1 2 , Marcia Simon 3 , Miriam Rafailovich 1 , Chung-Chueh Chang 2 Show Abstract
1 Materials Science and Engineering, Stony Brook University, Stony Brook, New York, United States, 2 ThINC facility, Advanced Energy Center, Stony Brook University, Stony Brook, New York, United States, 3 Oral Biology and Pathology, Stony Brook University, Stony Brook, New York, United States
It has been shown that stem cells isolated from the dental pulp (dental pulp stem cells (DPSCs)) can differentiate and express markers of odontoblasts, osteoblasts, adipocytes or neuronal cells when they are grown in specific inducing media. Adequate studies have shown that diverse scaffolds can provide biochemical and mechanical cues which can regulate or direct stem cell proliferation and differentiation. In our previous study, we have shown that monodisperse polybutadiene (PB) can be used to produce biocompatible flat thin films with different surface mechanics by simply altering the film thicknesses where surface chemistry remains the same. We have also shown that DPSCs can sense and adjust their cell mechanics accordingly to the underlying substrate mechanics after 4 days incubation. In addition, without the addition of inducing media, dexamethasone, biomineralized deposits and up-regulation of osteocalcin (OCN) gene marker were observed on hard PB surfaces. In contrast, extremely low level of biomineralized deposits and OCN were observed on the softer PB surfaces. The rise of nanotechnology also promotes the study on the effects of nanoparticles (NPs) on stem cell and shows that nanoparticles (NPs) can also offer a means of regulating cell function. However, stem cells are extremely sensitive to the extracellular signals where the stimuli from substrate mechanics and NPs should be studied simultaneously. Hence, in this study, we first investigate whether cells sensing the substrate mechanics alter the uptake of NPs and further impact the stem cell function. Briefly, TiO2 NPs (0.1 mg/mL) were added at either day 1 or day 4 (before and after cell sensing substrate mechanics) post-plating onto hard PB substrates. DPSCs were then grown in medium without dexamethasone. At week 1, cell moduli were measured using shear modulation force microscopy (SMFM). At week 4, biomineralization was characterized using SEM/EDS. Immunohistochemical staining was also performed to examine the expression of osteocalcin protein and was imaged by confocal microscopy. The results show that in the case of the addition of NPs at day 1 post-plating, the biomineralization was extremely low and the expression of osteocalcin protein was down-regulated in comparison with NPs-free cell controls. On the other hand, in the case of the addition of NPs at day 4 post-plating, biomineralization and the expression of osteocalcin protein are similar to those without NPs treatment. This result suggests that the fully response of the substrate mechanics plays a role on the effects of NPs on DPSCs differentiation which could be potentially applied to the drug delivery system where regulating the effects of NPs becomes necessary.
[We would like to thank the NSF-INSPIRE program (Grant #1344267).]
8:00 PM - BM04.06.09
Bulk Metallic Glasses for Orthopedic Applications
Ayomiposi Loye 1 , Jan Schroers 1 , Themis Kyriakides 1 Show Abstract
1 , Yale University, New Haven, Connecticut, United States
The regeneration of bone involves the complex orchestration of many biological events. In cases where large bone defects caused by trauma, infection, tumor resection and skeletal abnormalities occur, standard interventions using autologous grafts are utilized. These procedures involve the implantation of bone harvested from another site and are limited by clinical complications, quantity restrictions and high cost. In the creation of the ideal bone scaffold, the interaction of osteogenic cell populations, osteoinductive stimulus, osteoconductive matrix scaffolds and mechanical stability should be studied. The ideal material is osteoinductive (promotes precursor differentiation to osteoblasts), osteoconductive (supports bone growth), and osseointegrative (integrates into surrounding bone). In the clinic, bioactive ceramics are useful due to their high resorption rate, tunable nature, and elemental similarity to bone, but are impractical for load bearing applications because of their high porosity and low strength. Polymers made of collagen and hyaluronic acid have been explored but have limitations due to immunogenicity, sourcing and handling, risk of infection and mechanical weakness. Furthermore, titanium, the most clinically used metal, is a relatively bio-inert metal that requires excessive functionalization to encourage osteoinduction. In the creation of biomaterials for bone implants, there is room for improvement.
To study an alternative metal to titanium, we use Pt57.5Cu14.7Ni5.3P22.5 bulk metallic glass (Pt-BMG) and modify its surface through thermoplastic based nanopatterning techniques to influence differentiation of human mesenchymal stem cells (MSCs) and osteoclas precursor cells. BMGs are a unique class of high strength and elasticity amorphous materials often with high corrosion resistance. Such properties are paired with an unusual plastic-like processing opportunity. Analysis of MSC differentiation indicated that flat and nanopatterned Pt-BMGs promote osteogenic and adipogenic differentiation, respectively, suggesting that nanotopography can overcome the cues given by the high elastic moduli material in improving adipogenic differentiation.
Furthemore, MSCs on flat Pt-BMG showed enhanced osteogenic differentiation in comparison to titanium suggesting that the composition of Pt-BMGs is favorable for osteogenesis. Analysis of pathways involved in mechanotransduction revealed that, in comparison to nanopatterned Pt-BMG and titanium, flat Pt-BMG induced an increase in YAP/TAZ activation and focal adhesion formation. Our studies suggest that flat Pt-BMGs are mechanically strong, bioactive metals that could be used in orthopedic clinical applications. Moreover, the highly tunable nature of BMGs enables the creation of a wide range of surface topographies that can be reproducibly and systematically studied leading to the development of implants capable of modulating stem cell responses.
8:00 PM - BM04.06.10
Hydrolyzed Polyacrylamide Hydrogel as a Scaffolding Material for the Study of the Budding Yeast S. cerevisiae—Potential Biological and Industrial Applications
Marleine Tamer 1 , Maria Bassil 1 , Joseph Stephan 2 , Mario El Tahchi 1 Show Abstract
1 , LBMI, Department of Physics, Lebanese University, Faculty of Sciences II, Jdeidet Lebanon, 2 , Gilbert and Rose-Marie Chagoury School of Medicine, Lebanese American University, Byblos Lebanon
Wild microorganisms such as S. cerevisiae that do grow in laboratory conditions typically undergo adaptive domestication, with a resultant strain that markedly differs from its wildtype counterpart. In addition, typical laboratory growth conditions of yeast, whether in liquid media or on solid agar plates, presents several drawbacks. It is impossible to distinguish different genotypes in liquid culture, for instance, and growing cells on agar doesn’t allow for a rapid and localized change of environmental conditions such as usually occurs in the wild. To circumvent similar problems, several different growing matrices such as agar, agarose, gelatin and polyacrylamide have been evaluated to better recapitulate the natural environment of different types of cells.
In this study we will present the use of hydrolyzed Polyacrylamide (PAAM) as a new scaffolding material for the growth of the budding yeast, S. cerevisiae. Hydrolyzed Polyacrylamide hydrogel is a biocompatible, non-biodegradable material that offers a suitable substrate for cell culture, and holds many advantages over the previously mentioned matrices, including its high fluid content more representative of natural yeast habitats, as well as the possibility to more accurately manipulate experimental variables, both temporally and spatially.
In the first part of this study, we will evaluate the effect of hydrogel matrix parameter such as swelling, stiffness and pore size on the growth, adhesion and behavior of several yeast strains commonly used in laboratories around the world. We will then use the obtained results in developing research- and industry-based approaches and applications.
1. Palková, Zdena, Multicellular microorganisms: laboratory versus nature. EMBO reports, 2004. 5(5): p. 470
2. Haque, M.A., T. Kurokawa, and J.P. Gong, Super tough double network hydrogels and their application as biomaterials. Polymer, 2012. 53(9): p. 1805-1822.
8:00 PM - BM04.06.11
A Novel Methylcellulose/Gelatin/Calcium Phosphate-Based Thermoresponsive Injectable Bone Substitute
Öznur Demir Oğuz 1 , Duygu Ege 1 Show Abstract
1 Institute of Biomedical Engineering, Boğaziçi University, Istanbul Turkey
From the inspiration of the structural arrangement of the bone itself, this research focuses on the combination of calcium phosphate cements (CPC) with natural polymers. The CPC and polymer based injectable bone substitute (IBS) materials can overcome limitations of CPCs and can improve mechanical strength, viscosity, degradation, and injectability. In this study, methylcellulose (MC) was combined with CPCs because of MC’s thermoresponsive behavior and to tailor IBS’s viscosity and injectability. Also, gelatin (GEL) was incorporated to lower the phase transition temperature and to enhance cell adhesion. MC and GEL combination makes the liquid phase. Sodium citrate dehydrate was also used as the liquid component to crosslink MC and GEL and to reduce gelation time. The powder phase comprised of tetra calcium phosphate (TTCP), and dicalcium phosphate dehydrates (DCPD) and calcium sulfate dehydrates (CSD). TTCP and DCPD have the ability to transform hydroxyapatite in an aqueous environment and CSD was added to the powder phase to optimize the resorption rate as well as to enhance mechanical properties. TTCP was synthesized using a solid-state method. Gelation temperature and time were measured by test tube inversion method. Accordingly, the optimized MC concentration was chosen as 5 and 6 wt% while GEL concentration was kept constant at 2.5 wt% and gelation temperature was measured approximately 15 and 18 min, respectively. The addition of 50 mg/ml sodium citrate dehydrate solution to the optimized MC and GEL mixture reduce the gelation time and it was 5 min for both 5 and 6 wt% MC containing 2.5 wt% GEL samples. After that synthesized IBS was characterized by using X-Ray Diffraction (XRD), Fourier Transform Infrared Analysis (FTIR), Zeta Particle Size Analysis, rheometry, and thermogravimetric analysis. XRD and FTIR analysis proved that TTCP was successfully synthesized with particle size of 430.1 nm. FTIR spectra of liquid phase both amides I at 1633 cm-1 and β-glycosides bonds among saccharide units at 900-1230 cm-1 were indicated that there was a hydrophilic interaction between MC and GEL. Sodium citrate dehydrate was reacting with methyl groups in MC and amino groups in GEL to form ester crosslinking and inter-ionic attraction. According to XRD analysis of optimized powder and liquid phase mixture, only the peaks of TTCP, DCPD, and CSD were observed. Rheometer results showed that all the prepared formulations exhibited Newtonian flow. The mixture of powder and 6 wt% MC containing 2.5 wt% GEL liquid phase sample had the highest viscosity value due to its high concentration of MC. Results of thermogravimetric analysis showed two main decomposition steps for the liquid phase because of the hydrophilic interaction between MC and GEL. Overall, the synthesized thermoresponsive IBS represent promising platforms for future studies in bone tissue engineering and bone-defect repair.
8:00 PM - BM04.06.12
Nanotopography-Induced Neuromuscular Junction Assembly
Eunkyung Ko 1 , Seungjung Yoo 2 , Ziad Mahmassani 1 , Marni Boppart 1 , Sung Gap Im 2 , Rashid Bashir 1 , Hyunjoon Kong 1 Show Abstract
1 , University of Illinois at Urbana-Champaign, Urbana, Illinois, United States, 2 , Korea Advanced Institute of Science and Technology, Daejeon Korea (the Republic of)
Neuromuscular junctions (NMJ), a chemical synapse formed between the motor neurons and skeletal muscle fibers, serve a key role in skeletal muscle movement. There have been efforts to engineer NMJs in vitro by co-culturing motor neurons and skeletal muscle cells, yet majority of those studies were limited to observing the morphology of NMJs. There are still needs to improve functionality of engineered NMJs. In this study, we proposed that cell adherent nano-patterned substrates would enhance better myogenic differentiation of skeletal muscle cells and the resulting patterned muscle fibers would promote neural innervation of the neuronal cells into muscle fibers. To examine the hypothesis we compared the maturity of the myotubes formed on a flat substrate with those on different sizes of linearly patterned substrates and analyzed the neural innervation.
Polyurathane acrylate (PUA) nanopatterend substrates were prepared by applying PUA resin on the silicon master, covering the top with PET film, then curing the PUA resin with ultraviolet light. The resulting substrates were peeled off from the master, and coated with matrigel prior to seeding cells. Muscle cells were cultured cultured in growth medium for the first 3 days, and exchanged with differentiation medium to induce myotube formation. After a week, neural stem cells were seeded on the muscle layer to induce neuromuscular junction (NMJ) formation. NMJs were immunofluorescently stained to image the presence of acetylcholine receptors, and the functionality was analyzed by treating glutamic acid and curare.
The primary myoblasts cultured on the nano-patterend substrates were compared with those on a flat substrate. Morphometric study revealed that primary myoblasts cultured on the nano-patterend substrates align to each other better, and exhibit increased actomyosin expression. Moreover, the patterned substrates provided more focal adhesion points to the cells. Similarly, neural stem cells seeded on the aligned muscle layer expressed more neuronal marker compared to the muscle cells cultured on a flat substrate. Additionally, neuronal cells on the patterend myotubes followed the linear morphology of the muscle cells and aligned with the myotubes in parallel. We confirmed the NMJ formation by immunocluorescently using neurofilament, myosin, and acetylcholine receptor markers. As we analyzed the area of co-localization of 3 markers, expression level was higher on the patterned substrate group.
Our nano-patterend substrates can guide muscle and neural stem cells to form alignment and form NMJ in an orderly manner. The engineered NMJs on the patterned substrates show better functionaly compared to those on a flat substrate since the topography introduces a more in vivo-like environment to the cells. The engineered NMJs has potential as a drug screening tool as well as a platform that may allow us to study NMJ related diseases in molecular levels.
8:00 PM - BM04.06.13
Surface Pattern Formation on Silicone Elastomer Substrate through Photo-Initiated Graft Polymerization of Methacrylate Monomers
Tatsuo Aikawa 1 , Hiroaki Kudo 1 , Takeshi Kondo 1 2 , Makoto Yuasa 1 2 Show Abstract
1 Department of Pure and Applied Chemistry, Tokyo University of Science, Noda Japan, 2 Research Institute for Science and Technology, Tokyo University of Science, Noda Japan
Silicone elastomers have been used as a useful substrate for fabricating biomedical devices such microfluidic devices. Surface modification of silicone elastomers is an important technique for modulating surface properties (e.g., wettability, adhesiveness, optics) of biomedical devices. In addition, surface pattern formation can enhance the surface properties. We describe pattern formation on poly(dimethylsiloxane) (PDMS) through a photo-initiated polymerization of methacrylate monomers. The present study also investigated the influence of stiffness of the PDMS substrate and monomer type on the pattern formation. The expected mechanism for pattern formation in the present system is also discussed.
The PDMS substrate was prepared using a silicone elastomer kit (Sylgard 184) consisting of base and crosslinking regents. The stiffness of the PDMS substrate was controlled through controlling the concentration of the crosslinking regent. A benzophenone-based photoinitiator, 2-trimethylsiloxy-4-allyloxydiphenylketone (0.1 wt%) was added to the PDMS mixture. For photo-initiated polymerization, 15 mL of glycidyl methacrylate (GMA) solution (30 vol% vs. methanol) was added to a glass tube and then deoxygenated by bubbling with Ar for 20 min. A piece of the PDMS substrate (1 × 5 cm2) was plated in the monomer solution and irradiated at room temperature with ultraviolet light. The degree of polymer grafting (Dg) was calculated by Dg =(Wf − Wi)/Wi, where Wi and Wf are the weight of the PDMS substrate before and after graft polymerization, respectively.
Results and Discussion
The Dg value for grafting polyGMA increased with increasing UV irradiation time and the monomer concentrations (up to ~90%). Observation of IR absorption peaks (C=O stretching in the ester at 1731 cm−1 and C-O-C asymmetric stretching in the epoxy ring at 905 cm−1) proved grafting GMA on the PDMS substrate. It was found that pattern formation occurred when the Dg was greater than ~5%. The topology of the pattern was affected by Dg. Area of the convex components of the patterns was enlarged with increasing Dg. Pattern formation was observed on relatively soft PDMS substrates prepared by using 0.9 or 9.1 wt% of the crosslinking agent. The topology of the pattern was influenced by the stiffness of the substrate. The gap width between the convex regions of the pattern increased with substrate softness. In the case of the stiffer substrate, which was prepared using 50% of the crosslinking agent, characteristic patterns were not observed on the surface. The solubility of the monomers used in the polymerization was related to whether or not a pattern forms. When using GMA or benzyl methacrylate, which has relatively low solubility in the PDMS precursor, characteristic patterns were formed. In contrast, when using butyl or adamantly methacrylate, which is soluble in the PDMS precursor, the characteristic pattern was not observed on the PDMS surface.
8:00 PM - BM04.06.15
Effects of Plasma Treatment Power Levels and Treatment Time on Electrospun Polycaprolactone Scaffolds for Regenerative Tissue Engineering Applications
Sofia Donnecke 1 , Robert Chu 1 , Khal Gabriel 1 , Arthur Mcclelland 1 Show Abstract
1 , Harvard College, Cambridge, Massachusetts, United States
Polycaprolactone (PCL) is a widely used biocompatible and biodegradable polymer that has been studied for regenerative tissue engineering applications. Electrospun PCL fibers are hydrophobic as spun and are often plasma treated to increase hydrophilicity and biocompatibility. In this study, electrospun PCL scaffolds were systematically plasma treated with a low pressure, oxygen (20%)-argon (80%) gas mixture at a range of plasma powers and times (5 to 50 W; 15 s to 90 minutes) to study changes in surface chemistry and surface morphology with different treatment conditions. The surface chemistry was characterized using X-ray Photoelectron Spectroscopy (XPS), Attenuated Total Reflectance Fourier Transform Infrared spectroscopy (ATR-FTIR), Raman spectroscopy, contact angle measurements and scanning electron microscopy (SEM). XPS and FTIR data identified an overall increase in the surface oxygen content for all treated samples. No clear trends were seen with plasma treatment time under the conditions tested here.
SEM images show the morphology of the electrospun PCL fibers treated with plasma powers from 5 to 10 W as unchanged, the fibers treated with plasma powers of 15 to 20 W as having a cracked surface and squared off shape, while the PCL fibers treated at plasma power levels above 25W showed a round shape again and strong evidence of fiber melting. Interestingly, the contact angle of the scaffolds actually increased with plasma treatment until the 25W power level when it suddenly became strongly hydrophilic and completely wet. This suggests that the hydrophilicity of the PCL fibers is actually due to changes from the melting and recasting of the PCL fibers.
The sharp threshold that was observed in the contact angle measurements at 25W of power for 15s suggests the minimum effective plasma treatment that could be used to induce hydrophilicity in electrospun PCL scaffolds for regenerative tissue engineering applications.
8:00 PM - BM04.06.16
Characterization of Cell-Seeded Synthetic Scaffold for Esophageal Regeneration
Sherif Soliman 1 , Kalia Burnette 1 , Lena Kalenjian 1 , Elisaveta Todorova 1 , Greg Booker 1 , Arthur Mcclelland 2 , Saverio La Francesca 1 , Lucas Quesnel 1 Show Abstract
1 , Biostage, Holliston, Massachusetts, United States, 2 Center for Nanoscale System, Harvard University, Cambridge, Massachusetts, United States
Esophageal diseases may require resection of the damaged portion. Current standard of care requires the replacement of the esophagus with stomach or the intestine. Such procedures have high rates of mortality and morbidity and highly affect the quality of life of patients. The use of alternative conduits is needed. A tissue engineering approach that allows for the regeneration of esophageal tissues would have significant clinical application. In this study, we describe a bioengineered construct that is comprised of a synthetic scaffold laden with autologous cells that can be surgically implanted to guide regeneration of the esophagus.
The tubular synthetic scaffold was created with electrospun polycarbonate based polyurethane. This was designed to provide a microenvironment conducive to cellular proliferation, with special attention given to the morphological properties, microstructure characteristics, and surface chemistry. In our preclinical model, autologous adipose-derived mesenchymal stem cells were isolated, expanded, and seeded on the scaffold. Scaffolds were then incubated in a dispoasable bioreactor at 37 C to obtain an autologous combination construct. The 6-cm scaffold was implanted in big animal models in place of a 5-cm circumferential resection of the esophagus. Functional, biochemical and histological techniques tracked host tissue growth and stability.
In vitro, the construct dependably carried metabolically active cells that released bioactive molecules supportive of surgical repair and restoration of esophageal function. In vivo trials resulted in tissue growth that were observed to reconstitute the esophagus with a high degree of continuity and integrity after circumferential full thickness surgical resection. Full mucosal regeneration on the inner lumen was observed within a span of 2.5 months post-implantation
We describe an innovative bioengineeered construct that combines autologous cells with a synthetic scaffold for the treatment of patients with esophageal disease. The results demonstrate the feasibility of this approach to facilitate the regeneration of full thickness circumferential defects after esophageal resection as would be clinically required for esophageal malignancy. In addition to the esophagus, we expect the same approach to help patients with disorders of other hollow organs, such as the trachea and bronchus.
8:00 PM - BM04.06.17
Bovine Serum Albumin (BSA) Loaded Core-Shell Nanofibers Modified by Cold Atmospheric Plasma (CAP) for Bone Regeneration
Yangfang Zhou 1 , Mian Wang 1 , Michael Keidar 2 , Thomas Webster 1 3 4 Show Abstract
1 , Northeastern University, Boston, Massachusetts, United States, 2 , George Washington University, Washington, District of Columbia, United States, 3 , Wenzhou Institute of Biomaterials and Engineering, Wenzhou China, 4 , Wenzhou Medical University, Wenzhou China
Coaxial electrospinning is a novel technique for producing core-shell nanofibers which combine the advantages of having a multi-layer structure and the ability to deliver bioactive agents. In order to mimic the properties of the extracellular matrix, scaffolds were modified by cold atmospheric plasma (CAP) which is an ionized gas where the ion temperature is close to room temperature. It contains electrons, reactive oxygen species (ROS), reactive nitrogen species (RNS), radicals, and UV light . A previous study showed that plasma surface modification (PSM) could create a nano-structured surface and change surface hydrophilicity without affecting other surface chemicals . The CAP modification not only increased the surface roughness of the nanofibers , but it also changed the release profile of water-soluble agents that were loaded into the core fibers. The objective of the present student was to study the impact of cold atmospheric plasma (CAP) on core-shell nanofibers' cytocompatibility as well as their drug release profiles.
Nanofibers were electrospun by two different polymers. Bovine serum albumin (BSA) was dissolved in a polyvinyl alcohol (PVA) solution as the core polymer, while the shell polymer was poly-l-lactide (PLLA). Then, core-shell nanofibers were fabricated by a coaxial electrospinning technique. Control samples were made without loading BSA into the core polymer. Then, BSA loaded and non-BSA (control) loaded nanofibers were modified by CAP for 60s. Finally, four different samples were fabricated including BSA and non-BSA loaded nanofibers, CAP treated BSA and non-BSA loaded nanofibers.
Cell proliferation assays with fibroblasts and osteoblasts showed that BSA loaded nanofibers with CAP treatment significantly increased the density of both cells. Moreover, BSA loaded scaffolds and CAP modified scaffolds had similar effects on cell proliferation. Scanning electron microscope (SEM) images illustrated that CAP treatments could increase the pore size and pore number on the shell. These results proved the hypothesis that CAP treatments can improve cytocompatibility properties of scaffolds as a result from their roughness and hydrophilicity. Moreover, CAP treatments promoted the release of bioactive agents that were loaded in the core polymer because nanofibers that were treated by CAP for 60s and 90s released drugs faster than those modified by CAP for less time.
In conclusion, CAP modification can play an important role in enhancing the cytocompatibility of scaffolds as well as in controlling drug release, and, thus should be further studied for numerous bone tissue engineering applications.
References:  M. Wang, X. Cheng, W. Zhu, et al. Tissue engineering: part A, 2014, 5(20), 1060-1071.
 M. Wang et al. Acta Biomater, 2016, 12 (46): 256-265.
8:00 PM - BM04.06.18
Dental Pulp Stem Cell Growth and Differentiation on a Paramagnetic Polymer Composite
John Chen 1 , Zaiff Khan 1 , Anoushka Guha 1 , Rebecca Isseroff 1 2 , Simon Lin 2 , Juyi Li 2 , Kuan-Che Feng 2 , Linxi Zhang 2 , Marcia Simon 3 , Miriam Rafailovich 2 Show Abstract
1 , Lawrence High School, Cedarhurst, New York, United States, 2 Department of Materials Science and Engineering, Stony Brook University, Stony Brook, New York, United States, 3 , Stony Brook University School of Dental Medicine, Stony Brook, New York, United States
Human dental pulp stem cells (DPSCs) can differentiate, showing potential for regenerative medicine. Recently, Poly-lactic acid (PLA), a biocompatible polymer, is increasingly being implemented into biomedical application since it is easily adapted to FDM or 3D printing technologies. Since DPSCs have been proposed for regeneration of bone and teeth, and PLA is frequently used as a scaffold, we have investigated the influence of PLA and PLA nanocomposites on DPSC differentiation. We found that while the plating efficiency and proliferation of DPSC on PLA homopolymer was poor, incorporation of Graphene Oxide (GO) or partially Reduced Graphene Oxide (pRGO) increased both processes to levels comparable to those measured on tissue culture plastic (TCP) controls. Both processes were further enhanced when cells were grown in a 1 Tesla external magnetic field on PLA nancomposite substrates containing GO to which we added Iron pentacarbonyl (Fe-GO); or GO reduced together with Iron pentacarbonyl to form Fe-pRGO.
Magnetometry and TEM analysis indicated that Fe-pRGO particles were paramagnetic with Fe particles of approximately 6nm in diameter. PLA nanocomposite scaffolds were made by spin casting solutions of PLA mixed with GO, pRGO, Fe-GO, or Fe-pRGO particles onto silicon wafers and annealing them. DPSCs were plated and cultured on these scaffolds for up to 28 days, where large amounts of biomineralization or HA deposits (as determined by SEM and EDX) were found on all the nanocomposite scaffolds containing GO and pRGO, as well as the Fe-modified GO or pRGO. RT-PCR indicated up-regulation of OCN and ALP genetic markers occurring only on the cultures where biomineral deposition occurred, confirming that the deposits were associated with the differentiation of the DPSCs along osteogenic lineage in the absence of commonly used external differentiation triggers, such as dexamethosone. Biomineralization occurred regardless of the application of the external magnetic field; hence, its influence remains to be studied in greater detail. These results show that in contrast to neat PLA, an iron/graphene/PLA composite may be used to cultivate and differentiate DPSCs for bone replacement therapy.
8:00 PM - BM04.06.19
Bio-Inactive Material Design for Bone Regeneration Using Functional Polymers
Kazuhira Masuyama 1 , Tadashi Nakaji-Hirabayashi 1 2 , Ryuichi Moteki 1 , Hiromi Kitano 1 , Akira Taguchi 3 , Kazuaki Matsumura 4 Show Abstract
1 Graduate School of Science and Engineering, University of Toyama, Toyama Japan, 2 Graduate School of Innovative Life Science, University of Toyama, Toyama Japan, 3 , Hydrogen Isotope Research Center, University of Toyama, Toyama Japan, 4 , School of Material Science, Japan Advanced Institute of Science and Technology, Ishikawa Japan
A creation of biomaterials having an ability to regulate the response to cells and tissues contributes to the further development of not only cell-based therapies but also cell and molecular biology. We have developed two bio-inactive materials from the view point of charge-neutralization: (1) one is neutralized between polymers by mixing of anion/cation polymers and (2) the other is intramolecularly neutralized using zwitterionic polymers. The surface net-charges of both materials are neutral as the same as typical neutral polymer materials, whereas these materials enhanced uniform-construction of the rigid hydroxyapatite layer (enamelum) on the surfaces. It should be noted that the polymer surfaces significantly suppressed protein adsorptions, namely, an interfusion of proteins into HAp layer.
Briefly, we developed a composite material composed of chitosan (Ch) nanofiber, carboxymethyl cellulose (CMC) nanofiber and poly(vinyl alcohol) (PVA). We previously reported that the sheet material composed of with Ch and CMC nanofibers had strong mechanical property because of charge-neutralization between amine group of Ch and carboxyl group of CMC, whereas a flexibility of material was not sufficient [Ref: Colloids surf. B, 2016, 139, 95-9]. We could improve a flexibility of nanofiber material by constructing a polymer alloy with PVA. As described above, the surface of the composite material significantly suppressed protein adsorption and facilitated the construction of rigid HAp layer because of charge-neutralization. This composite material is expected to be applied as a bone prosthetic material.
Moreover, it was discovered that titanium alloy substrate covalently-modified with zwitterionic polymer, poly(carboxymethyl betaine) (PCMB), significantly suppressed the nonspecific adsorption of proteins and the construction of rigid HAp layer on the polymer were facilitated drastically. This technique will be applicable to the development of implant materials composed of titanium alloy (Ti6Al4V).
8:00 PM - BM04.06.20
Graphene Oxide-Containing Electrsopun Nanofibers for Cell Scaffold
Thiers Uehara 1 , Iêda Paino 3 , Fabricio Santos 1 , Vanessa Scagion 4 , Daniel Correa 2 , Valtencir Zucolotto 1 Show Abstract
1 Physics Institute of São Carlos, Univ of Sao Paulo, Sao Carlos, São Paulo, Brazil, 3 Physics Institute of São Carlos, University of São Paulo, São Carlos, São Paulo, Brazil, 4 National Laboratory for Nanotechnology in Agribusiness , Embrapa, São Carlos, São Paulo, Brazil, 2 National Laboratory for Nanotechnology in Agribusiness , Embrapa Instrumentation, Sao Carlos, Sao Paulo, Brazil
Graphene oxide (GO) has been widely explored in biomedical applications as active engineered materials for diagnosis and therapy. With the great potential of using these nanocomposites in biological systems, such as in the manufacture of materials for biotechnological application, it is very interesting to understand the impacts these materials could exhibit on living cells. In this study we investigated the interactions between C2C12 cells on random and aligned GO-containing poly(caprolactone) PCL nanofibers produced via electrospinning system. Plasma etching is important to modify the polymeric surface and to promote the interactions of the fibers with graphene oxide. Scanning Electron Microscopy and Fluorescence Microscopy revealed the morphology and orientation of C2C12 cells on PCL nanofibers (random and aligned). The investigation of the C2C12 cells on graphene oxide/PCL nanofibers may be relevant for medicine and nanotoxicological studies.
8:00 PM - BM04.06.21
Biomimetic Calcium Phosphates-Based Coatings Deposited on Binary Ti-Mo Alloys Modified by Laser Beam Irradiation for Biomaterial/Clinical Applications
Márcio Santos 1 , Carla Riccardi 2 , Edson Filho 3 , Antônio Guastaldi 3 Show Abstract
1 CCNH, Universidade Federal do ABC, Santo André - SP, SP, Brazil, 2 School of Agricultural Sciences, São Paulo State University, UNESP, São Paulo Brazil, 3 Chemistry, São Paulo State University, UNESP, Sao Paulo Brazil
Biomimetic Method has been widely used to prepare a calcium phosphate coatings on Ti and its alloys. This modification is based on a Synthetic/simulated Body Fluid (BSF) which facilitates the mimicking of the biological process in order to provide hard tissue repairs. The formation of HA and other calcium phosphates under biological medium and SBF occurs in the presence of Ca2+ and PO43- ions, as well as essential ions such as: Mg2+, HCO3-, K+ and Na+. Ti-15Mo alloy samples were irradiated by pulsed Yb: YAG pulsed laser beam under air and atmospheric pressure. Sequentially, calcium phosphate coatings were deposited on the irradiated surfaces by the biomimetic method. The biomimetic calcium phosphates-based surfaces were submitted to heat treatment conditions at 350°C and 600°C. The present study correlates two conditions of fluency (1,91 and 5,54 J.cm-2) as established have a sufficient energy to promote ablation on the laser beam irradiated surfaces. Likewise, it has been demonstrated the processes of fusion and fast solidification from the laser beam irradiation, under ambient atmosphere, inducing the formation of stoichiometric TiO2 and non-stoichiometric titanium oxides, including Ti3O5, TiO, Ti3O and Ti6O with different oxide percentages depending on the fluency applied. Besides that, laser modification has allowed a clean and reproducible process, providing no traces of contamination, an important feature for clinical applications. The physico-chemical and morphological analysis indicated the formation of a multiphase coatings depending on the heat treatment temperature performed to 350 °C (ACP1 and 2, HAD, HA phases) and 600 °C (HAD, HA and β-TCP phases). It is worth noting that multiphasic bioceramic systems has been gaining attention for biomedical applications. Thus, the laser beam irradiation associated to bioactive coatings of calcium phosphates of biological interest have shown to be promising and economically feasible for use in dental and orthopedic implants.
The authors thank to the grant 2014/05626-8, São Paulo Research Foundation (FAPESP) and CNPQ by the financial support.
8:00 PM - BM04.06.22
Human, iPSC-Derived Brain Endothelial Lumen in Multiscale Channels of Perfused Hydrogels
Jason Wang 1 , Shannon Faley 1 , Brian O'Grady 1 , Jung Bok Lee 1 , Emma Hollmann 1 , Ethan Lippmann 1 , Leon Bellan 1 Show Abstract
1 , Vanderbilt University, Nashville, Tennessee, United States
An accurate 3D in vitro tissue model of the brain neurovascular unit (NVU) implemented with physiologically representative human cells is an invaluable tool in the field of regenerative medicine to further the limited understanding of the biochemical and biomechanical interactions involved between the multitudes of cells relevant to studying neurovascular behavior. 3D tissue models of brain vasculature containing human cells have been difficult to develop due to limitations in obtaining primary neuronal cells that include 1) ethical considerations, 2) lack of cell lines that accurately represent physiological function, 3) difficulty in obtaining matching cell types needed to form a NVU (e.g. BMECs, pericytes, astrocytes, and neurons [1, 2]), and 4) fabrication of a platform that enables immediate perfusion of tissue-scale constructs .
Our aim is to overcome these challenges through the implementation of a stable source of matched iPSC-derived cells . These iPSC-derived cells are used to populate large, immediately-perfusable gelatin hydrogels through channel networks generated by sacrificial fabrication methods [5-7].
This work describes the formation and characterization of an endothelial lumen with barrier functionality using iPSC-derived human brain microvascular endothelial cells (iPSC-hBMECs) in a gelatin hydrogel containing macro- and micro-channels (mimicking arterioles and capillaries) using 3D printed and electrospun PNIPAM templates .
Our studies assess iPSC-hBMECs to determine: 1) cell viability and proliferation of cells in presence of PNIPAM, 2) formation of a true lumen within the macro and micro channels, 3) perfusion of cells throughout the hydrogel, and 4) verification of cell function post-perfusion through an efflux transporter assay and analysis of a dextran diffusion assay. The cell function is further reinforced through immunostaining for VE-cadherin, claudin5, and ZO-1 to verify the formation of tight junctions responsible for the barrier characteristics between the iPSC-hBMECs.
The ability to characterize and implement matching human neuronal cells in a biomimetic, perfusable hydrogel scaffold is a critical step towards developing a truly physiology-representative, tissue-scale in vitro model of the blood brain barrier. Future effort will involve the use of co-cultures of other iPSC neuronal cells (such as iPSC-astrocytes and iPSC-pericytes) in order to further improve the biomimetics of the neurovascular unit.
1. Gordon, J., S. Amini, and M.K. White,. Methods Mol Biol, 2013. 1078: p. 1-8.
2. Huang, Y., J.C. Williams, and S.M. Johnson,. Lab Chip, 2012. 12(12): p. 2103-17.
3. Griep, L.M., et al., Biomed Microdevices, 2013. 15(1): p. 145-50.
4. Lippmann, E.S., et al., Sci Rep, 2014. 4: p. 4160.
5. Bellan, L.M., et al., Adv Mater, 2012. 24(38): p. 5187-91.
6. Faley, S.L., et al., Biomicrofluidics, 2015. 9(3): p. 036501.
7. Lee, J.B., et al., Adv Healthc Mater, 2016. 5(7): p. 781-5.
8:00 PM - BM04.06.23
Ordered Honeycomb Petri Dish Used as a Scaffold for Cell Growth
Van-Tien Bui 1 , Ho Suk Choi 1 Show Abstract
1 , Chungnam National University, Daejeon Korea (the Republic of)
Three-dimensional (3-D) culture dish with ordered honeycomb pattern as an artificial human ovary where cells can behave as they do in vivo demonstrates a great application potentials in biotechnology. Here, we report on a very simple, low-cost, scalable and single-step method to directly create ordered controllable honeycomb patterns on polymeric substrates including the commercial Petri dish in normal air. The method based on the combination of the methanol accumulation-induced phase separation and the dip-coating technique in which a mixture of chloroform and methanol is exploited not only to induce a ternary solution but also to guarantee the formation of highly ordered honeycomb structure on the substrate. The surface topology of honeycomb substrate is effectively controlled by varying experimental conditions and the obtained honeycomb structure is indeed a part of the substrate revealing an increase in the structure’s stability. Moreover, honeycomb-structured Petri dishes with controllable pore arrays produced using this method shows good cell adhesion and proliferation as an evidence for use in biomedical applications.
8:00 PM - BM04.06.24
A Simple and Versatile Fabrication of a Tunable Nanofiber Membrane-Integrated Transwell Insert via Electrolyte-Assisted Electrospinning
Seongsu Eom 1 , Sang Min Park 1 , Seon Jin Han 1 , Dong Sung Kim 1 Show Abstract
1 , Pohang University of Science and Technology, Pohang Korea (the Republic of)
A transwell insert, which has a separating permeable membrane, is the most widely utilized in vitro cell culture platform to analyze chemotactic behavior and develop co-culture model. Although the commercially available transwell inserts have low cost and great accessibility, the flat porous membrane of transwell insert has limited capability, due to its structural difference from in vivo environment. The integration of Transwell assay with an electrospun nanofiber membrane, which has a structure similar to the natural extracellular matrix, shows a significant potential in chemotactic assays and co-culture models. However, the previous studies have utilized multi-step process to make nanofiber membrane-integrated transwell insert, which includes peeling off nanofibers from metal collector, cutting and attaching the membrane on the well insert wall. These additional processes after electrospinning might lead to misalignment and low reproducibility. To overcome this limitation, we firstly developed a simple and versatile fabrication process of tunable nanofiber membrane-integrated transwell inserts by adopting electrolyte-assisted electrospinning that utilizes electrolyte solution as a collector instead of a metal. The fabrication process enables simultaneous fabrication and integration of a nanofiber membrane on a transwell insert without manual manipulation. Moreover, this process has the capability to modulate the properties of nanofiber membrane such as diameter of the nanofibers and porosity and thickness of the nanofiber membrane. On the nanofiber membrane-integrated transwell inserts, bEnd.3 cells were cultured and the trans-endothelial electrical resistance(TEER) values were analyzed, to validate the biological relevance.
8:00 PM - BM04.06.25
The Potential of L-Arginine/Hydroxyapatite Coatings to Enhance Osseointegration of Implants
Ilayda Duru 1 , Duygu Ege 1 Show Abstract
1 Institute of Biomedical Engineering, Bogazici Univ, Istanbul Turkey
Bone bonding is a crucial process for cementless hip implants and bioactive ceramic coatings have been widely used on these implants in order to induce the bone ingrowth. However, these coatings may suffer from delamination and the healing after surgery still takes too long. L-arginine (L-arg) is of great interest in biomedical studies due to its promising effect on osteogenic cell activity and apatite formation. The studies show that L-arg can have electrostatic interaction with cell membranes and assist the cellular uptake of attached particles. L-arg/hydroxyapatite (HA) coatings possess self-repairing ability. In the present study, we introduce a new route to produce L-arginine/hydroxyapatite coatings and investigate their potential to promote osteogenesis on hip implants.
Firstly, the surfaces of Ti6Al4V substrates were polished and then etched in a concentrated solution of HCl/H2SO4. ZrO2 was deposited on Ti6Al4V as an intermediate layer between HA and Ti6Al4V by reactive magnetron sputtering. Reactive magnetron sputtering was processed at 200 W and 200 °C for 4 h. HA coating on ZrO2 layer was produced by electrophoretic deposition at 30 V in 205 s. suspension was obtained by ultrasonic and magnetic agitation of the solution of 1 g HA in 35 ml isopropanol for 3 h. After the coating process, HA coatings were dried at room temperature and sintered at 600 °C for 1 h in ambient atmosphere in a furnace. HA-coated samples were immersed in L-arg solution in phosphate buffered saline and incubated at room temperature for 24 h. SEM, AFM, Raman, XRD, FTIR and XPS results were examined to characterize L-arg/HA coatings on ZrO2-coated Ti6Al4V implants. Finally cell culture studies were carried out with mesenchymal cells.
SEM and AFM images of etched substrates demonstrated the rough topography and average Ra value was found around 4.0 µm. Raman and XRD spectra of produced ZrO2 layer revealed the characteristic peaks of monoclinic structure. The primary bands on Raman spectrum were a doublet at 178 and 192 cm-1, a broad peak at 475 cm-1. Moreover, XRD spectrum showed that the coating had a single-phase structure composed of monoclinic ZrO2. FTIR spectrum of L-arg/HA coating revealed the significant peaks at 1224 cm-1 and 1739 cm-1 which was accompanied by a small peak at 1719 cm-1. These bands represent the C-N bond and protonated carboxyl group. XPS analyses demonstrated the increase of C/Ca ratio with the addition of L-arg. Ca/P ratio was decreased which showed the vacancies of calcium at the surface of hydroxyapatite. C1s spectrum of L-arg/HA coating also demonstrated the C-C, C=O and C-N bonds. It might be concluded that L-arg/HA coatings were formed by attachment of the carboxyl group of L-arg to calcium atoms of HA. Overall, L-arg/HA coatings were successfully coated on ZrO2-coated Ti6Al4V hip implants. Finally, cell culture study with mesenchymal cell on the study groups showed high potential of these coatings for biomedical applications.
8:00 PM - BM04.06.26
Potential Enhancement of Osseointegration with Carbon Nanotube/Sulfonated Poly(ether ether ketone)
Hatice Kaya 1 , Osman Bulut 2 , Duygu Ege 1 Show Abstract
1 Institute of Biomedical Engineering, Boğaziçi University, Istanbul Turkey, 2 Faculty of Civil Engineering, Istanbul Technical University, Istanbul Turkey
In recent years, L-arginine has been studied extensively to improve biological response of biomaterials and polyether ether ketone (PEEK) has arisen as an alternative to metallic biomaterial for load-bearing implant application, which overcame the adverse effects of the conventional implants such as stress shielding, release of toxic ions and radiotherapy interference. However, as PEEK is a bioinert polymer, it leads to poor osseointegration and restricts its potential applications where the direct contact with bone is required. Chemical modification techniques have been developed to enhance hydrophilicity of PEEK without compromising the physical properties of the bulk material. In this study, sulfonated PEEK (SPEEK) and carboxyl functionalized multi-walled carbon nanotube (f-WMCNT) composite films were fabricated by solvent casting. 10 wt. % of SPEEK with three different concentration of f-MWCNT (0.5, 1 and 2 wt. %) were dissolved in N,N-Dimethylmethanamide (DMF). The surface of the obtained nanofilms were covalently conjugated with L-arginine. The samples were characterized with Proton Nuclear Magnetic Resonance Spectroscopy (H-NMR), Fourier Transform Infrared Spectroscopy (FTIR), X-ray Photoelectron Spectroscopy (XPS), Contact Angle Analyzer and Dynamic Mechanical Analysis (DMA).
A distinct peak was observed in the H-NMR spectrum at 7.5 ppm (HE) which revealed the presence of sulfur (-SO3H) groups in the hydroquinone ring of repeated PEEK unit. The degree of sulfonation of PEEK was calculated as 81%. The FTIR bands were observed at 1020 cm-1, 1076 cm-1 and 1254 cm-1 which were assigned to S=O stretching, symmetric and asymmetric stretching vibration of (O=S=O), respectively. The band at 1486 cm-1 corresponding to the aromatic C–C band in PEEK was observed to split into two new absorption bands at 1471 cm-1 and 1491 cm-1 for SPEEK that was attributed to the substitution upon sulfonation. Therefore, the FTIR spectra confirmed successful sulfonation of PEEK. After L-arginine modification, XPS revealed increase of the atomic concentration of nitrogen (N) from 1.13 % to 3.12 %, which indicated covalent conjugation of L-arginine on the surface of the films. In addition, high resolution XPS spectrum showed a peak at 399.56 eV which may indicate binding energy of the N-C new bond formation. The contact angle of CNT/SPEEK films were 60.3°, 56.57° and 50.8° for 0.5, 1 and 2 wt. % CNT and that of pure SPEEK films and PEEK 67.77° and 103.5°, respectively. Therefore, wettability of CNT/SPEEK films were much greater than pure PEEK, which is advantages for biomedical applications. Finally, advanced mechanical strength was proved by DMA analysis. The storage modulus at 37°C for 0, 0.5, 1 and 2 wt. % are 1188 MPa, 1500 MPa, 1819 MPa and 2261 MPa, respectively. This study indicates that L-arginine modified CNT/PEEK films are promising candidates for femoral replacement applications.
8:00 PM - BM04.06.27
3D Printed iPSC-Derived Neurobionic Constructs for Neurodevelopmental Disorder Modeling and Therapeutic Screening
Kiavash Kiaee 1 , Yasamin Aliashrafi Jodat 1 , Sudeep Joshi 1 , Manu Mannoor 1 Show Abstract
1 Mechanical Engineering, Stevens Institute of Technology, Hoboken, New Jersey, United States
Neurodevelopmental disorders (NDs) such as autism spectrum disorders (ASD), Down syndrome and intellectual disability (ID), are impairments that affect the development and growth of the human brain during embryonic and early postnatal life. Existing substandard prognosis and ineffective therapeutic interventions in such cases can be attributed to: (i) poor understanding of the developmental etiology of NDs (ii) inefficiency of rodent models as well as postmortem samples, failing to recapitulate disease specific phenotypes for these complex network disorders, and (iii) inability of 2D cell cultures systems to reproduce essential biological characteristics of the complex and dynamic 3D in vivo brain environment, namely, cell function, differentiation, drug metabolism, gene expression, morphology and viability.
Human induced pluripotent stem cells (iPSCs) are a promising new tool to study neurological disorders since this cell type can be derived from patients as a genetically tractable and renewable source for cells that are otherwise difficult to acquire, such as neurons and astrocytes. Here we present an approach based on 3D printing of iPSC-derived neuronal cells to model human brain development and study complex neurodevelopmental features. Astrocytes and neurons differentiated from donor derived iPSCs were seeded into hydrogels, and 3D printed into spatially organized and confined configurations using a customized micro-extrusion pneumatic system. Therefore, maximizing the potential of iPSCs to create three dimensional in vitro cellular constructs that faithfully simulate human brain tissue. The printed co-culture was analyzed for several phenotypes including migration characteristics, synapse formation and soma size.
Remarkably, such 3D printed cerebral organoids could serve as a reductionist model to study the mechanisms of early in utero human brain development under controlled conditions. Taken together, this work will lead to the creation of 3D multicellular brain organoids derived from human iPSCs that faithfully recapitulate the program of early neurological development in humans and in addition, serve as ideal platforms for screening various therapeutic agents.
Gulden Camci-Unal, University of Massachusetts Lowell
Surya Mallapragada, Iowa State University
Matteo Moretti, IRCCS Insituto Ortopedico Galeazzi
Pamela Yelick, Tufts University
Multifunctional Materials | IOP Publishing
BM04.07: Polymeric Biomaterials for Regenerative Engineering II
Wednesday AM, November 29, 2017
Sheraton, 2nd Floor, Independence West
8:00 AM - BM04.07.01
Multi-Photon Direct Laser Writing and 3D Imaging of Nano-Scale Resolution 3D Architectures for Cell Colonization
Angelo Accardo 1 , Marie-Charline Blatche 1 , Remi Courson 1 , Isabelle Loubinoux 2 , Christophe Thibault 1 , Laurent Malaquin 1 , Christophe Vieu 1 Show Abstract
1 , LAAS-CNRS, Toulouse France, 2 , ToNIC, Toulouse NeuroImaging Center - INSERM, Toulouse France
Engineering of efficient scaffolds for enhancing and guiding the growth of cells is an issue that several scientific communities are addressing in the last decade. Although most of the current methodologies are still involving 2D architectures [1,2], the transition from cell monolayers to 3D cultures is motivated by the need to work with cellular models that mimic the features of natural tissues and by the possibility to employ in perspective such engineered tools as bio-implants to repair damaged areas of our body. There is therefore the urgent need to develop high-resolution 3D fabrication approaches and 3D imaging protocols able both to favor ideal cell proliferation features and to provide a clear scenario of the cell colonization mechanisms.
Here, we report the combination of advanced 3D fluorescence imaging techniques and 3D architectures realized by a direct laser writing (DLW) fabrication approach  featuring a much higher level of accuracy at the micro- and nano-scale compared to other 3D fabrication technologies such as single-photon stereolithography, extrusion based approaches and inkjet printing . For achieving freestanding architectures with sub-micrometric resolution (≈ 200 nm), we exploited a two-photon polymerization technique. To assess the colonization efficiency around and within the developed architectures, we choose a fast growing neuroblastoma cell line (N2A) known to express several properties of neurons such as the formation of neuritic extensions or interconnections. For unveiling the localization and the morphology of the cells not only around the 3D scaffold but also within the scaffold’s core, SEM (Scanning Electron Microscopy) characterization was associated with light sheet fluorescence microscopy (LSFM) and two-photon confocal imaging (2PI) able to “shine light” in the most inaccessible regions of the architecture. The multi-technique characterization approach revealed an optimal cell invasion both around the fabricated architecture and within its core regions as well as the capability to form freestanding cellular networks and long neuritic extensions (up to ≈ 60 μm length) especially around the nano-gratings induced by the DLW fabrication.
The developed 3D fabrication/3D imaging protocols open therefore a wide scenario of neuroscientific applications especially in the field of neural tissue engineering as well as for evaluating the action of biomedically relevant biomolecules on neural networks in 3D in-vitro environments.
 L. Vaysse, A. Beduer, J. C. Sol, C. Vieu, I. Loubinoux, Biomaterials 58 (2015) 46-53.
 F. Cesca, T. Limongi, A. Accardo, A. Rocchi, M. Orlando, V. Shalabaeva, E. Di Fabrizio, F. Benfenati, RSC Adv. 4 (2014) 45696-45702.
 A. Accardo, M.-C. Blatché, R. Courson, I. Loubinoux, C. Thibault, L. Malaquin, C. Vieu, Small, in press (2017) DOI: 10.1002/smll.201700621 (Selected as Back Cover)
 M. Guvendiren, J. Molde, R. M. D. Soares, J. Kohn, ACS Biomater. Sci. Eng. 2 (2016) 1679-1693.
8:15 AM - BM04.07.02
Self-Folding 3D Nanoladder as a New Model Scaffold to Control the Directionality Regeneration after Spinal Cord Injuries
Yimin Huang 1 , Chen Yang 1 Show Abstract
1 , Boston University, Brookline, Massachusetts, United States
Spinal cord injury (SCI) often leads to permanent paralysis and loss of sensation below the site of injury because of the inability of damaged axons to regenerate in the adult central neuron system (CNS). Although various strategies were tested to stimulate axonal regeneration at a site of SCI, the growth of axons is generally disorganized and random. Biocompatible scaffolds that guide and maintain the native organization of axons regenerating through an injury site could be of importance in enhancing recovery of the nervous system after injury. Inspired by the natural structure of spinal cord, here we report a nanoladder scaffold to mimic the multi-scale hierarchically organized axonal bundles in the spinal cord. In this study, a nanoladder scaffold is fabricated on the cover glass, via photolithography and reactive ion etching. Cultured embryonic neuron cells on the nanoladder scaffold demonstrated a significant neurite elongation and alignment of the neuron in parallel with the nanoladder direction. Further, using organotypical spinal cord slices as injured model, an enhanced axonal regeneration together with the functional connection can be monitored, which demonstrates the efficacy of using such scaffold in facilitating and guiding directional neuronal regeneration after SCI. Taking consideration of the 3D nature of spinal cord, a bilayer self-folded 3D nanoladder scaffold has also been developed and studied. A simple and robust approach is used to fabricate the Si-based nanoladder structures using photolithography in combination with electron-beam evaporation. Because of its high biocompatibility, visible light induced cross-linkable gelatin is used as a self-folding film. By combining two components together, a bilayer self-rolled tube, which can be irreversibly folded at 37°C, is obtained in response to external thermal stimuli to form complex 3D structures. Such 3D scaffold has the potential to be injected into the injured site. In all, we suggest that our nanoladder scaffold could be used as efficient strategies for the design and manipulation of various functional platforms for advanced tissue regenerative applications.
8:30 AM - BM04.07.03
Nanofibrous Scaffolds Produced by Electrospinning, Rotary-Jet Spinning and Airbrush for Orthopedic Regeneration
Paria Ghannadian 1 , Mirian De Paula 2 , Siddhi Kankaya 4 , Akhil Agarwal 1 , Anderson Lobo 3 , Thomas Webster 1 Show Abstract
1 Chemical Engineering, Northeastern University, Boston, Massachusetts, United States, 2 , Universida de do Vale do Paraiba, São José dos Campos, Sao Paulo, Brazil, 4 Pharmaceutical Sciences, Northeastern University, Boston, Massachusetts, United States, 3 , Universidade Brasil, Itaquera, Sao Paulo, Brazil
Polycaprolactone (PCL) is a bioresorbable polymer with potential applications for bone and cartilage repair. In this study, polycaprolactone fibers (with and without hydroxyapatite nano particles (nHAp) and carbon nanotubes (CNT)) were produced using three different methods: electrospinning, rotary-jet spinning and airbrush. The scaffolds were characterized using contact angles, atomic force microscopy, differential scanning calorimetry, scanning electron microscopy, and transmission electron microscopy and were subjected to cell culture, bacterial assays and mechanical (tensile) testing. The biological and material properties were studied to understand how the various fabrication techniques and nanoparticles affect fibroblasts, gram positive and gram negative bacteria growth on samples. Experiments showed no toxic effect on cells and a decrease in bacterial density (in range of 4% in electrospinning to 80% in rotary-jet spinning) by adding nHAp and CNT to the PCL scaffolds without using growth factors or antibiotics.
8:45 AM - BM04.07.04
Peptide Based Materials for Cardiac Regeneration—2D Patch and 3D Injectable Technologies
Aline Miller 1 Show Abstract
1 , University of Manchester, Manchester United Kingdom
Heart failure (HF) is a debilitating condition that has serious implications for individuals affected in terms of expectancy and quality of live. HF can have many causes including hypertension, coronary artery disease and myocardial infarction (heart attack). The improvement in acute myocardial infarction (AMI) survival rate is thought to be one of the major factors contributing in the dramatic increase in HF cases seen in recent years. Adult cardiomyocytes have a limited capacity to regenerate, the lost myocytes are normally replaced by fibroblasts and myofibroblasts which form scar tissues. The formation of this non contracting fibrous scar alters the workload of the remaining myocardium resulting in remodelling of the heart leading ultimately to congestive HF. Regeneration of the myocardium is therefore essential for the recovery and survival of patients. Here, I will outline how we capitilised on our 15 years of experience of designing self-assembling beta-sheet rich peptide hydrogels to optimise hydrogel property and function to deliver of cardiac cells for the direct regeneration of heart tissue using two novel approaches; 1) the development of a 3D injectable peptide based hydrogel, and 2) cardiac patch, for the regeneration and repair of infarcted heart.
The 3D injectable hydrogel system is a beta-sheet fibrillar hydrogel composed of self-assembling octa-peptide FEFEFKFK (F: phenylalanine; K: Lysine; E: Glutamic acid). This hydrogel is well known to support a number of different cell types, and here Cardiac Progenitor Cells (CPCs) were encapsulated and explored the hydrogels biological properties in vitro, in terms of biocompatibility, cell adhesion and survival, before demonstrating its injectiability and ability to deliver CPCs in vivo using a small animal model. Of particular interest was the ability of the gel to remain at the site of injection and retain its ‘cargo’ for up to 14 days as cells differentiated and formed cardiac tissue. We also noted the ability of the system to support cell homing.
In terms of the patch, we have developed a novel biodegradable biocompatible cardiac patch based on a 3 component concept to provide a structural reinforcement to infarcted heart, which presents tissue remodelling and can also be used for the topical delivery of drugs and cells. I will outline how we demonstrated this in vitro before going onto show in vivo evidence of suturability of the patch, cell migration from the patch to the heart, and the capacity of the device to withstand large blood pressure variations where rats, pigs and sheep were all used in a sequential study.
In each case, injectable or patch delivery of CPCs, resulted in a > 50% reduction in heart chamber volume in studies using rats in comparison to the control, thus demonstrating the potential of both technologies for addressing the big issue of HF.
9:00 AM - *BM04.07.05
Electroconductive Hydrogels for Biosensors and Printable Bioelectronics
Anthony Guiseppi-Elie 1 2 Show Abstract
1 , Texas A&M University, College Station, Texas, United States, 2 , ABTECH Scientific, Inc., Richmond, Virginia, United States
Electroconductive hydrogels (ECHs) are bioactive and stimuli responsive polymers for use as neural prostheses, soft electronics, biosensors, and electro-stimulated drug eluting devices. ECHs are nanocomposites that combine the redox switching and electrical properties of inherently conductive electroactive polymers (CEPs), such as the polypyrroles, polyanilines and polythiophenes, with the inherent biocompatibility and fast ion transport characteristics of hydrated natural (e.g. Chitosan) and synthetic (e.g. poly(HEMA-co-PEG) hydrogels. The synthesis and in vitro characterization of 3-D scaffolds of these hybrid polymers were studied in relation to engineered physicochemical properties. Interpenetrating networks of potentiostatically electropolymerized polypyrrole (PPy-co-PyBA) were formed within poly(hydroxyethylmethacrylate)-based (poly(HEMA-co-PEG) hydrogels on chemically modified gold surfaces. Separately synthesized polyaniline-chloride (PAn-Cl) nanofibers were dispersed within chitosan (CHI) to yield nanocomposites with tunable ionic and electronic conductivity. The influence of ECH composition on the electrical, electrochemical and mechanical properties and post synthesis processing to influence porosity were investigated in relation to the morphology and viability/proliferation of rat pheochromocytoma neural progenitor cells (PC12) and murine embryo fibroblasts (NIH/3T3).
Electropolymerizing pyrrole within the hydrogel introduced high electrical conductivity and switchability; however, this also increased the elastic modulus and altered the water content, both factors known to influence cellular interactions. The degree of hydration and free versus bound water distribution was also different in the new ECH which affects protein adsorption and cell adhesion. Considerable growth and proliferation of PC12 was seen on Au*|gel-PPy scaffold when compared to controls of Au*, Au*|Gel and Au*|PPy scaffolds and this scaled with the electropolymerization charge density.
PAn-Cl/CHI composites (0-100 wt%) were slow dried or frozen and lyophilized to produce dense or highly porous membranes respectively. SEM showed that composition greatly affected 3-D foam formation with the 50/50 wt% producing the most reticulated structure. Electrical impedance spectroscopy (EIS) with equivalent circuit modeling in air, DI-water and in physiological buffers (PBS, HEPES) allowed the same modified Randles equivalent circuit of low chi-square values (<0.05) to be used to describe all compositions and resulting morphologies. However, membranes changed from being highly resistive (100 wt% PAn-Cl, as cast) to being highly capacitive (50/50 wt% PAn-Cl/CHI, lyophilized) and displayed a percolation threshold at ca. 30wt%. Cyclic voltammetry in 0.1 M HCl from -0.2 V to 1.0 V showed that all compositions and morphologies were electroactive. Enhanced cell proliferation accompanied increased electroactivity. However, factor-less differentiation of PC12 was not observed.
10:00 AM - *BM04.07.06
Engineering and Exploring the Cell-Material Interface for Regenerative Engineering
Molly Stevens 1 Show Abstract
1 Department of Materials and Department of Bioengineering, Imperial College London, London United Kingdom
An important aim of regenerative medicine is to restore tissue function with implantable, laboratory-grown constructs that contain tissue-specific cells that replicate the function of their counterparts in the healthy native tissue (1,2). In this talk I will describe our recent work in the development of implantable scaffolds and in the development of state of the art technologies for monitoring and elucidating the cell-material interface (3,4,5).
1. E.Pashuck et al. (2012) Designing Regenerative Biomaterial Therapies for the Clinic, Science Translational Medicine.
2. E. Place et al. (2009) Complexity in biomaterials for tissue engineering, Nature Materials.
3. S.Bertazzo et al. (2013) Nanoanalytical electron microscopy reveals fundamental insights into human cardiovascular tissue calcification, Nature Materials.
4. C. Chiappini et al. (2015) Biodegradable silicon nanoneedles delivering nucleic acids intracellularly induce localized in vivo neovascularization, Nature Materials.
5. P. Howes et al (2015) Colloidal nanoparticles as advanced biological sensors. Science.
10:30 AM - BM04.07.07
Porous Piezoelectric Bone Scaffolds with High Aspect Ratio
Gian Nutal Schädli 1 , Robert Baumann 1 , Sotiris Pratsinis 1 Show Abstract
1 , ETH Zurich, Zurich Switzerland
Smart bioactive engineered bone scaffolds are important for the progressively aging world population and for treating bone diseases. Piezoelectricity contributes to the process of bone growth and remodeling . Therefore, composite bone scaffolds containing piezoelectric BaTiO3 is a step towards mimicking natural bone. Current strategies for bone scaffold design pursue polymer-ceramic composites that have similar mineral composition as well as mechanical and structural properties to bone. Often hydroxyapatite (HA) - poly(lactic-co-glycolic acid) (PLGA) composites are used to mimic natural bone. Most commonly, these composites (scaffolds) are fabricated using solvent casting particulate leaching (SCPL), a widely applied method that, however, is limited to thin scaffold structures with low aspect ratio (length:diameter) .
Here, BaTiO3 particles 50 nm in average diameter with specific surface area (SSA) of 20 m2/g  are used as piezoelectric filler and compared to nano-sized HA particles (SSA = 44 m2/g) for fabricating reinforced piezoelectric and non-piezoelectric PLGA scaffolds, respectively. For the first time to our knowledge, cylindrical HA - PLGA scaffolds are produced with a high 2:1 aspect ratio by pressurized SCPL while maintaining high mechanical strength, overcoming previous limitations of SCPL. The mechanical and piezoelectric properties of these scaffolds are analyzed by compressive strain and force measurements as well as a piezometer, respectively. Bone scaffolds made with HA – PLGA at 1:1 weight ratio have a compressive Young’s modulus of 17 MPa that is 6 times larger than those made without applying pressure during casting. Piezoelectric scaffolds made with BaTiO3 – PLGA at the same weight ratio have (because of the lower surface area) a smaller modulus of 10 MPa that is promising for further studying the contribution of piezoelectricity to bone growth.
 Noris-Suárez K, Lira-Olivares J, Ferreira A M, Feijoo J L, Suárez N, Hernández M C and Barrios E 2007 In vitro deposition of hydroxyapatite on cortical bone collagen stimulated by deformation-induced piezoelectricity Biomacromolecules 8 941–8
 Stevens B, Yang Y, Mohandas A, Stucker B and Nguyen K T 2008 A review of materials, fabrication methods, and strategies used to enhance bone regeneration in engineered bone tissues J. Biomed. Mater. Res. Part B Appl. Biomater. 85B 573–82
 Schädli G N, Büchel R and Pratsinis S E 2017 Nanogenerator power output: influence of particle size and crystallinity of BaTiO3 Nanotechnology 28 275705
10:45 AM - BM04.07.08
Comprehensive In Vitro and In Vivo Studies of Novel Melt-Derived Nb-Substituted 45S5 Bioglass Reveal Enhanced Bioactive Properties for Bone Healing
Joao Lopes 1 2 , Celso Bertran 2 , Lucas De Souza 3 , Angelo Camilli 3 Show Abstract
1 School of Chemical Engineering, University of Campinas (UNICAMP), Campinas, Sao Paulo, Brazil, 2 Department of Physical Chemistry, Institute of Chemistry, University of Campinas (UNICAMP), Campinas Brazil, 3 Institute of Biology, University of Campinas, Campinas Brazil
Despite growing evidence of the osteogenic potencial of Nb-substituted silicate glass, its effect over pluripotent cells and bone regeneration has still not been investigated. To determine the cytotoxicity and bioactive properties, such as, osteoinduction, osteoconduction and osteostimulation of this material we carried out in vitro and in vivo experiments. The in vitro approach consisted of treating human embryonic stem cells (hESCs) with the dissolution products of different compositions of Nb-substituted silicate glass derived from 45S5 bioglass. Toward this purpose, two series of bioactive glasses using a SiO2–Na2O–CaO–P2O5-Nb2O5 system were studied. The first series (I) substituted P2O5 with Nb2O5, while the second series (II) substituted SiO2 with Nb2O5; in both series, the Na2O:CaO ratio was fixed. Their ionic leaching profiles were determined by ICP-OES analysis and we verified the effect of these leached ions over the cellular viability and osteogenic differentiation of the hESCs. The solid state 31P MAS NMR spectroscopy was used to investigate the kinetic of calcium phosphate layer formation of two series of bioactive glass before and after soaking in SBF solution. Solid state 31P MAS NMR proved to be very sensitive technique to elucidate the in vitro chemical reactivity and bioactivity of niobium bioactive glasses. 31P MAS NMR showed that glasses containing lower quantities of Nb2O5 exhibited enhanced early glass apatite formation, whereas for glass compositions containing higher amount of Nb2O5 an opposite effect was observed. In the in vivo trial glass rods were implanted into circular defects at rat’s tibias. The area of subperiosteal and trabecular new-formed bone was quantified, as well as the thickness of a bone layer formed onto the material’s surface, by means of histomorphometry. Our results showed that Nb-substituted silicate glasses are not toxic to hESCs. Moreover, adding up to 1.3% of Nb2O5 to 45S5 bioglass increased the release of Si, P, Na and Ca species by the glass, which may have been responsible for the observed enhanced osteogenic and osteostimulative properties. We conclude that due to this greater biological performance Nb-substituted silicate glasses may be an interesting alternative for biomedical applications.
J.H. Lopes, A. Magalhaes, I.O. Mazali, C.A. Bertran, Effect of Niobium Oxide on the Structure and Properties of Melt-Derived Bioactive Glasses, J. Am. Ceram. Soc., 97 (2014) 3843-3852.
11:00 AM - BM04.07.09
Bioactive Antimicrobial and Immunomodulatory Coating
Philippe Lavalle 1 , Angela Mutschler 1 , Lorène Tallet 1 , Cynthia Calligaro 1 , Julien Barthes 1 , Helena Knopf-Marques 1 , Leyla Kocgozlu 1 , Engin Vrana 2 , Pierre Schaaf 1 Show Abstract
1 , INSERM, Strasbourg France, 2 , Protip Medical, Strasbourg France
All implantable biomedical systems face several risks once in contact with the host tissue : i) excessive immune response to the implant (1); ii) development of bacterial biofilms and iii) yeast and fungi infections. A multifunctional surface coating which can address all these issues concomitantly would significantly improve clinical outcomes. We develop here for the first time a coating that address these three issues simultaneously. We hypothesized that polyarginine (PAR), a synthetic highly cationic polypeptide, can act on macrophages to control innate immune response because arginine is an important component of macrophage metabolism. Moreover, PAR is susceptible to act as an antimicrobial agent due to its positive charges. We developed a new polyelectrolyte multilayer films (2) based on PAR and hyaluronic acid (HA). The layer-by-layer PAR/HA films have a strong inhibitory effect on the production of inflammatory cytokines released by human primary macrophages subpopulations (3). This could reduce potential chronic inflammatory reaction following implantation. Next, we show that PAR/HA films were very effective to inhibit Gram- positive and Gram-negative pathogenic bacteria associated with infections of medical devices. We demonstrate that exclusively films constructed with poly(arginine) composed of 30 residues (PAR30) acquire a strong antimicrobial activity (4, 5). The cytocompatibility of the PAR/HA films was assessed with several cell types playing a major role in tissue engineering. This all-in-one system that limits strong inflammation and prevent pathogen's infections on implants constitutes an original strategy easy to scale up.
(1) Kzhyshkowska, J.; Gudima, A.; Riabov, V.; Dollinger,, C.; Lavalle, Ph.; Vrana, N.E. J. Leuk. Biol. 2015, lb.5VMR0415-166R.
(2) Knopf-Marques H., Singh S., Su Su Htwe S. S., Wolfova L., Buffa R., Bacharouche J., Francius G., Voegel J.-C., Schaaf P., Ghaemmaghami A. M., Vrana N. E., Lavalle Ph., Biomacromolecules, 2016, 17, 2189–2198.
(3) Özçelik, H.; Vrana, N.E.; Gudima, A.; Riabov, V.; Gratchev, A.; Haikel, Y.; Metz-Boutigue, M.H.; Carradò, A.; Faerber, J.; Roland, T.; Klüter, H.; Kzhyshkowska, J.; Schaaf, P.; Lavalle, P. Adv. Healthc. Mater. 2015, 4, 2026-36.
(4) Mutschler, A.; Tallet, L.; Rabineau, M.; Dollinger, C.; Metz-Boutigue, M.-H.; Schneider, F.; Senger, B.; Vrana, N. E.; Schaaf, P.; Lavalle, P. Chem. Mater. 2016, 28, 8700-8709.
(5) Mutschler A., Betscha C., Ball V., Senger B., Vrana N. E., Boulmedais F., Schroder A., Schaaf P., Lavalle Ph. "Nature of the polyanion governs the antimicrobial properties of poly(arginine)/polyanion multilayer films", Chem. Matter., 2017, 29, 3195–3201.
11:15 AM - BM04.07.10
Thermally Drawn Porous Constructs for Tissue Engineering
Dena Shahriari 1 , Yoel Fink 1 2 , Polina Anikeeva 1 2 Show Abstract
1 Research Laboratory of Electronics, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States, 2 Department of Materials Science and Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States
When using biomaterials in tissue engineering application, it is essential to create constructs biocompatible to the tissue of interest and to control the porosity of the material. More specifically, it is necessary to control properties such as chemistry (material selection), permeability and mechanical properties. As an example, in the case of nerve repair when neurons need to be guided along the nerve gap using a scaffold, the implant should have mechanical properties such as stiffness and strength optimized and have interconnected porosity. The pore size distribution and percentage need to be optimized so that nutrition, oxygen and waste are permeable while neurons grow on the surface of the material without penetrating the pores to ultimately bridge the nerve gap and reach the distal end of the injury.
We advanced a thermal drawing technique to create porous materials from a variety of polymers at unprecedented large scales (each experiment was done in about 4 hours and produced 100s of meter-long constructs). We mixed a sacrificial crystalline with a thermoplastic, and thermally drew the composite above the glass transition of the polymer but below the crystalline melting temperature to keep the crystalline structure intact during the drawing. By changing the polymer/crystalline percentage, we were able to create polymers with pore sizes at low micrometer scale and porosity percentages of 0-70 vol%. Corresponding to the porosity percentage, the mechanical properties of the materials consistently changed (stiffness was reduced and strength was increased as more porosity was introduced). We created these materials in different shapes such as hollow tubes with circular and square cross-sections to create porous structured biomaterials at large scales. We next investigated the growth of primary neural cells through the porous tubes with 300 µm diameter. We were able to tune neural growth rates based on the polymer chemistry, porosity percentage and materials geometry. The porous tubes are therefore viable options for use in bridging nerve gaps as well as being used in tissue engineering applications that require a structured biomaterial.
11:30 AM - BM04.07.11
Metallic 3D Woven Lattices Coated with Hydroxyapatite as Bio-Scaffolds
Ju Xue 1 , Yunfei Wang 1 , James Guest 2 , Warren Grayson 3 , Shoji Hall 1 , Timothy Weihs 1 Show Abstract
1 Material Science and Engineering, Johns Hopkins University, Baltimore, Maryland, United States, 2 Civil Engineering, Johns Hopkins University, Baltimore, Maryland, United States, 3 Biomedical Engineering, Johns Hopkins University, Baltimore, Maryland, United States
Many researchers are focused on developing the next generation of bio-scaffolds that combine biologically activated coatings with porous scaffolds that possess fluidic permeabilities and mechanical properties that mimic those of native tissues. Here we present studies of 3D lattices that are woven with 304 stainless steel and Ti6Al-4V wires and then are coated uniformly with hydroxyapatite. The architectures of the 3D weaves are designed to optimize a combination of fluidic permeability and mechanical stiffness, and the 100 to 150 micron diameter wires are coated with hydroxyapatite after weaving to improve bioactivity and osteointegration. The hydroxyapatite coatings are electrochemically deposited onto the weaves using a phosphate salt aqueous solution containing and . The coating temperature and the applied voltage were varied to maximize the uniformity and adhesion of the coating. X-ray diffraction (XRD) and energy dispersive spectroscopy (EDS) were employed to characterize the crystal structures and chemical compositions of the coatings, and scanning electron microscopy (SEM) and micro-CT have been used to investigate the morphology and the uniformity of the coatings across the 3mm thick weaves. Permeability of the woven lattices was also measured both before and after deposition. Lastly, we will describe the growth and morphology of the hydroxyapatite coating as it is immersed in a simulated body fluid (SBF) solution, as a function of time, using mass gain, pH variation, and SEM imaging.
11:45 AM - BM04.07.12
Silver Palladium and Silver Platinum Nanoparticles as New Antimicrobial Agents
Aida Lopez Ruiz 1 , Caterina Bartomeu Garcia 1 , Thomas Webster 1 , Nicole Bassous 1 Show Abstract
1 , Northeastern University, Boston, Massachusetts, United States
The increasing use of antibiotics has led to the emergence of antibiotic resistant bacteria. According to the Centers for Disease Control (CDC), in 2017 at least 2 million people will become infected with drug resistant bacteria and at least 23,000 people die each year as a direct result of these infections. The increasing of getting infections in hospitals, in special in immunocompromised patients, as cancer patients by antibiotic resistant bacteria, reduce the options for treatment against bacteria.
In this manner, we have been pioneers in the use of nanoparticles to kill bacteria and treat infections. The antimicrobial activity of nanoparticles is based on their small size and high surface area capable of penetrating biofilms as well as bacteria to influence intracellular mechanisms (such as the function of thiol-containing proteins and reactive oxygen species). The composition chosen for the presently fabricated nanoparticles is an alloy of silver palladium and silver platinum. This composition was selected due to the high antimicrobial effect of silver and for the imaging and anticancer properties that palladium and platinum have, thus, representing a new generation of theranostic nanoparticles for infection treatment and at the same time anticancer treatment.
Results showed a remarkable antimicrobial effect for silver palladium and silver platinum nanoparticles. Antibacterial activity was tested with three different bacteria: S. aureus, P. aeruginosa and E. coli multi drug resistant; determining bacteria colony-forming units and growth-curve assays. For colony-counting at 11h of culture, between 100-200 µg/mL of AgPdNPs decreased the bacteria population by 99%. For the anticancer effect with glioblastoma at 1 and 3 days, between 150-250 µg/mL of AgPdNPs and AgPtNPs decreased the cancer cells population by 90%.
BM04.08: Polymeric Biomaterials for Regenerative Engineering III
Wednesday PM, November 29, 2017
Sheraton, 2nd Floor, Independence West
1:30 PM - BM04.08.01
Dual Sensor for Oxygen and Temperature Sensing and Specific Ion Effects on the Water Solubility of the Dual Sensor
Ziyun Yang 1 , Weizhen Wu 1 , Tingting Pan 1 , Yanqing Tian 1 Show Abstract
1 Materials Science and Engineering, Southern University of Science and Technology, Shenzhen, Guangdong, China
Dissolved oxygen and temperature have marked effect on cell metabolism and respiration. Measurement of these two parameters has caused significant interests for understanding physiological and biological applications. Herein, we will report a random copolymer, poly(MEO2MA-co-OEGMA-co-PtPophrin), consisting of 2-(2-methoxyethoxy) ethyl methacrylate (MEO2MA), oligo(ethylene glycol) methacrylate (OEGMA) for temperature sensing and platinum (II) porphyrin (M1) for oxygen sensing. The lower critical solution temperature (LCST) value of this copolymer was controlled at 37 degrees Celsius by adjusting the mole fractions of MEO2MA and OEGMA. LCST affected the polymer’s aggregation status, which in turn affected the nanostructures, fluorescence intensities, and responses to dissolved oxygen. Thus the polymer was enabled to functionalize as a dual temperature and dissolved oxygen optical sensor. The thermo-response was investigated by measurements of optical transmittance and laser particle size analysis as well as fluorescence intensity change along the temperature change. The oxygen-responsive property was studied under different dissolved oxygen concentrations. The results illustrated that this sensor has high-resolution to temperature, good repeatability, which will act as a potential instrument in biological fields.
Poly(MEO2MA-co-OEGMA-co-PtPophrin) is a macromolecular whose precipitation is sensitive to the concentration of the ions present in solution, especially anions. The effect of anion on the LCST of the sensor was investigated by measurements of optical transmittance along the type or the concentration of anion change. The result showed that anions can lower the LCST of the sensor and the ability of a specific anion to lower the LCST generally followed the Hofmeister series.
1:45 PM - BM04.08.02
3D-Painted Salts—A Highly Tailorable Extension of the “3D-Painting” Process for Fabricating Highly Porous and Structurally Robust Architectures from a Wide Variety of Materials
Adam Jakus 1 , Ramille Shah 1 Show Abstract
1 , Northwestern University, Chicago, Illinois, United States
In this presentation, we will be introducing a new modality to the recently established, material-independent 3D-Painting process that integrates salt-leeching methods to additively manufacture structures with high material porosity without compromising mechanical integrity. “3D-painting” is the collective name recently given to a newly introduced, materials-centric, room-temperature, 3D-printing process developed at Northwestern University. This process has resulted in materials such as the highly osteogenic Hyperelastic “Bone”, neurogenic 3D-Graphene, Organ Specific “Tissue Papers”, and new approaches to 3D-print metals, alloys, conductive and semi-conductive materials, and more. 3D-paints are created by mixing a solvent mixture composed of evaporate, surfactant, and plasticizer with a biomedical elastomer (10-40% by solids volume) and suspended powder (60-90% by solids volume), where the powder used defines the resulting 3D-paint. In the current work, the powder component of the 3D-paint is entirely or partly composed of water soluble salts. Leveraging the ability to create high-particle content 3D-painted architectures with established salt leeching methods, we demonstrate that 3D-painted salt structures can be readily leeched via water washing to create mechanically robust structures with upwards of 95-98% porosity without compromising mechanical properties. We briefly discuss how this process can be utilized to add tailored porosity to 3D-paintable materials such as bioceramics, graphene, metals and alloys, and extracellular matrices (ECMs), for a variety of biological and non-biological applications but primarily focus on the properties of materials and structures 3D-printed from paints containing only salt (CuSO4) powders at 25, 50, and 70 volume% solids loading (remaining solids is medical grade polylactide-co-glycolide; PLGA), that have been leeched prior to use. The material groups retain their as printed architectures, but exhibit distinct mechanical, structural, and biological properties. Microstructurally, the most porous groups resemble decellularized ECM. Elastic moduli range from 5-250 MPa; absorbency ranges from 200-800%; and all materials can be rolled, folded, sutured, cut, etc. In vitro studies on 3D-printed scaffolds with human mesenchymal stem cells over 28 days show that the low porosity group does not permit cell adhesion, viability or proliferation. This is in stark contrast to the high porosity groups that are cell friendly and promote cell adhesion, proliferation, and matrix production. We also present in vivo results from adult rat critical sized cranial defects treated with the highest porosity PLGA material over the course of 8 and 12 weeks. Finally, we demonstrate that the highest porosity PLGA variant derived from this 3D-painted salt system can act as a non-specific-biocative scaffold/carrier for difficult to surgically handle liquids and gels, such as those derived from ECMs or peptide amphiphiles.
2:00 PM - BM04.08.03
Bioactive Glass Containing Silicone Elastomers—Possible Material for Percutaneous Driveline Exit Sites
Nicholas Cohrs 1 , Konstantin Schulz-Schönhagen 1 , Dirk Mohn 1 2 , Wendelin Stark 1 Show Abstract
1 , ETH Zurich, Zurich Switzerland, 2 , University of Zurich, Zurich Switzerland
Due to the limited amount of donor hearts, an increasing number of ventricular assist devices (VADs) are implanted into patients who suffer from heart failure. VADs are small, continuously pumping and electrically driven artificial blood pumps, which support the weakened heart. One major drawback of the therapy is the need for a percutaneous driveline, which permanently penetrates the skin and provides the VAD with the required electricity. This driveline exit site is most susceptible to infection and constitutes as an entry point for germs . For instance, patients implanted the HeartMate II device suffer from a driveline infection rate of 37% per patient-year . A stable and durable connection of the driveline’s material, e.g. silicone, to the skin is desired to prevent pathogens from entering the body. Bioactive glass (BG) is an amorphous material whose property to form a stable bond to bone is well known. However, an increasing number of studies are being published, which examine BG in contact with soft tissue – with promising results . As silicone elastomers are inert in contact with living tissue, BG could be a suiting material to improve the elastomer’s bioactive properties, thus enhancing skin biointegration and giving a more stable driveline exit site. We investigated whether the incorporation of different BG45S5® particles, varying in particle size, into medical grade silicone elastomers could form a bioactive composite . BG45S5® nanoparticles were produced using flame spray synthesis , while two commercially available BG microparticles with primary diameters of 4 µm and 54 µm were purchased. In vitro tests of the BG containing silicone composites at different particle concentrations in simulated body fluid revealed the hydroxyapatite forming ability of the composites, thus proving its bioactivity. Cell proliferation tests with primary human dermal fibroblasts showed that BG significantly improves the cytocompatibility of medical grade silicone elastomers.
The study showed the potential of bioactive glass particles with silicone elastomers for possible driveline exit sites. Cell proliferation measurements were conducted to assess the cytocompatibility of the material, however do not allow to make a statement regarding tissue adhesion of the composite. Further in vivo studies are in process and address biocompatibility and tissue adhesion.
 D. Dean, F. Kallel, G.A. Ewald, et al., J Heart Lung Transplant 34(6) (2015) 781-789.
 S.P. McCandless, I.D. Ledford, N.O. Mason, et al., Cardiovasc Pathol 24 (2015) 71-75.
 V. Miguez-Pacheco, L.L. Hench, A.R. Boccaccini, Acta Biomater 13 (2015) 1-15.
 N.H. Cohrs, K. Schulz-Schönhagen, F. Jenny, et al., J Mater Sci 52 (2017) 9023-9038.
 T.J. Brunner, R.N. Grass, W.J. Stark, Chem Commun 13 (2006) 1384-1386.
2:15 PM - BM04.08.04
Phthalocyanine-Based Thin-Film Coatings by Layer-by-Layer Assembly for In Vitro Photodynamic Therapy
Yonca Belce 1 , Fevzi Cebeci 1 2 Show Abstract
1 Faculty of Engineering & Natural Sciences, Sabanci University, Istanbul, Tuzla, Turkey, 2 Nanotechnology Research & Application Center, Sabanci University, Istanbul, Tuzla, Turkey
Photodynamic therapy (PDT) is one of the promising treatment methods for cancer treatment since 1900. Once a photosensitizer (PS) is activated by light, the generated singlet oxygen leads to cell death. Photosensitizer substituted by hydrophobic phthalocyanine needs to be carried by nanoparticles or polymer micelles to the target tissue. However, as the photosensitizer is activated by near-infrared (700-2500nm) or visible (400-700nm) light to initiate the photodynamic reaction only some of the encapsulated photosensitizers can arrive to the target tissue. Unfortunately, light at that wavelength cannot penetrate deeper than 1-3mm through skin surface. In addition, due to the short life time of reactive singlet oxygen PDT is commonly recommended for tumors on skin or just under the skin. Therefore, there is a need for improvement on the direct efficiency of photosensitizers on tumor cells. Stable cofacial complexes formed by porphyrins and phthalocyanines are also studied to increase efficacy.
In this study, it is aimed to fabricate thin film coatings containing photosensitizers with controlled release mechanism and with known material amount onto the materials that are commonly used in medical treatment. Thin film structures are assembled by dip-spin and spray layer-by-layer (LbL) techniques. Photosensitizer amount in the film system is quantitatively determined once it is coated on absorbable, light transmittable and porous gelatin sponge or on micro needle bandage structures. Depending on the absorption spectrum of PS, thin film structures are exposed to photodynamic reaction. Hydrophobic PS is encapsulated by amphiphilic polymers for homogenous distribution in water. Hence, it is possible to obtain tetra layer thin film structure with photosensitizers by LbL method. Oppositely charged polyelectrolytes are placed to the outer layer so that it can make a strong interaction (hydrogen bonding, ionic bonding) with the active material in photodynamic therapy and it will also protect the system from external factors. QCM-D measurements are performed to determine the quantity of photosensitizers coated on surface and for further characterizations of the coatings profilometer and ellipsometer are used. The distribution of photosensitizer and the surface morphology of the film are investigated by scanning and transmittance electron microscopy. Mechanical and optical properties of the films are also analyzed.
3:30 PM - BM04.08.05
Designing 3D Cell Niches Exploiting Peptide Self-Assembly for Regenerative Engineering
Deepak Kumar 1 , Victoria Workman 2 , Felicity Rose 3 , Alberto Saiani 2 , Julie Gough 1 Show Abstract
1 School of Materials, University of Manchester, Manchester United Kingdom, 2 School of Materials and Manchester Institute of Biotechnology, University of Manchester, Manchester United Kingdom, 3 Centre of Biomolecular Sciences, School of Pharmacy, University of Manchester, Manchester United Kingdom
We have developed a platform for the design of 3D scaffolds, i.e.: hydrogels, exploiting so-called β-sheet forming peptides whose properties can be tailored to accommodate different cells’ needs. We have used these novel materials for the culture of a variety of cells including chondrocytes, osteoblasts, neurons, Schwann cells as well as embryonic and mesenchyme stem cells. Our results clearly demonstrate that our peptides offer great promise for the design of specific cell niches due to their low immunogenicity and the ability we have to control and tailor their properties.
Recently [under review in Avd Func Mat] we investigate the possibility of using these materials to devise a strategy to prevent formation of oesophageal strictures after endoscopic mucosal resection (EMR). EMR is a surgical treatment used in Barrett’s oesophagus to remove pre-cancerous tissue. Formation of oesophageal strictures poses a major problem to patients. In this study, we evaluate the response of oesophageal epithelial cells and stromal fibroblasts to several β-sheet forming peptide hydrogels, each with distinct sequence, charge and mechanical properties, as a potential material to be delivered after EMR to support normal healing and tissue regeneration and prevent stricture formation.
A panel of peptide hydrogels (composed of 8-10 amino acids) were investigated to assess their ability to support oesophageal cell types. Oesophageal stromal fibroblasts (rOSFs) were incorporated into peptide hydrogels, whereas oesophageal epithelial cells (mOECs) were seeded on the surface, mimicking the in vivo arrangement of these cell types. mOEC viability, proliferation and migration across the hydrogel surface were evaluated; whereas rOSF viability, morphology and homogenous distribution within the hydrogels were assessed.
Live/dead analysis revealed that the soft & positively charged hydrogels were optimal for supporting mOEC viability, expansion, migration and the formation of an epithelial sheet. Cultured mOECs also retained similar ZO-1 and cytokeratin expression relative to controls. On the other hand stiffer & positively charged hydrogels were optimal for supporting rOSFs and their homogenous incorporation after day 7 and 14 of culture. Culture of a composite of both cell types in their respective optimal hydrogels revealed successful recapitulation of the in vivo oesophageal submucosal layer with complete epithelialisation and positive detection of involucrin and AE1/AE3 after 7 and 14 days of co-culture.
This study has identified the optimal peptide hydrogels to support oesophageal tissue specific cell activity and function with adequate mechanical properties to match native tissue mechanical properties. These hydrogels also promoted the formation of an uninterrupted epithelial sheet. This provides exciting opportunities to permit their use as a minimally invasive endoscopic therapy to prevent oesophageal strictures.
3:45 PM - BM04.08.06
Handheld In Situ Bioprinter
Navid Hakimi 1 , Richard Cheng 1 , Nazihah Bakhtyar 2 , Marc Jeschke 2 1 , Saeid Amini-Nik 2 1 , Axel Guenther 1 Show Abstract
1 , University of Toronto, Toronto, Ontario, Canada, 2 , Sunnybrook Research Institute, Toronto, Ontario, Canada
3D bioprinting strategies aim at reconstituting structural elements of native tissues by controlling the position of different cell types and extracellular matrix components. The provided microenvironment and spatial organization influence cell migration, elongation, clustering, proliferation, differentiation, and function. Current bioprinting platforms offer promising in-vitro results but are not yet compatible with clinically relevant settings. Higher print rates, reduced preparation and wait times, and compact solutions for on-site deposition or transfer of organ-scale printed tissues are required to ultimately treat patients with acute and complex wounds that are amongst the most impactful clinical and economical challenges. We present a handheld in-situ bioprinter that overcomes these limitations by in-situ formation of wound-adhesive skin substitutes. The handheld printer’s microfluidic cartridge deposits epidermal and dermal cells within biopolymeric precursor solutions in a bi-layered fashion. Upon enzymatic or ionic crosslinking, architected, cell-laden skin substitutes are obtained and characterized in-vitro. A porcine excisional wound model serves as a case study to demonstrate feasibility for in-vivo application. Widely used and clinically approved biomaterials (collagen type 1, fibrinogen, hyaluronic acid, and alginate) were selected in this work. Skin substitutes were formed in a one-step manner that does not require multi-axis printhead translation or manipulation of printed constructs. We expect the handheld bioprinter platform to enable the in situ delivery of a wide range of architected biomaterials, wound healing adhesives and cells.
4:00 PM - BM04.08.07
3D Bioprinting of Liver-Mimetic Construct with Alginate-Cellulose Nanocrystal Bioink
Yun Wu 1 , Shirley Tang 1 Show Abstract
1 , University of Waterloo, Waterloo, Ontario, Canada
Bioprinting offers the potential to create three dimensional (3D) artificial tissues with hierarchical structures reproducibly and with ease. To realize such potential, though, one must design proper bioink formulations to address the challenges in both printability and post-print biocompatibility. Rheological properties of the bioink (pre-gel), gelation mechanism, mechanical and chemical properties of the gel matrix (post-print) are important factors to consider while designing a bioink. Here, we report our work in developing a bioink composed of cells suspended in a buffered solution of alginate and cellulose nanocrystals (CNC). The alginate-CNC hybrid ink exhibits outstanding shear thinning behavior with a fast crosslinking mechanism. The pore size and porosity of the hybrid hydrogel matrix, after printing and gelation, are suitable for making 3D cell-laden tissue constructs, as confirmed by SEM. Bioink printability and pattern fidelity were evaluated by printing a liver-mimetic honeycomb structure, with NIH-3T3 fibroblasts (stromal) surrounding hepatoma cells. Cell viability in the printed constructs and a reference sample (molded, no printing) was recorded during a 7-day culture. There was no significant difference in cell viability between the molded and the bioprinted constructs, revealing that the extrusion printing process, through a nozzle with a 100 μm inner diameter, has no side effect on cell viability, due to the strong shear thinning property of the bioink composed of 2% (w/v) alginate with 4% (w/v) CNC. Overall, our alginate-CNC hybrid bioink can be used for 3D printing of artificial tissues with micro-structures made of different cell types.
4:30 PM - BM04.08.09
Cytocompatible and Reversible Phospholipid Polymer Hydrogels for Controlling Stem Cell Functions
Kazuhiko Ishihara 1 , Haruka Oda 1 , Tomohiro Konno 1 Show Abstract
1 , University of Tokyo, Tokyo Japan
As the field of regenerative medicine starts to play an important role in the new generation of bioengineering, the cells are starting to be treated as one of the materials to be controlled and optimized. The cells performance is affected by its microenvironment; not only by its chemical signals but also from its physical cues. We aimed to control cellular function through the physical cues from its environment. The stiffness of the cellular environment were in the range of 0.1 to 10 kPa, which is most likely to be seen in the living tissue. Cytocompatible and property tunable phospholipid polymer hydrogels were prepared as cellular environment. 2-Methacryloloxyethyl phosphorylcholine (MPC) polymers show no interaction with the proteins and cells. By constructing the cellular microenvironment with MPC polymers, it made it possible to discuss the physical cues of environment to the encapsulated cells with negligible effect on chemical cues such as nutrition or signal proteins. We have reported that the initial storage modulus of the MPC polymer hydrogel controls proliferation of the encapsulated stem cells. Here, we report the proliferation control by changing the storage modulus of the hydrogel during the encapsulation of stem cells.
Poly(MPC-co-n-butyl methacrylate(BMA)-co-p-vinylphenylboronic acid (VPBA)) (PMBV) was synthesized with radical polymerization method. When the PMBV was mixed with poly(vinyl alcohol)(PVA), hydrogel was formed spontaneously with in 10 sec without any chemical treatment. During this process, cells could be encapsulated. Proliferation and differentiation of mouse mesenchymal stem cell, C3H10T1/2 were examined with relation of stiffness of the hydrogel.
The PMBV in cell culture medium (DMEM) solution was mixed with PVA-DMEM solution in 60/40 vol. ratio to give PMBV/PVA hydrogel with storage modulus of 1.1 kPa. This storage modulus is known to suppress encapsulated cell proliferation. The PMBV/PVA hydrogel was then swollen to enlarge its volume to 1.5-hold. The storage modulus of the hydrogel decreased to 0.70 kPa and stable 3D polymer network was confirmed. When the storage modulus is between 0.50 and 1.0 kPa, the encapsulated cells are known to show slow proliferation. The C3H10T1/2 cells did not proliferate in the 1.1 kPa PMBV/PVA hydrogel, and the cell cycle was converged into G1 phase. The fraction was up to 95%. As the storage modulus was lowered to 0.70 kPa, the cells restarted its proliferation and cell cycle restarted as well. This is a novel way to control large amount of cell proliferation. This leads to high sensitivity to differentiation inducing active protein BMP-2, and higher efficiency in differentiation of the cells to bone cells was also confirmed, that is, 4.2-times higher by a conventional cell culturing procedure.
4:45 PM - BM04.08.10
TAT-Functionalized Fusogenic Liposomes for the Treatment of Bacterial Meningitis
Caterina Bartomeu Garcia 1 , Di Shi 1 , Thomas Webster 1 Show Abstract
1 , Northeastern University, Boston, Massachusetts, United States
Brain inflammatory diseases such as bacterial meningitis have recently become a global concern in clinical care due to the recent emergence of antibiotic-resistant bacteria, making it increasingly difficult to treat these infections. The overuse of broad-spectrum antibiotics in clinical care is prevalent and often ineffective, leading to increase drug resistance in bacteria and poorer patient care outcomes.
To address this need, we report the use of fusogenic liposomes to deliver targeted antibiotics at the site of infection, notably inside the bacterial cell using cell penetrating peptides (TAT 47-57). Liposomes have recently been demonstrated as effective drug carriers, though combining this approach with TAT peptides to deliver drugs inside bacteria as a way to combat bacterial meningitis which has yet to be fully explored and would be a novel means of treatment. To this end, we investigated the effect of TAT-functionalized liposomes to inhibit the growth of bacteria commonly associated with bacterial meningitis, including Streptococcus pneumoniae, methicillin-resistant Staphylococcus aureus (MRSA), and Escherichia coli.
Liposomes were prepared by lipid film rehydration, functionalized with TAT (47-57) using wet chemistry. Liposomes were loaded with one of three commonly used antibiotics to treat these bacterial infections, including vancomycin, methicillin, and ampicillin. The inhibitory effect on bacterial growth of the liposomes was determined using bacterial growth curves and determining bacteria colony forming units, and this system was compared against non-TAT-functionalized liposomes and against free antibiotic.
The results from this study demonstrated excellent growth inhibitory effects of TAT-functionalized liposomes loaded with methicillin for treating MRSA, with a reported minimum inhibitory concentration (MIC) of 1.7 µg/mL, well below the needed dosage for free antibiotic (5 µg/mL). A similar decrease in bacteria population was also demonstrated using TAT-functionalized liposomes loaded with vancomycin to treat S. pneumoniae (MIC of 1 µg/mL). The TAT-functionalized liposomes presented the highest antibacterial activity in all assays performed, followed by non-functionalized liposomes, and lastly free antibiotics. Cytotoxicity assays with endothelial cells and astrocytes demonstrated a notable increase in percent cell viability using functionalized and non-functionalized liposomes in comparison with free antibiotics. Results obtained with a methicillin concentration of 5 µg/mL showed only 20% cell viability for the free antibiotic, while for liposomes it was significantly higher at 90%.
All results obtained for TAT-functionalized liposomes, including their low MIC with respect to the use of free antibiotics plus their remarkable high percentage of cell viability with astrocytes and endothelial cells, provides evidence that this liposomal system can be a safe, alternative means for treating bacterial meningitis.
BM04.09: Poster Session III: Biomaterials for Regenerative Medicine
Wednesday PM, November 29, 2017
Hynes, Level 1, Hall B
8:00 PM - BM04.09.01
Tunable Nanostructured Multilayer Assembly Using Silica Forming Engineered Mussel Glue for Accelerating Bone Growth of Titanium Implants
Chang Sup Kim 1 , Yun Kee Jo 2 , Hyung Joon Cha 2 Show Abstract
1 , Yeungnam University, Gyeongsan Korea (the Republic of), 2 , Pohang University of Science and Technology, Pohang Korea (the Republic of)
Silica nanoparticles (SiNPs) have been utilized to construct bioactive nanostructures comprising surface topographic features and bioactivity that enhance the activity of bone cells onto titanium-based implants. However, there have been no previous attempts to create microrough surfaces based on SiNP nanostructures even though microroughness is established as a characteristic that provides beneficial effects in improving the biomechanical interlocking of titanium implants. Herein, we propose a protein-based SiNP coating as an osteopromotive surface functionalization approach to create microroughness on titanium implant surfaces. A bioengineered recombinant mussel adhesive protein fused with a silica-precipitating R5 peptide (R5-MAP) enables to directly control the microroughness of the surface through the multilayer assembly of SiNP nanostructures under mild conditions. The assembled SiNP nanostructure significantly enhances the in vitro osteogenic cellular behaviors of preosteoblasts in a roughness-dependent manner and promotes the in vivo bone tissue formation on a titanium implant within a calvarial defect site. Thus, the R5-MAP-based SiNP nanostructure assembly could be practically applied to accelerate bone tissue growth to improve the stability and prolong the lifetime of medical implantable devices.
8:00 PM - BM04.09.02
Tough Double Network Hydrogel with High Bioactivity Promotes Bone Formation
Junchen Liao 1 , Dingshan Liang 1 , Chuanxin Zhong 2 , Shiyuan Liu 1 , Fuzeng Ren 1 , Ju Fang 1 , Bi Wang 1 Show Abstract
1 Material Science and Engineering, Southern University of Science and Technology, Shenzhen, Guangdong, China, 2 Biomedical Engineering, Southern University of Science and Technology, Shenzhen China
Hydrogels can mimic critical properties of the native extra-cellular environment (ECM). Their highly porous structure and water retention properties allow hydrogels to be applied as one of the best material for cell culturing and tissue engineering. However, the potential for traditional hydrogels were limited in repairing hard tissues due to the low mechanical behavior and bioactivity. Here, a novel double-network (DN) hydrogel, was prepared based on biocompatible Polyethylene glycol (PEG) and chitosan extracted from the natural chitin. The tough PEG network was constructed by thiol-ene crosslinking between 4-armed PEG and linear PEG monomers, forming the first network of this DN hydrogel. Chitosan was introduced as a second cross-linked network for its high bioactivity and antimicrobial properties. Then the DN hydrogel was nanoreinforced by biphasic calcium phosphate (BCP/a mix of hydroxyapatite and β-TCP). Scanning electron microscopy (SEM) results revealed that the multiple porous network structures of this hydrogel composites and the combined BCP were adhered tightly due to the porous and crosslinking structure. This DN hydrogel exhibits excellent mechanical properties compared to the traditional hydrogels. The in vitro and in vivo analysis confirm that the resulted hydrogel highly promotes the attachment and proliferation of preosteoblast as well as accelerating the bone formation rate of rabbits. In addition, the whole processes of synthesis were carried out in aqueous medium without any toxic chemical crosslinkers or catalysts. Therefore, this study provides a green approach to obtain hydrogel composites with strong mechanical properties and good bioactivities for hard-tissue reparation.
This work is financially supported by Science and Technology Innovation Foundation for the Undergraduates of Guangdong province(2017S09).
 Mehrali M, Thakur A, Pennisi C P, et al. Nanoreinforced Hydrogels for Tissue Engineering: Biomaterials that are Compatible with Load-Bearing and Electroactive Tissues[J]. Advanced Materials, 2016, 29.
8:00 PM - BM04.09.04
Silicon Nanowire Scaffold for Optical Pacing of Cardiomyocytes
Kelliann Koehler 1 , Ramya Parameswaran 1 , Michael Burke 1 , Bozhi Tian 1 Show Abstract
1 , University of Chicago, Chicago, Illinois, United States
Developments in cardiac tissue engineering have resulted in a wide range of both synthetic and biological polymer based scaffold materials with mechanical and biological cues for cardiomyocyte viability. These materials offer advancements over traditional methods for treatment of heart damage. Despite advances, most synthetic scaffolds materials serve as passive insulating supports for cardiac tissue. Here we report a silicon nanowire based scaffold with the potential for localized optical, wireless, stimulation of cellular components for regeneration of natural cardiac activity. Silicon is an optically responsive semiconductor, the silicon nanowires can function as a photothermal or photovoltaic stimulation platform to wirelessly direct beating of cells interfaced with the scaffold. Recently, our lab has demonstrated that silicon nanostructures can provide localized optical control of cellular membrane potential in excitable cells, such as neurons, through both photothermal and photoelectric stimulations. For this work, we applied this semiconductor functionality to pace primary neonatal rat cardiomyocytes. Cells were seeded into the scaffold and were optically stimulated at varying frequencies to train cells to beat at that frequency. Fluo-4 was used to monitor calcium transients to determine trained beating frequency. Immunofluorescent staining with connexin 43 and cardiac troponin I antibodies verified that cells were viable, aligned, and were electrically connected to neighboring cells via gap junctions. After optical pacing was demonstrated with isolated cardiomyocytes, we applied this platform to pace an isolated perfused heart via the Langendorff model. These findings demonstrate a significant advancement towards a novel platform towards wireless, active, synthetic tissue constructs for treatment of cardiac tissue damage.
8:00 PM - BM04.09.05
3D Printing Graphene Based Biomaterials for Regenerative Medicine Applications
Priscilla Stachera 1 , Olga Tsigkou 1 , Brian Derby 1 , Suelen Barg 1 Show Abstract
1 , University of Manchester, Manchester United Kingdom
The prevalence of peripheral nerve injuries accounts for over 600,000 people affected in the US and Europe on an annual basis. Peripheral nerves comprise a bundle of nerves that runs down carrying signals, back and forth, between the body and the brain. Once a nervous signal is not able to travel past an injured area of the body, movements from bellow of this area are lost. Even with the possibility of nerve suturing allowing peripheral nervous system self-regeneration, a total recovery of the lesion cannot be obtained. In order to persuade enough axons to regenerate across the lesion to bring back substantial neurological function it is in most cases necessary to construct a bridge over which axons can grow and signal can travel.[2,3] It is proven that materials that signal can be sufficient to allow nervous signal to travel, with previous studies showing promising results with the use of different metallic materials and carbon nanotubes. However, barriers have appeared for such studies continuity and evolution. In that matter, once voltage stimulus can induce axonal regrowth, graphene, being an atomically thin nanomaterial with exceptional electrical properties, appears as an alternative to open up oportunities in the field.[4,5] However, its practical utilization will depend on the ability to integrate its 2D sheets into 3D structures of practical dimensions, while controlling structural features at multiple length scales. Starting with the design and characterization of a graphene based bio-ink (graphene/F127-DA), done through AFM, SEM and FTIR analysis, with precise and optimized rheological properties, this study focus on developing scientific and technological capabilities for 3D printing bridges. It concerns also the design of hierarchical and complex structures in different scales (concerning porosity in different dimensions) and their manufacturing through the use of an extrusion based printer, while constantly evaluating the structures-cells interactions and behavior through in vitro analysis and through their mechanical and electrical responses.
 P.N. Mohanna, R.C. Young, M. Wiberg, G. Terenghi, A composite pol-hydroxybutyrate-glial growth factor conduit for long nerve gap repairs, J. Anat. 203 (2003) 553–565.
 G. Stoll, S. Jander, R.R. Myers, Degeneration and regeneration of the peripheral nervous system: From Augustus Waller’s observations to neuroinflammation, J. Peripher. Nerv. Syst. 7 (2002) 13–27.
 L.A. Pfister, M. Papaloïzos, H.P. Merkle, B. Gander, Hydrogel nerve conduits produced from alginate/chitosan complexes, J. Biomed. Mater. Res. - Part A. (2006) 932–937.
 C. Gardin, A. Piattelli, B. Zavan, Graphene in Regenerative Medicine: Focus on Stem Cells and Neuronal Differentiation, Trends Biotechnol.(2016) 1–3.
 T. Sedaghati, S.Y. Yang, A. Mosahebi, M.S. Alavijeh, A.M. Seifalian, Nerve regeneration with aid of nanotechnology and cellular engineering, Biotechnol. Appl. Biochem. 58 (2011) 288–300.
8:00 PM - BM04.09.06
Fibrillar Polycaprolactone/Gelatin Homogeneous Composite Scaffold for Skin Tissue Engineering Applications
Gina Prado-Prone 1 2 , Masoomeh Bazzar 3 , Phaedra Silva-Bermudez 2 , M.L. Focarete 3 , Clemente Ibarra 2 , Cristina Velasquillo-Martinez 2 Show Abstract
1 , Universidad Nacional Autonoma de Mexico, Mexico city Mexico, 2 , National Institute of Rehabilitation (INR), Mexico city Mexico, 3 Department of Chemistry, University of Bologna (Unibo), Bologna Italy
Major skin injuries are commonly treated by using cadaver or animal skin grafts; however, these therapies are limited by immune rejection and insufficient skin sources. Tissue engineering (TE) provides a new strategy for treating extensive skin wounds by means of skin substitutes developed from biodegradable scaffolds pre-seeded with dermal cells (fibroblasts) to populate the wound after its cutaneous application aiming to promote and accelerate the wound healing while reducing immune rejection, scar formation, and increasing availability. An ideal scaffold should mimic the natural extracellular matrix (ECM) as much as possible, promoting cell adhesion and proliferation while providing mechanical integrity to easily transfer the engineering graft to the wound. Electrospinning technique (ET) enables the production of continuous polymeric fibers ranging from nanometers to submicrons resembling the ECM morphology. In this work, electrospun fibers of a mixture of polycaprolactone (PCL) and Gelatin (Gel) were fabricated combining the properties characteristics of both polymers. PCL is a biocompatible synthetic polymer with well structural stability but the lack of functional groups (hydrophobic surface) and neutral charge (low bioactivity) limit its use as cell scaffold. Gel is a natural biocompatible polymer derived from collagen hydrolysis possessing functional groups present in ECM leading to a low immunogenicity, high hydrophilicity and a better environment for cell adhesion and proliferation. However, the physicochemical differences of both polymers often generate poorly blended immiscible fibers resulting in weak molecular interactions. To overcome this limitation, electrospun PCL/Gel fibers were fabricated using acetic acid (AcAc) as the solvent which is potentially less toxic that solvents commonly used to obtain PCL-based fibers and its high acidity can protonate amino and carboxyl groups of Gel molecules becoming positively charged and stretching its molecular chains easily penetrating in PCL chains, forming a miscible blend. Electrospun PCL/Gel mats were studied in their morphology, chemical composition, water wettability, crystalline structure, mechanical properties and thermal degradation behavior by means of scanning electron microscopy, Fourier transform infrared spectroscopy, energy dispersive X-ray spectroscopy, X-ray diffraction, water contact angle, stress-strain curves and thermogravimetric and differential scanning calorimetric analysis. Furthermore, fibroblasts viability and adhesion was evaluated using calcein/ethidium-homodimer and MTT colorimetric analysis. Preliminary results showed that electrospun PCL/Gel fibers using AcAc as the solvent, produced fibrillar homogenous composites with suitable mechanical properties for medical handling. Moreover, electrospun PCL/Gel fibers showed high cell viability and adhesion on their surface indicating their potential to be use as skin scaffold for TE applications.
8:00 PM - BM04.09.07
Smart Phenytoin Loaded Nanofibrous Wound Dressing for Wound Healing and Detection of Healing Progression
Islam Khalil 1 2 , Isra Ali 2 , Ibrahim El-Shebiny 2 Show Abstract
1 Department of Pharmaceutics and Industrial Pharmacy, Misr University for Science and Technology, 6th of October City,, Giza Governorate, Egypt, 2 Nanomaterials Laboratory, Center for Material Science, Zewail City of Science and Technology, 6th of October City,, Giza Governorate, Egypt
Skin, the soft organ that protects the human internal body from surrounding environment, could be lost due to accidental injuries and burns. Skin tissues could heal naturally after injuries and burns, however normal reorganization of new tissues still represent a challenge especially in complicated wounds such as the diabetic ones. Diabetic wounds possess high alkalinity that obstructs wound healing rapidly. Topical application of phenytoin shows promising wound healing activity. However, Phenytoin is available in the market in a spray form which is not convenient for the patient to get his/her dose accurately. Therefore, we propose to fabricate a smart responsive highly porous electrospun nanofibrous matrix loaded with the appropriate dose for wound healing to act as: (a) wound dressing, (b) scaffolding material for regeneration of damaged skin and (c) sensor for detecting the progress of healing underneath the dressing.
The wound dressing would be fabricated in the form of a three-layer sandwich structure, where each layer has a specific role. First, a fabricated thin layer of electrospun carbapol-based nanofibers would be in direct contact with the diabetic wound. This layer is highly dissolvable, with adhesive property to wound and neutralize wound pH in order to facilitate wound healing. The second layer is composed of Phenytoin-loaded PLLA based nanofibers that could deliver the required dose in a well-controlled and sustained manner. Finally, a sensor layer would be added containing a humidity sensor dye that could respond to change in humidity during wound healing through changing its color. This would help the patient to monitor healing progression and remove the dressing when healing is complete. The three layers were prepared successfully through electrospinning technique after tailoring different parameters to produce highly porous wound dressing mats in order to facilitate oxygen and nutrient diffusion for cell. Scanning electron microscope revealed that diameter of carbapol based nanofibers and PLLA based nanofibers were around 124 ± 19 nm and 267 ± 37 nm respectively. FTIR, DSC and TGA instruments were used to confirm the chemical and thermal stability of the fabricated nanofibers. Physicochemical characterization showed that PLLA based nanofibers showed swelling maximum after 6 hours, where their initial weight increased by 50-60%. In addition, biodegedability test showed that the nanofibers were capable of losing more than 25% of their initial weights in 14 days. Finally in vitro drug release profile showed that the nanofibers could deliver Phenytoin in a well-controlled manner and prolonged period along more than 10 days. Hence, the fabricated system could be a smart wound dressing material for controlled treatment of wounds as well as detecting the healing progress through monitoring loss of humidity upon wound closure.
8:00 PM - BM04.09.08
Is Oxygen a Requirement for Mesenchymal Stem Cell Function? Implications for Scaffold Design
Fiona Lau 1 , Nicoletta Eliopoulos 2 1 , Jake Barralet 1 Show Abstract
1 , McGill University, Montreal, Quebec, Canada, 2 , Lady Davis Institute, Montreal, Quebec, Canada
INTRODUCTION: A key limitation for the development of bulk tissue engineered scaffolds is the inability to deliver oxygen to cells found further within the scaffolds – which can result in hypoxia, and tissue necrosis. Benefits of cell growth without the presence of vasculature will reduce chances of hypoxia and tissue necrosis within these regions. Mesenchymal stem cells (MSCs), found in various tissues such as bone marrow and adipose tissues have various regenerative medicine applications due to their innate ability to differentiate into various cell types, and most importantly due to their paracrine actions. MSCs are plentiful and accessible in humans of all ages, can be obtained with minimal morbidity, easily handled, expanded and gene-enhanced in vitro. Therefore, these cells are desired cellular vehicles for the continuous delivery of beneficial gene products. We hypothesize that in the absence of oxygen, MSCs will alter their metabolic pathway to survive without oxygen. To determine if absence of oxygen in culture media affects cell viability and protein production, we cultured unmodified and Erythropoietin (Epo) gene-modified MSCs in the absence of oxygen (anoxia) and measured viability and Epo production.
METHODS: 1x104 unmodified and Epo-gene modified C57Bl/6 mouse bone marrow-derived MSCs were cultured in 35mm tissue culture plates in 2mL α-MEM culture media (supplemented with 1% Penicillin/Streptomycin, 10% Fetal Bovine Serum and 1% L-Glutamine) for a period of 7 days in normoxic (21% oxygen) and anoxic (<1% oxygen) conditions. Anoxia was created by first deoxygenating media, placing the tissue culture plates inside glass jars and flushing with a mixture of carbon dioxide and nitrogen for a total of 10 minutes per sample and then sealing. The conditioned media of each plate were collected and used to measure Epo levels by ELISA (R&D Systems). The cells from each plate were trypsinized and cell viabilities determined using Trypan Blue.
RESULTS (preliminary): Anoxia showed significant decrease in the total cell number of unmodified MSCs compared to normoxic control (P<0.01), while no significant decrease was found for Epo-modified MSCs. However, within anoxic conditions, unmodified MSCs are significantly reduced compared to Epo-modified MSCs (P<0.01), whereas compared to normoxic conditions no difference can be found. Contrary to expectation, Epo production was not eliminated by absence of oxygen, however no significant difference was detected.
CONCLUSION: Our results indicate that MSCs could survive for up to 7 days in the absence of oxygen. Epo production is unaffected by the absence of oxygen when compared to normoxic controls. This indicates that gene modified MSCs could sustain protein production in anoxia. Future directions will include experimental studies on the effects of nutrient (mainly glucose) on Epo production to further test Epo gene-modified MSCs.
8:00 PM - BM04.09.09
Highly Efficient Targeting of the Pulmonary Microvasculature through Bio-Inspired Functionalization of Low Molecular Weight Polyethylenimine
Andrew Dunn 1 , Donglu Shi 1 Show Abstract
1 , University of Cincinnati, Cincinnati, Ohio, United States
The pulmonary microvasulature plays a key role in not just gas exchange but also in clinically relevant pulmonary vascular diseases such as pulmonary hypertension and pulmonary fibrosis, which may occur at all stages of human life. Efficient targeting of the pulmonary microvascularture is therefore needed for effective therapeutic treatment. Nucleic acid delivery in-vivo provides a powerful tool for therapeutic delivery of small interference or small hairpin ribonucleic acids (siRNA and shRNA respectively), complementary deoxyribonucleic acids (cDNA), or clustered regularly interspaced short palindromic repeats and the associated protein 9 (CRISPER/Cas9) for targeted genomic editing but delivery must be achieved through a tailored excipient. Here we present a bioinspired colloidal system based off of functionalizing polyethylenimine with biological fatty acids using a one-pot, green chemistry synthesis scheme for extremely efficient targeting of the pulmonary endothelial lineage. This colloidal system has achieved greater than 90% targeting in-vivo by whole lung flow cytometry 24 hours post intravenous injection in adult mice while targeting less than 10% of hematopoietic, epithelial, and lineage negative populations. This highly efficient targeting is further retained in neonatal mice with greater than 85% targeting of endothelial cells. Immunofluorescence reveals nanoparticles disseminated throughout the microvasculature, a critical criteria for therapeutic treatment of microvascular diseases.
8:00 PM - BM04.09.10
Development of Aligned Nanofiber Mat Assisted Microfluidic Platform for In Vitro Neuro-Muscular Junction Model with Mature and Functional Muscles
Bumchang Kim 1 , Seongsu Eom 1 , Sang Min Park 1 , Seon Jin Han 1 , Dong Sung Kim 1 Show Abstract
1 Mechanical Engineering, Pohang University of Science and Technology, Pohang Korea (the Republic of)
Investigating the signaling mechanism and physiological analysis of neuro-muscular junction (NMJ) is important to elucidating undisclosed parts of neuro-muscular diseases. In this respect, in vitro NMJ model has received great attention as a simple and accessible way to understand the physiological mechanism of NMJ, like contractile force, excitatory potentials of muscles and molecular signaling between nerve and muscle generated by nerve stimulation. Especially, microfluidic platforms compartmented by micro-channel have been commonly utilized as the in vitro model to understand each function of neuron and muscle independently. Though the previous studies showed successful formation of neuro-muscular junction and investigations of interactions between neuron and muscles, the in vitro model has a limitation in providing in vivo microenvironment to obtain the NMJ with mature and functional muscle. At this point, it can be a valuable approach to develop an advanced microfluidic platform to achieve a more realistic in vitro NMJ model with a mature and functional muscle. To achieve this, aligned nanofiber mat was firstly adopted to develop an advanced microfluidic platform for the NMJ model. The developed microfluidic platform is compartmentalized by the micro-channel into muscle culture-side and nerve-culture side. At bottom of the muscle culture-side, aligned nanofiber mat was integrated to promote the alignment of the muscle cells. The aligned nanofiber mat was easily integrated onto the PMMA substrate by using electrolyte-assisted electrospinning process, without requiring additional steps to peel off and transfer electro-spun nanofiber from metal collector by mechanical or chemical method. And then a polydimethylsiloxane (PDMS) microfluidic channel is attached beside the aligned nanofiber mat by (3-Aminopropyl)triethoxysilane (APTES) bonding. The NMJ formation with mature and functional muscle in the developed platform was verified through the PC12 and C2C12 cell culture.
8:00 PM - BM04.09.11
Spectroscopic Study of Intermolecular Interactions between Collagen and Heparin
Tadatomo Kawai 1 , Shuhei Hamamura 1 , Yasutada Imamura 1 , Yuzo Itoh 1 Show Abstract
1 School of Advanced Engineering, Kogakuin University, Tokyo Japan
The extracellular matrix which including collagens, fibrin, fibronectin, proteoglycans, glycosaminoglycan, etc., plays an important role in wound healing. Collagen family, especially type I collagen, have been researched for tissue engineering, as tissue scaffolds, dermal matrices and wound dressings for a long time. Type V collagen obstructs to form type I collagen fibril. For fibril formation, heparin inhibit a formation of type I collagen fibril. The type V collagen strongly interacts with heparin. If it can be regulation of collagen fibril assembly by the intermolecular interaction, it may control the cell adhesion, chemotaxis, and migration. In this study, we research the intermolecular interaction between the collagen and heparin by FT-IR and Raman spectroscopy.
FT-IR spectra were obtained using a FTIR microscopy (FT/IR-4600 equipped with a IRT5200, JASCO) by accumulation of 100 scans with a resolution of 4 cm-1. Raman spectra were measured with a Raman microscopy of the laser 488.0 / 514.5nm (NRS-2000, JASCO).
The several peaks of IR spectra of the mixture of collagen V and heparin were shifted, comparing with those of collagen V and those of heparin. The shifted peaks were assigned to amino groups of collagen V and sulfuric acid groups of heparin. The IR spectra of the model compounds were calculated by ab initio molecular orbital method and the normal vibration calculation. It was confirmed that the trend of the peak shifts of the calculated IR spectra of the model compound accorded with the experimental IR spectra. We thought that this peak shifts were caused by intermolecular interaction between amine groups of collagen V and sulfuric acid groups of heparin.
8:00 PM - BM04.09.12
Lead-Free Perovskite (Ba1−xCax)(ZryTi1−y)O3 Thin Film Obtained by Laser Techniques for Biocompatible Applications
Nicu Scarisoreanu 1 , Valentina Dinca 1 , Floriana Craciun 2 , Valentin Ion 1 , Andreea Carmen Andrei 1 , Adrian Bercea 1 , Maria Dinescu 1 , Madalina Icriverzi 3 Show Abstract
1 , National Institute for Lasers, Plasma and Radiation Physics, Magurele Romania, 2 , CNR Istituto dei Sistemi Complessi, Rome Italy, 3 , Institute of Biochemistry of the Romanian Academy, Bucharest Romania
The new lead-free piezoelectric and ferroelectric materials such as (Ba1−xCax)(ZryTi1−y)O3 (BCZT) are key materials in latest dielectric material research due to their high dielectric constant and strong piezoelectric response. Employing laser based techniques to obtain non-toxic (Ba1−xCax)(ZryTi1−y)O3 -BCZT as biomaterial which supports cellular adhesion and proliferation of different kind of cells, can have an important impact on the integration of this material in different biomedical applications. In this work we report the synthesis of functional biocompatible piezoelectric (1-x)Ba(Ti0.8Zr0.2)TiO3 – x(Ba0.7Ca0.3)TiO3, x = 0.45 (BCZT 45) thin films with high piezoelectric properties. Pulsed laser-based techniques, classical pulsed laser deposition (PLD) and matrix-assisted pulsed laser evaporation (MAPLE) have been used to synthesize the thin BCZT 45 films. The second technique has been employed in order to ensure growth on polymer flexible Kapton substrates. The BCZT 45 thin films grown by both techniques show similar structural properties and high piezoelectric coefficient coupling between mechanical loading and electrical potential. While it has long been shown that electrical potential favours biological processes like osteogenesis, the assessment of cell adhesion and osteogenic differentiation onto BCTZ materials has not yet been demonstrated. The BCTZ thin films obtained PLD technique showed high dielectric properties (relative permittivity of about 2200 and tangent loss ~ 1-1.5% at frequency of 10 KHz) measured by dielectric spectroscopy. The local piezoelectric properties (d33~280 pm/V), polarization dynamics and switching characteristics of the samples were investigated by piezoresponse force microscopy technique (PFM). Moreover, the applicability of piezoelectric active layers of BCZT thin films deposited by Matrix-Assisted Pulsed Laser Evaporation (MAPLE) in biotechnology has been demonstrated. Starting from a frozen target of nanopowders of BCTZ and methanol, piezoelectric active thin films of BCTZ have been obtained on Pt-coated kapton substrates, the piezoelectric characterization of the films being made by PFM technique. The mechanisms by which BCZT-based biofilms facilitate cellular adhesion were investigated for different types of cells such as epithelial embryonic kidney HEK 293 cells, human malignant melanoma A 375 cells or human bone marrow-derived mesenchymal stem cells (hBM-MSCs). The cell proliferation studies have been made using human bone marrow-derived mesenchymal stem cells (hBM-MSCs) which were seeded onto BCTZ biomaterial. The results suggest that BCTZ material led to an increased proliferation level and it has been proved that BCZT coatings on Kapton polymer substrates provide optimal support for osteogenic differentiation and viability of hBM-MSCs cells. The hMSC cells are of great interest as progenitors of bone cells, especially in regenerative medicine.
8:00 PM - BM04.09.13
Fast Degradable PGS-Based Shape Memory Polymer Synthesis for Vascular Anastomosis Device
Sun Woong Han 1 , Hong Koo Baik 1 Show Abstract
1 , Yonsei Univ, Seoul Korea (the Republic of)
We demonstrated the synthesis and characterization of a novel poly(glycerol sebacate)(PGS)-based biodegradable shape memory polymer for a novel vascular anastomosis device.
The esterification reaction of glycerol, sebacic acid and fatty acid lead to the obtaining of the biodegradable polymer which can be controllable mechanical and biodegradation properties. This synthesis is relatively simple polymerization that can be carried out under mild conditions without addition of toxic catalysts or crosslinking reagents. Furthermore, the shape memory polymer having a melting temperature of 32°C enables keep its temporary shape at room temperature and recover its original shape at 37°C up to 90%. This biodegradable shape memory polymer has a potential for use in implantable devices, in particular the engineering of small-diameter blood vessels. We fabricated a small-diameter tube (2 mm in diameter) using the synthesized fast degradable PGS-based shape memory polymer for an anastomosis device. The device is advantageous for decreasing surgical time, difficulty and risks due to its non-invasive property.
8:00 PM - BM04.09.14
Creation of GelMA Hydrogels with Physiologically Relevant Compressive Moduli (200-2000 Pa) for Use as 3D In Vitro Cancer Models Using Unique UV Polymerization Source
Sara Hopper 1 , Emily Wilgocki 1 , Kayla Barton 1 , Susan Pomilla 1 , Jason Nichol 1 Show Abstract
1 , Endicott College, Beverly, Massachusetts, United States
The extracellular matrix plays a key role in cancer cell phenotype. It is expected that cancer cells originating from tissues of different stiffnesses will sense and respond to their environment differently, which could yield important information in better understanding cancer cell physiology. Gelatin methacrylate (GelMA) is a UV-crosslinkable hydrogel that has been shown to be effective in the 2D and 3D culturing of cells in a wide variety of applications. Cells can easily bind to GelMA 2D surfaces and within 3D structures, and can proliferate, elongate, and remodel their surroundings due to the presence of natural binding and enzymatic degradation sites in the gelatin backbone. GelMA is a highly elastic material with mechanical stiffnesses demonstrated in the 1 to 30 kPa range through variation of gel concentration and degree of methacrylation, however it has been difficult to produce GelMA hydrogels with compressive moduli below 1 kPa. It has been established that normal breast tissue has a quite low compressive modulus of roughly 200 Pa, whereas precancerous regions have a modulus of roughly 600 Pa and cancerous regions can be as high as 1-2 kPa. Three major aims were investigated in these studies: to create robust GelMA hydrogels in these 3 stiffness ranges using 1 w/v concentration (thus changing only the methacrylation degree), to investigate the differences in behavior, morphology and gene expression of breast cancer cells encapsulated in these hydrogels, and to validate a novel UV polymerization system using a common microscope fluorescent light source with a DAPI/UV filter to avoid the need for an expensive standalone system. An Olympus IX81 microscope equipped with a Lumen Dynamics X-Cite 120Q Fluorescence Light Source and standard UV filter set was used to polymerize the GelMA hydrogels. Validation studies with NIH 3T3 cells determined the parameters and conditions (exposure time, lamp power, photoinitiator etc) whereby cells could be 3D encapsulated with a high degree of early viability that was maintained for at least 1 week. In parallel studies, GelMA concentration was varied from 3-5% w/v, and the methacrylation degree was varied (low, medium, high) to determine which parameters would yield robust hydrogels, while the compressive moduli were determined using an Instron 5944 with 10N load cell. Results demonstrated that it was possible to create and accurately measure GelMA hydrogels with moduli as low as 200 Pa using this system. Finally a breast cancer cell line (MDA-MB231), was used to encapsulate cells within 3D GelMA hydrogels of these 3 stiffness ranges and the cell morphology was tracked using phase contrast images, while RNA was harvested upon completion. Preliminary results suggest that the cells react differently to levels of mechanical stiffness in terms of proliferation and morphology. Ongoing experiments will aim to further quantify these observed effects and to determine differences in gene expression.
8:00 PM - BM04.09.16
Hyaluronic Acid Microfiber Based on Microfluidic Chip
Yun-Kyung Lee 1 , Kwang-Ho Lee 1 Show Abstract
1 , Kangwon University, Chuncheon Korea (the Republic of)
Motivation, and Innovation
To date, the tissue engineering has been introduced to restore or replace damaged tissues. Hyaluronic acid (HA) belonging to the natural polymeric materials was been applied to cosmetic and regenerative medicine fields because HA has the excellent biocompatibility and ability to contain moisture up to several hundred times its own weight. In previous researches, there have been efforts to form such HA as structure like the fillers. However, since the biodegradation rate was fast and shape control was difficult, development of a 3-dimensional structure such as microfibers has been limited. Here, we employed a co-axial microfluidic chip to make rapid synthesis and produce a 3-dimensional HA microfibers. Our approach enabled to overcome the limitations of the HA microfiber fabrication process in previous researches.
We used a polydimethylsiloxane (PDMS)-based microfluidic chip composed of center channel and sheath channel to make of HA microfibers. HA solution was mixed with NaOH and 1, 4-buthanediol diglycidyl ether (BDDE) as a crosslinking agent and then prepared solution was injected into the center channel of the microfluidic chip. Ethanol solution was injected into the sheath channel that designed to induce extraction of the HA solution surrounding the center channel. To evaluate the applicability of HA fiber as a filler, the surface characteristics, chemical compositions, tensile strength, fluorescence material immobilization, swelling properties and cell compatibility test for HA microfibers were performed.
We suggested that the diameter of HA microfibers depends on the flow conditions of the HA solution and these conditional control was easily done in the microfluidic channel. In order to verify the drug immobilization of HA microfibers, we proved that fluorescent particles of 10 μm and solutions of 10kDa could be stably immobilized in the HA microfibers. The tensile strength of 0.0152 MPa, bending test, and swelling test showed that HA microfibers can be applied as medical threads. Also, we cultured cells on the HA microfiber for 5 days and analyzed the cell viability to about 98%. These high cell proliferation and survival rate verified the cell compatibility of HA microfibers.
In this research, we described innovative fabrication methods of HA microfibers produced by the 3D co-axial microfluidic chip. We solved the side effect such as degradation of mechanical properties when the HA was applied in the liquid state to the living body. The results of various analysis showed that HA microfibers can be safely used as dermal filler in tissue engineering.
Acknowledgements: This research was supported by a grant from the National Science Foundation (NRF) funded by the Ministry of Education (2017R1D1A1A02019351).
8:00 PM - BM04.09.17
Injectable Cryogel Scaffolds with Inherent Antimicrobial Properties
Kasturi Joshi Navare 1 , Pierre Villard 1 , Mahboobeh Rezaeeyazdi 1 , Thibault Colombani 1 , Devyesh Rana 1 , Nicole Bassous 1 , Thomas Webster 1 2 , Sidi Bencherif 1 3 4 Show Abstract
1 Chemical Engineering, Northeastern University, Boston, Massachusetts, United States, 2 , Wenzhou Institute for Biomaterials and Engineering, Wenzhou China, 3 School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts, United States, 4 , Sorbonne University, UTC CNRS UMR 7338, Biomechanics and Bioengineering (BMBI), University of Technology of Compiègne, Compiègne France
The use of biocompatible 3D polymeric scaffolds is set to revolutionize regenerative medicine. Hydrogels are a class of highly hydrated polymeric materials used as 3D scaffolds in tissue engineering due to their unique properties such as high-water content, softness, flexibility, and biocompatibility. Recently, we have developed highly elastic and syringe-injectable biomimetic cryogels. Injectable cryogels are a class of polymeric hydrogels with unique properties including large and interconnected pores, mechanical robustness, and injectability with shape memory properties. However, hydrated environment can also facilitate microbial infections. There is high prevalence of antibiotic-resistant microbial infections, which often result in impaired wound healing and biomedical implant failure. In this situation, it is critical that gels possess antimicrobial characteristics in addition to serving their primary functional role. Here in order to impart antimicrobial properties, we have incorporated radicals generating particles into cryogels while retaining cytocompability. These radical generating gels resulted in the inhibition of superbugs found in implant associated infections, namely multidrug resistant Escherichia coli, Methicillin resistant Staphylococcus aureus, Pseudomonas aeruginosa, Staphylococcus epidermidis. The biocompatibility and mechanical properties of antimicrobial cryogels were assessed and found to be suitable for tissue engineering applications. The cryogels made thus address two main challenges faced by gel implants, such as biomaterial associated infections while potentially providing suitable three-dimensional microarchitectural features for guided tissue regeneration and biointegration.
8:00 PM - BM04.09.18
Studies on the Interaction of Type Ii Collagen-Lipid with DPPC Using Langmuir Monolayers
Lucineia Ceridório 1 , Mikaela Santos 1 , Lilian Ramos 1 , Luciano Caseli 1 Show Abstract
1 , Universidade Federal de São Paulo, Diadema (SP) Brazil
Type II collagen is the predominant structural protein in the extracellular matrix of the articular cartilage. The deposit of layers of ordered collagen molecules with nanometric scale is interesting for several applications in the biomedical area. The Langmuir-Blodgett technique has been proposed for the deposition of oriented collagen layers onto solid supports. In this work, phospholipid Langmuir monolayer is used for the study of the properties of type II collagen at the air-water interface in order to investigate the molecular interactions between molecules of cell membranes and proteins. The effects of type II collagen on dipalmitoylphosphatidylcholine (DPPC) monolayer were studied by kinetics measurements, surface pressure-area isotherms, compression-decompression curves, polarization modulation infrared reflection-absorption spectroscopy (PM-IRRAS) and Brewster angle microscopy (BAM). Solutions of type II collagen in acetic acid and phosphate buffer solution (pH 7.2) were employed as subphase of the lipid monolayer. The addition of collagen in the subphase of DPPC monolayer shifted the isotherms to higher molecular areas, indicating the adsorption of the protein in the phospholipid monolayer. Successive cycles of the compression-decompression lead the isotherms to lower values of molecular area due to the progressive removal of the protein from the DPPC monolayer. With the collagen in the subphase, PM-IRRAS showed changes in the DPPC spectra, mainly in the regions for the CH-stretching modes in the acyl chains (between 2850 -2970 cm-1), and in the region for the CO-stretching mode of the polar head group (~ 1730 cm-1), and also for the amide I band (~ 1614 cm-1). These results suggested that intermolecular forces guide the adsorption of the type II collagen in lipid monolayer. The BAM images revealed that the morphology of DPPC monolayer was affected by the protein. The data demonstrated the type II collagen is a molecule that interact with DPPC monolayer enabling the transfer to solid supports by the Langmuir-Blodgett technique to be used for deposition of type II collagen for bioinspired materials.
Acknowledgments: This work was supported by CNPq, CAPES and FAPESP.
1- Pastorino, L; Dellacasa, E, Scaglione; et all, Oriented collagen nanocoatings for tissue engineering. Colloids and Surfaces B: Biointerfaces. 114 , 372– 378, 2014
2- Mady, M. M.; Biophysical Studies on Collagen-Lipid Interaction. Journal of of Bioscence and Bioengineering. 104, 144-148, 2007.
8:00 PM - BM04.09.19
Engineering Conductive Fibrous Patches for Cardiac Tissue Regeneration
Brian Walker 1 , Chu Yu 1 , Ehsan Shirzaei Sani 1 , William Kimball 1 , Nasim Annabi 1 2 3 Show Abstract
1 , Northeastern University, Boston, Massachusetts, United States, 2 Biomaterials Innovation Research Center, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts, United States, 3 Harvard-MIT Division of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States
Tissue engineering approaches particularly using bioengineered scaffolds have been applied for the treatment in patients suffering from heart failure after myocardial infarction (MI). Electrospinning is one of the most widely utilized techniques for engineering fibrous scaffolds for tissue engineering applications. In this study, we aimed to engineer a fibrous scaffold with tunable conductivity and physical properties based on combining a photocrosslinkable biopolymer, gelatin methacryloyl (GelMA) and a bio-ionic liquid (Bio-IL), choline acrylate. GelMA-based biomaterials possess tunable mechanical properties, favorable biodegradation, and cell binding sites, which makes them suitable for fabrication of tissue engineering scaffolds. In addition, the Bio-IL used in this study is conductive and non-cytotoxic. To engineer the fibrous scaffolds, different concentrations of GelMA prepolymer solutions in hexafluoroisopropanol (HFIP) were prepared, and fibrous GelMA mats were then formed using electrospinning technique. The Bio-IL was then added to cold water in different ratios, injected on the frozen GelMA fibers to penetrate among the fibers. Lastly, the samples were exposed to UV light for 300 sec to crosslink the GelMA/Bio-IL scaffolds.
Our results demonstrated that the engineered electrospun GelMA/Bio-IL scaffolds showed tunable mechanical strength and conductivity, based on GelMA and Bio-IL concentrations. The elastic modulus and conductivity of engineered materials increased concomitantly with an increase in the concentration of Bio-IL. For example, the elastic modulus of scaffolds containing 12.5% GelMA increased from 75.25 ± 5.59 kPa to 173.9 ± 11.88 kPa by increasing the Bio-IL concentration from 33% to 66% (v/v). Furthermore, the conductivity of a 10% GelMA/Bio-IL patch also increased from 1.23 ± 0.16 S/m to 3.56 ± 0.47 S/m when the Bio-IL concentration increased from 33% to 66%. The in vitro cytocompatibility of the fibrous scaffolds was confirmed by seeding primary rat cardiomyocytes (CMs) on the engineered scaffolds. Taken together, our results demonstrate the remarkable potential of GelMA/Bio-IL fibrous scaffolds for the engineering of cardiac tissue patches.
8:00 PM - BM04.09.20
Biomaterial Instructive Cues for CNS Interventions
Virginia Ayres 1 , Volkan Tiryaki 2 , Ijaz Ahmed 3 , David Shreiber 3 , Jesus Garcia 1 Show Abstract
1 , Michigan State University, East Lansing, Michigan, United States, 2 , Siirt University, Siirt Turkey, 3 , Rutgers, The State University of New Jersey, Piscataway, New Jersey, United States
Until recently, therapeutic implants such as stents, shunts, probes, wafers and scaffolds have been mainly viewed as passive vehicles for the delivery of physical, pharmacological and cellular interventions. Recent research, however, indicates that therapeutic implants create local physical environments that supply directive cues in their own right that work in conjunction with biochemical cues and produce a jointly-directed outcome. In our research, we quantitatively investigate the nanoscale directive cues presented by synthetic electrospun nanofibrillar scaffolds that have demonstrated promise for regenerative repair of central nervous system (brain and spinal cord) injuries [1-3]. The nanophysical cues presented by the nanofibrillar scaffolds are determined using advanced scanning probe microscopy and contact angle measurements to quantitatively assess nanoscale elasticity, surface roughness, work of adhesion and surface polarity. We couple this information with quantitative analyses of neural cell system responses from key cellular players: astrocytes, neurons and oligodendrocytes. Confocal and super-resolution microscopy-based immunocytochemistry and atomic force microscopy are used to quantify spreading and other morphological response and protein responses to simulated quiescent and injury situations. Cluster analyses are used to assess the impacts of directive nanophysical cues on cellular responses within an n-dimensional parameter space.
Our current work focuses on how a scaffold-based intervention could eventually contribute new options for the treatment of perforating/penetrating central nervous system injuries. This is an extremely challenging problem, with limited treatment options for surviving patients beyond costly and uncertain rehabilitation. One hurdle is the formation of a glial scar, in which reactive astrocytes surround a wound site and develop tightly interwoven processes while expressing inhibitory proteoglycans and tenascins, which biomechanically and biochemically block axonal elongation and reconnection. Our recent research indicates that it may be possible to design biomaterials with instructive nanoscale cues that cooperate with the local environment to steer astrocytes away from the reactive state and into a permissive state that allows neuron reconnection across a wound site and resumed function.
 VM Tiryaki, VM, U Adia-Nimuwa, VM Ayres, I Ahmed, DI Shreiber. Texture-based segmentation and a new cell shape index for quantitative analysis of cell spreading in AFM images. Cytometry: Part A, 2015, 87(12): 1090-1100.
 VM Tiryaki, VM Ayres, I Ahmed, DI Shreiber. Differentiation of reactive-like astrocytes cultured on nanofibrillar and comparative culture surfaces. Nanomedicine, 2015 10(4): 529–545.
 VM Tiryaki, VM Ayres, AA Khan, I Ahmed, DI Shreiber, S Meiners. Nanofibrillar scaffolds induce preferential activation of Rho GTPases in cerebral cortical astrocytes. Int. J. Nanomedicine. 2012 7:3891-3905.
8:00 PM - BM04.09.21
Characterization of Tissue-Engineered Blood-Brain Barrier Microvessels
Jason Luo 1 , André Adams 1 , Kyle DiVito 1 , Sumati Sundaram 2 , Katherine Rogers 3 , Monique van Hoek 2 , Kylene Kehn-Hall 2 , Cynthia de la Fuente 2 Show Abstract
1 , United States Naval Research Laboratory, Washington, District of Columbia, United States, 2 , George Mason University, Manassas, Virginia, United States, 3 , University of Maryland, College Park, College Park, Maryland, United States
The blood-brain barrier (BBB) is a highly selective semipermeable membrane which restricts nearly all molecular transport between the circulatory and neurological systems to highly regulated intracellular processes. As such, an ideal in vitro BBB model has applications in the development of neurologically therapeutic molecules that can more effectively penetrate the BBB. Additionally, diseases of the blood-brain barrier itself, manifesting in a loss of molecular selectivity, are correlated with neurodegenerative diseases such as Alzheimer’s and multiple sclerosis. An in vitro BBB model thus has additional applications in understanding the development and restoration of barrier function, which might in turn yield novel therapies for various neurodegenerative diseases.
Initial work was performed in 2D Transwell cultures in a manner consistent with other 2D BBB models from the literature, and protocols were established to induce the development of barrier properties in both human umbilical vein endothelial cells (HUVECs) and brain microvascular endothelial cells (BMECs) derived from induced pluripotent stem cells. Barrier properties were quantified via immunostaining for the intracellular proteins ZO-1, claudin, and occludin, as well as tracking the diffusion of size-specific fluorophores across the cell membrane: in treated HUVEC monolayers, 20-KDa FITC and 70k-KDa FITC dyes experienced 67% and 85% reductions in transmembrane mobility, respectively. Additionally, given that the BBB greatly reduces paracellular ionic transport, its development is directly correlated with an increase in the electrical resistance across the thickness of the cell layer. Quantification of this transendothelial electrical resistance (TEER) was performed using an EVOM2 four-terminal sensor: treated HUVECs and BMECs displayed up to 80 Ωcm2 and 1500 Ωcm2 increases in TEER over time, respectively.
These barrier-inducing protocols were then translated to engineered microvessels fabricated using a sheath-flow microfluidic system that forms tubular PEG/gelatin-methacrylamide scaffolds with luminal walls lined with an endothelial monolayer; the lumens themselves are 100µm in diameter. A finite element model of the engineered vessel was developed and used to predict permeability kinetics and transendothelial resistance as a function of barrier development. Barrier properties in the monolayer were confirmed statically via immunostaining and dynamically by measurement of fluorescent dyes’ transendothelial diffusion kinetics. TEER measurements were performed using the same EVOM2; microelectrode geometry for performing the three-dimensional assay was optimized in simulation. Finally, under dynamic mechanical analysis it was noted that under a 3% strain applied sinusoidally at 1Hz, vessels displayed a 25% increase in Young’s modulus and 33.3% increase in their stress:strain phase lag as the BBB layer developed, corresponding with the formation of a structurally sound basement membrane.
8:00 PM - BM04.09.22
Autoclavable Cryogel Scaffolds for Biomedical Applications
Pierre Villard 1 , Mahboobeh Rezaeeyazdi 1 , Kasturi Joshi Navare 1 , Thibault Colombani 1 , Sidi Bencherif 1 2 3 Show Abstract
1 , Northeastern University, Boston, Massachusetts, United States, 2 , Harvard University, Cambridge, Massachusetts, United States, 3 , University of Technology of Compiègne, Compiègne France
Sterilization is a critical step in medical device manufacturing which affects the safety and efficacy of medical devices as this process kills, deactivates, or eliminates all forms of life and other biological agents. Toward this end, sterilization of biomaterials is critical to assure the hygienic requirements for various biomedical applications. Sterilization can be achieved with several techniques including heat, chemicals, irradiation, high pressure, and filtration. One of the most popular sterilization method used in the laboratory is heat sterilization and more particularly autoclaving. However, autoclaves use steam heated up to 134 °C under high pressure which could be detrimental to a number of biomaterials including polymeric hydrogels. Hydrogels have become very popular in the biomedical field due to their unique properties such as high water content, softness, flexibility and biocompatibility. However, hydrogels are typically not sterilized but rather sanitized, especially in academic institutions, as they are unable to overcome the drastic autoclave sterilization process. Recently, we have developed mechanical robust injectable cryogels. Injectable cryogels are a class of hydrogels with unique properties including large and interconnected pores, mechanically robust sustaining up to 90% deformation, and injectability with shape memory properties. A series of cryogels prepared with various biopolymers have been autoclaved and tested not only for their degrees of sterilization but also for their physico-chemical integrities (injectability, physical properties, and intrinsic biological properties). Our preliminary data suggest that unlike conventional hydrogels, injectable cryogels are resilient to the aggressive steam sterilization conditions.
Acknowledgement: This research was financially supported by Northeastern University (Tier 1 Provost grant).
8:00 PM - BM04.09.23
Ag/BSA Nanoparticles Encapsulated Hydrogel for Potential Wound Dressing Application
Berhanu Zewde 1 , Dharmaraj Raghavan 1 Show Abstract
1 , Howard University, Washington, District of Columbia, United States
Nanocomposite hydrogels (NCH) have been increasingly studied for biomedical applications. However, most of the NCH lack control over the release of nanoparticles in the region of interest. The triggered release of nanoparticles from NCH is influenced by a number of parameters. Here, we investigate the release profile of nanoparticles from crosslinked NCH as a function of the external stimuli and the overall structure of the NCH. In this regard, we synthesized NCH by insitu encapsulation of preformed Ag/BSA nanoparticles while the norborenene hyaluronic acid (NorHA) reacted with PEGylated tetrazine (PEG-Tz macromer) via click chemistry. FTIR, NMR and TGA was used to characterize the hydrogel, and PEG-Tz macromer as well as NorHA precursor prior to crosslinking so as to establish that during hydrogel formation the precursor functionality indeed reacted. Results from FTIR support the disappearance of tetrazine functionality at 1667 cm_1 upon reaction of PEG-Tz macromer with NorHA. The encapsulation of Ag/BSA nanoparticles in the hydrogel was established by quantifying the silver content in the NCH using SEM/EDAX, AAS and TGA techniques. Desorption studies of nanoparticles release from hydrogel in aqueous medium showed a sudden burst in nanoparticles release followed by a gradual release. Additionally, we were able to regulate the release of nanoparticles from hydrogel by varying the concentration of NorHA in the formulation of nanocomposite hydrogel. These findings can have a strong bearing in acheiving regulated release of nanoparticles from potential wound dressing applications because released silver nanoparticles can exert positive effects on wound healing through their antimicrobial properties, reduction in wound inflammation, and modulation of fibrogenic cytokines. Further studies are underway to investigate the dependence of nanoparticles release from nanocomposite hydrogel as a function of external stimuli.
Acknowledgement : US Army/MIT-ISN W911NF-13-D-0001
8:00 PM - BM04.09.24
Bijels-Derived Hydrogel Hybrid Membranes for Medical Applications
Haoran Sun 1 , Min Wang 1 Show Abstract
1 Department of Mechanical Engineering, The University of Hong Kong, Hong Kong Hong Kong
Bicontinuous interfacially jammed emulsion gels (bijels) are new structures discovered recently. They are composed of two interwoven networks, with each being a liquid-phase material that is immiscible with the other. Bijels can be solidified through processes such as polymerization, resulting in materials with bicontinuous structures. Since distinctive release behaviors of bio-agents with different functions from the same carrier are often required in the biomedical field, the bicontinuous architectures make bijels-derived materials very attractive for applications such as controlled release in tissue engineering. Traditionally, bijels are made via thermal quenching. However, this technique can be applied only to certain pairs of liquids and delicately prepared colloids with relatively low thermal stability. The batch process of thermal quenching also limits the potential of bijels. Solvent transfer-induced phase separation (STRIPS), which provides a facile way to continuously fabricate bijels in various shapes (particles, fibers and films) with high thermal stability, thus becomes very appealing. A major issue for applying bijels-derived materials in tissue engineering is the biocompatibility of most of current bijels systems. To make bijels-derived materials useful in tissue engineering, biocompatible hydrogels may be used in bijels systems. In this study, the fabrication of biocompatible hydrogel hybrid membranes via bijels using the STRIPS technique was investigated. To produce bijels structures, a ternary liquid mixture was made by adding ethanol, hexanedioldiacrylate (HDA), 2-hydroxy-2-methylpropiophenone, distilled water, Ludox TMA, and CTAB in ethanol (0.2 M). HCL was added to this mixture to adjust pH. A polystyrene plate was immersed in the ternary mixture for forming a mixture film on its surface. It was taken out and then immersed in a water bath (10-3 M CTAB) to form bijels film. A high-intensity UV light cured HDA monomer, solidifying the bijels structure. After curing, bijels films were immersed in Na-alginate solution, taken out and immersed in CaCl2 solution for crosslinking. The products were freeze-dried. The bicontinuous structure of bijels films (before introducing Na-alginate) could be clearly seen under SEM. This microstructure was composed of solidified HDA and liquid water. The water phase was evaporated after freeze-drying, leaving continuous channels in bijels-derived structure. The channel diameter was 3-10 µm. After adding Na-alginate and crosslinking, these channels were filled with cross-linked Ca-alginate hydrogel, leading to successful fabrication of bijels-derived hydrogel hybrid membranes. For controlled release, a bio-agent could be incorporated in Ca-alginate hydrogel. It could be released when a dilute sodium citrate solution broke down Ca-alginate. This study has demonstrated that it is feasible to produce biocompatible bijels-derived membranes for potential biomedical applications.
8:00 PM - BM04.09.25
High Pressure Engineering for Biological Scaffolds—Xenogeneic Acellular Small-Diameter Vascular Grafts and Autologous Inactivated Dermis
Tetsuji Yamaoka 1 , Naoki Morimoto 2 , Atsushi Mahara 1 Show Abstract
1 , National Cerebral and Cardiovascular Center Research Institute, Suita Japan, 2 Plastic and Reconstructive Surgery, Kansai Medical University, Hirakata Japan
We have studied the effect of high hydrostatic pressure (HHP) ranging from 100 to 1,000 MPa (from 1,000 atm to 10,000atm) on various cells and tissues. By pressing at 200 MPa for 10 min, all types of mammalian cells died in suspension form, in monolayer culture, and even inside the native tissue, while the cells pressed at 170MPa adhered and grew in a similar manner to those of untreated cells. When HHP increased over 500 MPa, the cell membrane was destroyed and ECM started to denature, which results in increased decellularization efficacy from the tissue. We utilized this HHP treatment for preparing xenogeneic acellular small-diameter vascular grafts and autologous inactivated dermis
First, the ostrich carotid arteries with the inner diameter of 2mm and length of 30cm were decellularized with HHP at 1,000MPa and the blood vessels were transplanted in porcine femoral-femoral cross over bypass surgery as a control experiment. Since the decellularized blood vessels contains collagen as a main component, they lead to the severe clotting resulting in lower patency in large animal models. Then, we established a novel EPC (endothelial progenitor cell) capturing technology using a synthetic peptides composed of collagen-binding sequence and EPC capturing sequence. The affinity of the modified tissue were quantitatively analyzed in vitro system and they were transplanted to minipigs. Surprisingly, in three weeks transplantation, the modified grafts were completely patent, which was the first report for this thin and long blood vessels. The luminal surface of the grafts did not have clotting but covered by the endothelial marker positive and progenitor marker double positive cells, which is a proof of EPC capturing onto the modified surface. We are now evaluating artery replacement model of goats and followed one year results.
Next, we tried to utilize this HHP treatment technology as a novel tumor treatment. As mentioned above, HHP at 200 MPa killed the cells in the tissues completely without resulting in ECM denature, which raised an idea to treat resected tumor tissue with HHP and transplant back to the same position of the patients. Since the treated tissue is autologous, removal of cellular debris is not needed. We tried to inactivate the human giant congenital melanocytic nevi (GCMN) tissue which is very hard to treat by the conventional methods. The pressed nevus tissues were analyzed in detail and also transplanted into nude mice subcutaneously for 1 year to confirm complete cell killing. Our result suggested that GCMN can be completely inactivated by HHP and the inactivated tissue can be used as the autologous implantable dermal substitute. The clinical trials were just started.