Meetings & Events

spring 1998 logo1998 MRS Spring Meeting & Exhibit

April 13 - 17, 1998 | San Francisco
Meeting Chairs: John A. Emerson, Ronald Gibala, Caroline A. Ross, Leo J. Schowalter









Symposium CC—Biomaterials Regulating Cell Function and Tissue Development

Chairs 

Kevin Healy 
Division of Biological Materials 
Northwestern Univ 
Chicago, IL 60611-3008 
312-503-4735

Yoshito Ikada
Research Ctr for Biomedical Engr
Kyoto Univ
Kyoto, 606 JAPAN
81-75-7514115

Antonios Mikos 
Inst of Biosciences & Bioengineering 
Rice Univ 
Houston, TX 77251 
713-285-5355

David Mooney
Dept of Bio & Matls Sci & Chem Engr
Univ of Michigan
Rm 3074 Dow Bldg
Ann Arbor, MI 48109-2136
313-763-4816

Robert Thomson 
W. L. Gore & Assoc 
Flagstaff, AZ 86002-0300 
520-779-2771 

Proceedings published as Volume 530 
of the Materials Research Society 
Symposium Proceedings Series.
 

* Invited paper

SESSION CC1: BIOMATERIALS IN TISSUE ENGINEERING 
Chairs: Antonios G. Mikos and David Mooney 
Monday Morning, April 13, 1998 
Pacific A
8:30 AM *CC1.1 UROLOGICAL APPLICATIONS OF AUTOLOGOUS CELL THERAPY. Daniel R. Omstead and Frank T. Gentile, Reprogenesis, Inc. Cambridge, MA. 

Nearly 8 million surgical procedures are performed annually in the United States to treat organ and tissue deficiencies. These surgeries result in approximately 40-90 million hospital days with cost of treatment, support, and loss of productivity estimated at over $400 billion. Reprogenesis has developed an autologous cell therapy to treat a number of disorders. The first clinical application of this work is to develop a treatment for two urological disorders: vesicoureteral reflux in pediatric patients and urinary stress incontinence in adults. This product contains autologous cells (chondrocytes) suspended in a crosslinked alginate hydrogel which is injected submucosally at critical sites surrounding the ureteral orifice (in reflux) and the urethra and bladder neck (in incontinence). In reflux, the cell-alginate matrix serves as a bulking agent to prevent retrograde flow of urine from the bladder to the ureter and kidney. In incontinence, the bulking agent allows proper closure of the urethral sphincter. The key manufacturing components of this product are: 
1) isolation of chondrocytes from an auricular cartilage biopsy, 
2) expansion and possible storage (cryopreservation) of those cells in vitro, 
3) formulation of these cells into an alginate hydrogel matrix, and 
4) mixing of the cell-alginate suspension with crosslinking agents that permits injection if the formulation as a partially crosslinked hydrogel that then becomes a fully crosslinked extracellular matrix after placement within the submucosal tissues. Based on preclinical studies (Atala et al., 1993; Paige et al. 1995 and preclinical studies performed by Reprogenesis Inc., it is expected that the crosslinked alginate hydrogel will be replaced by natural cartilage matrix constituents produced by the implanted cells such that the neocartilage is formed in vivo. A process has been developed by Reprogenesis, Inc. To produce the individual components of this product, as well as the final product (Chondrocytes-Alginate Gel Suspension) in a controlled and repeatable fashion so as to produce a product that is safe and predicable. Similar approaches (i.e. autologous cells injected in a hydrogel matrix) are under active investigations in a number of other therapeutic areas. 

9:00 AM *CC1.2 
DEVELOPMENT EXPERIENCES WITH CULTURED SKIN. Janet Hardin-Young, Organogenesis, Canton, MA. 

Formation of a functional tissue requires complex interactions between matrix proteins and different cell populations which comprise the tissue. Organotypic cultures create a permissive environment for these interactions to occur. The human skin equivalent (HSE) is an organotypic skin contruct which consists of a contracted collagen lattice containing dermal fibroblasts overlaid with a cornified epidermal layer. In clinical studies, the HSE is effective in the treatment of chronic and acute wounds. The HSE provides immediate coverage to close wounds and provides cytokines and matrix components which are necessary for wound healing. The HSE has been useful in examining the influence of cell-matrix interactions on cell differentiation, basement membrane assembly, and in vivo performance. When wounded in vitro, it displays a normal healing response in regard to cytokine production, keratinocyte migration, re-epithelialization and differentiation resulting in in vitro ``healing''. These results demonstrate that the HSE is an interactive tissue capable of responding to the surrounding wound bed. Understanding which interactions between cells and matrix proteins are important for in vitro tissue development is a vital step in utilizing these interactions to improve tissue-engineered grafts. 

9:30 AM CC1.3 
A NEW APPROACH TO CARTILAGE TISSUE ENGINEERING USING HUMAN DERMAL FIBROBLASTS SEEDED ON THREE-DIMENSIONAL POLYMER SCAFFOLDS. Steven B. Nicoll, Univ. of California, Joint Bioengineering Graduate Group, Berkeley and San Francisco, CA; Anna Wedrychowska, Univ. of California, Laboratory of Connective Tissue Biochemistry, San Francisco, CA; Nancy Smith, California State University, Microscopy and Graphical Imaging Center, Hayward, CA; Rajendra S. Bhatnagar, Univ. of California, Joint Bioengineering Graduate Group, Berkeley and San Francisco, CA. 

Current methods for correcting articular cartilage defects are limited by a scarcity of cartilage cells. Here we describe a novel method for the conversion of human dermal fibroblasts to chondrocyte-like cells and the potential application of this methodology to cartilage tissue engineering. Human neonatal foreskin fibroblasts (HFFs) were seeded on two-dimensional, tissue culture polystyrene (TCPS) in high density micromass cultures in the presence staurosporine (50-200 nM), a protein kinase C (PKC) inhibitor, and lactic acid (40 mM) to induce functional hypoxia. HFFs were similarly cultured on three-dimensional polymer scaffolds composed of a non-woven polyglycolic acid (PGA) fiber mesh reinforced in a dilute solution of poly(L-lactic acid) (PLLA). At 24 hours, northern analysis revealed a staurosporine dose-dependent increase in aggrecan core protein expression in lactate-treated micromass cultures on TCPS, while type I collagen gene expression was virtually abolished in all cultures supplemented with staurosporine. The cells in these cultures displayed a rounded, cobblestone-shaped morphology typical of differentiated chondrocytes (most pronounced at 200 nM staurosporine and 40 mM lactate), and were organized into nodules which stained positively with Alcian blue. When seeded on PGA/PLLA matrices under identical conditions as described for TCPS, a chondrocyte-like morphology was observed in cultures treated with lactate and staurosporine in contrast to the flattened sheets of fibroblast-like cells seen in untreated controls. Taken together, the above findings suggest that staurosporine treatment coupled with high density micromass culture in a functionally hypoxic environment induces chondrogenic differentiation in HFFs, and that these cells may be used in concert with three-dimensional polymer scaffolds for the repair of articular cartilage lesions. 

9:45 AM CC1.4 
USE OF ALLOGRAFT DERMAL MATRIX FOR INTRAORAL RESURFACING. Craig D. Friedman, John Andrew Ridge, Fox Chase Cancer Center, Department of Surgery, Philadelphia, PA; Peter D. Costantino, Dept of Otolaryngology, Mt. Sinai Medical Center, New York, NY. 

Neoplastic, traumatic, and inflammatory diseases often lead to full-thickness mucosal defects in the oral cavity. The standard reconstruction of significant mucosal defects has been an autologous split thickness skin graft. Defects will heal through re-epithelialization by peripheral mucosa with significant patient discomfort, delay in healing, and scarification which results in loss of function of the intraoral tissues. Successful skin grafts result in a metaplastic squamous epithelium covering the recipient site, and associated donor site morbidity. Since defects re-epithelialize by the migration of peripheral mucosal cells, the use of an appropriate matrix which supports cell migration might represent an alternative to autologous skin grafting for intraoral resurfacing. Allograft dermal matrix (AlloDerm), an acellular processed tissue graft, was applied to eighteen patients with full thickness defects of the oral cavity as an alternative to autologous split thickness skin graft. Patients had full thickness mucosal defects of the oral cavity and no prior history of radiation therapy. Defects were reconstructed with the AlloDerm dermal matrix in the same technical fashion as with an autologous skin grafting for intraoral resurfacing. Patients were evaluated for functional return time to re-epithelialization, associated pain and discomfort, and location within the oral cavity. Overall success rate was 90% with complete epithelialization noted on clinical examination within four weeks. Allograft dermal matrix may be considered a useful reconstructive option for patients with oral mucosal defects. 

10:15 AM *CC1.5 
ACELLULAR BIOLOGICAL MATRICES AS TEMPLATES FOR TISSUE RECONSTITUTION. Stephen Livesey, John Harper, Abhijit Nag, Lawrence Boerboom, Chris Coleman, Sy Griffey, LifeCell Corporation, The Woodlands, TX. 

The extracellular matrix of tissues is now recognized as a highly complex, three-dimensional array of specific components. These components interact via specific cell surface receptors to regulate cell phenotypic expression and hence, tissue maintenance and remodeling. LifeCell's technology involves the processing and preservation of biological tissues to deliver structurally and biochemically intact acellular tissue matrices. Specific components of the technology involve decellularization, matrix stabilization and preservation. Application of this approach to date has been in the areas of dermis, heart valves and vascular conduits. Transplantation studies of the acellular dermal matrix (AlloDerm) have demonstrated in animals the ability of the matrix to undergo revascularization and cellular repopulation without specific or nonspecific host immune response. Clinical use of AlloDerm has been in the treatment of full thickness burns, dental surgery and reconstructive plastic surgery; either as a skin graft or as an implant to correct soft tissue loss. Heart valves and vascular conduit processing has resulted in matrices which in animal studies do not calcify and undergo cellular repopulation. Analysis for cell differentiation markers has been consistent with the repopulation of specific matrix locations with cells expressing appropriate phenotype. Analysis of host-matrix interaction has indicated the importance of matrix components in directing the host cell response and hence tissue maintenance and remodeling. 

10:45 AM CC1.6 
A BIORESORBABLE SCAFFOLD FOR AUGMENTATION CYSTOPLASTY IN THE DOG MODEL. Stephen F. Badylak, Timothy B. McPherson, Hongyu A. Liang, Jason P. Hodde, Purdue University, Hillenbrand Biomedical Engineering Center, West Lafayette, IN; Bradley P. Kropp, University of Oklahoma, Department of Urology, Oklahoma City, OK. 

Xenogeneic small intestinal submuscosa (SIS) has been shown to function as a resorbable scaffold around which new urinary bladder tissue forms. The newly deposited tissue is indistinguishable either functionally or structurally from normal urinary bladder tissue after three to four months. The originally implanted SIS biomaterial can no longer be recognized as early as 28 days after surgery using hematoxylin and eosin (H&E) staining. A monoclonal antibody (MoAb) identified as 8C8 against porcine SIS has been developed and can be used as a marker to track the presence of this biomaterial over time. The anterior dome (approximately 40% of the total bladder surface area) was surgically excised in nine dogs and replaced with porcine origin SIS. Three dogs were sacrificed four weeks after surgery, three dogs were sacrificed eight weeks after surgery, and the final three dogs were sacrificed twelve weeks after surgery. Tissues were collected for staining with H&E and Masson's trichrome stain. Immunohistochemistry with 8C8 MoAb was also performed. At 28 days there was a complete transitional epithelial cell lining on a basement membrane structure in the remodeled SIS. The underlying connective tissue was very vascular and contained a mixture of partially organized collagen and scattered bundles of smooth muscle cells. Scattered islands of 8C8-positive staining tissue were present in the excised grafts. At eight weeks after surgery, the submucosal tissue consisted of organized connective tissue with greater amounts of smooth muscle. There was no evidence for remaining SIS at eight weeks. By twelve weeks after surgery, there were organized bundles of smooth muscle present and the wall of the urinary bladder was of normal thickness. Thin bands of collagenous connective tissue were randomly interspersed throughout the bundles of smooth muscle. 

11:00 AM CC1.7 
INVESTIGATING CELL IMMUNOISOLATION PARAMETERS USING MICROFABRICATED MEMBRANES. Tejal A. Desai, Derek J. Hansford, and Mauro Ferrari, Biomedical Microdevices Center, University of California, Berkeley, CA. 

The clinical success of immunoisolation membrane technology has thus far been limited since it is still not entirely clear what physical parameters are required for the complete blockage of immune components. Several studies have shown that for effective immunoisolation, transplanted cells should be prevented from interacting with host C1q, IgM and IgG. Therefore, the diffusion of IgG, IgM, and C1q molecules through microfabricated inorganic membranes with uniform and well-controlled pores sizes in the tens of nanometer range was investigated using a two compartment rotating diffusion chamber over a period of a week. It was determined whether membranes with pore sizes in the tens of nanometers could provide sufficient immunoprotection based on size exclusion without hindering necessary nutrient diffusion 

11:15 AM CC1.8 
COMPARISON OF RELEASE RPOFILES OF VARIOUS GROWTH FACTORS FROM >BIODEGRADABALE CARRIERS. Yasuhiko Tabata, Masaya Yamamoto, Yoshito Ikada, Research Center for Biomedical Engineering, Kyoto University, Sakyo-ku, JAPAN. 

Various growth factors act on different cells to regulate their >proliferation and differentiation. However, upon applying the growth factor with a short in vivo half-life to tissue regeneration, it is necessary to utilize a matrix system to release biologically active growth factor at a >controlled rate over an extended time period. In this study, we compared the in vitro and in vivo release of several growth factors from biodegradable gelatin carriers. Recombinant human growth factors used included basic fibroblast growth factor (bFGF), transforming growth factor beta 1 (TGF), and bone morphogenetic protein 2 (BMP). Hydrogel carriers were prepared through chemical crosslinking of acidic and basic gelatins with isoelectric points (IEPs) of 5.0 and 9.0, respectively. The radioactivity of each radioiodinated growth factor was measured to evaluate its release profile. Basic bFGF and TGF were hardly released from the acidic gelatin hydrogel carrier under the in vitro non-degradation condition, whereas their rapid release was observed from the basic gelatin hydrogel. This could be explained in terms of polyion complexation between the growth factors and gelatin. However, there was no such influence of BMP release on the gelatin IEP. Animal study revealed that the radioactivity of growth factor incorporated in the hydrogel gradually decreased with time. As bFGF-incorporating gelatin hydrogels were degraded in the body, the biodegradation of carrier could be controlled by changing their crosslinking extent. A good correlation of in vivo retention was found between the hydrogel and bFGF. This indicates that bFGF was released with time in vivo, accompanied with hydrogel biodegradation. In addition, we studied the release profile of other growth factors to assess feasibility of our biodegradable carriers for tissue engineering. This work was supported by a grant of ``Research for the Future'' Program from the Japan Society for the Promotion of Science (JSPS-RFTF96I00203) 

11:30 AM CC1.9 
INFLUENCE OF BIOMATERIAL MICROSTRUCTURE ON CYTOKINE SECRETION BY MACROPHAGES. Mary Lee Amirpour, Michael Pishko, Texas A&M University, Department of Chemical Engineering, College Station, TX. 

Neovascularization is critical to the success of many engineered tissues and transplanted cell-containing devices. Macrophages, as part of the host response to an implanted material, play a key role in the regulation of neovascularization through the secretion of cytokines that both promote and suppress angiogenesis. We investigated the relationship between the microstructure of biomaterials and the role of macrophages on the surface of these materials in the secretion of cytokines relevant to neovascularization, such as vascular endothelial growth factor (VEGF), interleukin-1 (IL-1), and interferon- (IFN-). Using a common salt-casting technique, we fabricated polymer scaffolds with different microstructures using biodegradable (polylactide-co-glycolide) and non-degradable (polysulfone) polymeric biomaterials. The microarchitecture of the scaffolds were characterized using electron microscopy and image analysis. Mouse peritoneal macrophages were then cultured on these scaffolds and the media assayed by ELISA for the presence of the secreted cytokines. In addition, we investigated how hypoxia, in combination with material structure, affects secretion of VEGF, IL-1, and INF-. Cell viability on the scaffolds was determined using a chemiluminescent assay and cell morphology was determined by electron microscopy and fluorescence microscopy. Our overall goal is to predict what level of neovascularization will occur for a given biomaterial based on its microstructure. 

11:45 AM CC1.10 
A MODEL FOR OXYGEN TRANSPORT IN MICROENCAPSULATED ISLETS. Hiroo Iwata, Young-Gun Park, Yoshito Ikada, Kyoto Univ, Kyoto, JAPAN. 

Bioartificial pancreas (BAP) is a medical device enclosing islets of Langerhans (islets) into a semi-permeable membrane. The device has been expected to facilitate islets to be transplanted without immunosuppressive therapy. Many research groups have been attracted by microcapsule type BAP, because it provides a safe, simple technique for implanting immuno-isolated islets into various sites of patient's body. The membrane for microcapsulation has to be permeable to small molecules such as oxygen, nutrients, glucose, and insulin, but impermeable to lymphocytes and large molecules such as immunoglobulins and complement proteins. Although numerous studies have been devoted to examine immuno-isolative effectiveness of the microcapsule, little attention has been paid to oxygen supply to islets through the membrane. Since oxygen supply has been reported to be a limiting factor that determines the islet biofunction and survival after transplantation. The oxygen concentration C(r) in the microcapsule was derived as a function of the radial distance using the Fick's diffusion equation in this study. It can be expressed by: C(r)=C(R2)+v/(6D1)*(r2-R22)+v*R13/(3D1)*(1/r-1/R2) (in the islet) C(r)=Co+v/(3D2)*(R23-R13)*(1/R3-1/r) (in the microcapsule) where Co is the oxygen concentration at the transplanted site, D1 and D2 the oxygen diffusion coefficients in the islet and the microcapsule, respectively, v the oxygen consumption rate by unit volume of cells, R1, R2, and R3 the diameter of the necrosis area, the living islet, and the microcapsule, respectively. When Co is 0.47*10-4 mmol/ml and both D1 and D2 are assumed to be 1.4*10-5 cm2/s, and v is 1.2*10-5 mmol/ml/sec, the equations predict that the diameter of the microcapsule should be less than 0.054 cm for the islet (0.025 cm in the average diameter) survival without central necrosis. 

SESSION CC2: CONTROLLING CELL INTERACTIONS WITH BIOMATERIAL CHEMISTRY OR SURFACE PROPERTIES 
Chairs: Kevin E. Healy and Yoshito Ikada 
Monday Afternoon, April 13, 1998 
Pacific A
1:30 PM *CC2.1 
ENGINEERED PROTEIN POLYMER BIOMATERIALS FOR TISSUE ENGINEERING. Joseph Cappello, France Ferrari, Erwin Stedronsky, Protein Polymer Technologies, Inc., San Diego, CA. 

Protein-based materials afford many advantages over synthetic biodegradable polymers in applications of tissue engineering or wound healing. In tissue engineering applications, biomaterials are often intended to be seeded in-vitro with cells and maintained in aqueous culture medium for extended periods of time. During this time polymeric materials sensitive to hydrolysis will begin breaking down and an immediate change in physical and mechanical properties will result. Water stable protein-based materials may avoid physical change during processing while still providing for ultimate resorption when implanted into the body. The degradation products of protein-based implants are natural amino acids and peptides which cause no known toxicity. Additionally, proteins can be engineered to include amino acid sequence components that mediate cell adhesion and other cellular differentiation functions. We describe the physical and biological properties of synthetically engineered protein polymers in forms such as injectable gels, porous sponges, and durable coatings. The protein designs include protein structural elements from natural silk and elastin proteins. Protein polymers can be designed to resorb over various time periods after implantation by adjusting their amino acid sequences. Inclusion of amino acid sequences serving as protease cleavage sites can increase their specific proteolytic degradation by 10 fold. Implant studies have been used to evaluate material biocompatibility, resorption, and cell and tissue integration. 

2:00 PM CC2.2 
MODIFICATION OF SILK FIBROIN FILM WITH A CHIMERA FIBROIN FRAGMENT FOR IMPROVEMENT OF CELL ADHESION. Yasushi Tamada, National Institute of Sericultural and Entomological Science, Tsukuba, Ibaraki, JAPAN. 

The main component of silkworm silk proteins is fibroin which is one of structural proteins like collagen and keratin. The silk fibroin would be suitable as the scaffold for cell culture, transplantation, and tissue engineering because of the mechanical properties. However, the silk fibroin has less function on regulating cell function and tissue development. Immobilization of biological active molecules like proteins and peptides to substrate for cell have been reported as one of the promising techniques to control cell behaviors like cell adhesion, cell growth, cell metabolism, and cell differentiation. Many immobilization techniques have been studied and most of them require to use the cross-linking chemicals which are usually harmful to cells and tissues. Silk fibroin have a characteristic phase transition property by a conformational change of the protein from a random coil to a beta-sheet. During the phase transition, the biological molecules can be stably entrapped into silk fibroin without any chemicals. A new immobilizing technique which is utilized the phase transition mechanism of silk fibroin with a chimera fibroin fragment was designed. The chimera fibroin fragment was constructed by binding a functional peptide to fibroin fragments including crystal regions. This technique consist of immobilizing the fused functional peptide to the crystal region in silk fibroin molecules through co-crystallization during the phase transition process. As a preliminary study, RGD peptide sequence of synthetic oligonucleotides, which is cell adhesion promoting peptide, was fused to fibroin fragment gene by inframe gene fusion technique, and the chimera fibroin (RGD-fibroin) gene was expressed by E.coli expression system. Further, the RGD-fibroin fragment was immobilized to fibroin film by the phase transition technique. In this paper, the construct of the chimera fibroin gene, the expression, and the results of cell adhesion onto the modified fibroin film will be presented. 

2:15 PM CC2.3 
STUDY OF EMPTY SPACES IN COLLAGEN BIOPOLYMER MICROSTRUCTURE BY POSITRON ANNIHILATION LIFETIME MEASUREMENTS. Serge Siles, Laboratoire de Biophysique, Faculte de Medecine, La Timone, Marseille, France; Gerard Moya, Xiao-Hua Li, Denise Siesse-Moya, Laboratoire de Physique des Materiaux, Faculte des Sciences St Jerume, Marseille, FRANCE; Pierre Moser, D.R.F.M.C./SP2M/NM, CEA Grenoble,Grenoble, FRANCE. 

The microstructure of collagen is made of crystalline and amorphous regions which contain empty spaces. The study of these empty spaces is important for the understanding of biomechanisms in proteins. The lifetime measurement in Positron Annihilation Spectroscopy (P.A.S.) is applied to study the free volume collagen properties as a function of concentration in collagen solutions. The lifetimes of positrons are obtained by a conventional fast fast coincidence system. All data are fitted in three components by using the computer programme Positron Fit and resolved. The longest component can be associated with a pick-off annihilation of ortho-positronium trapped in free volumes of amorphous regions. This component varies continuously with the collagen concentration while the average lifetime and the intensity of the longest lifetime remain constant. Such a behaviour has been already observed in some polymers and attests the existence of amorphous areas. Using the formalism of Eldrup, one can deduce the average size of empty spaces present in these amorphous areas. This investigation shows the potential of P.A.S. in the study of biopolymers microstructure 

2:30 PM CC2.4 
PEG CROSS-LINKED ALGINATE HYDROGELS WITH CONTROLLED MECHANICAL PROPERTIES. Petra Eiselt, Jon A. Rowley, David J. Mooney, University of Michigan, Depts of Chemical, Biomedical, and Biological and Materials Science Engineering, Ann Arbor, MI. 

Reconstruction of tissues and organs utilizing cell transplantation offers an attractive approach for the treatment of patients suffering from organ failure or loss. Highly porous synthetic materials are often used to mimic the function of the extracellular matrix (ECM) in tissue engineering, and serve as a cell delivery vehicle for the formation of tissues in vivo. Alginate, a linear copolysaccharide comprised of D-mannuronic acid (M-block) and L-guluronic acid (G-block) units is widely used as a cell transplantation matrix. Alginate is generally considered to be biocompatible, and hydrogels are formed in the presence of divalent cations such as Ca2+, Ba2+ and Sr2+. However, ionically cross-linked alginate gels continuously lose their mechanical properties over time with uncontrollable degradation behavior. We have modified alginate via covalent coupling of cross-linking molecules to expand and stabilize the mechanical and swelling property ranges for these gels. Several amino terminated PEG molecules of varying molecular weight (200, 400, 1000, 3400) were synthesized utilizing carbodiimide chemistry. They were covalently coupled to alginate, and mechanical properties of the resulting hydrogels were determined. The elastic modulus of the cross-linked alginates depended on the molecular weight of the cross-linking molecules, and ranged from 10 - 110 kPa. The cross-link density in the hydrogels was also varied from 3 to 47 % (relative to the carboxylic groups in the alginate) and the mechanical properties were measured. The elastic modulus increased gradually and reached a maximum at a cross-link density of 15 %. Further increases in cross-link density led to a decline in modulus. We believe this decrease can be attributed to an increase in single end anchorage, and the corresponding decrease of intermolecular cross-link reactions. In summary, covalently coupled hydrogels can be synthesized which exhibit a wide range of mechanical properties, and these materials may be useful in a number of tissue engineering applications. This work was sponsored by Reprogenesis. 

2:45 PM *CC2.5 
CONSTRUCTION OF BIOMIMETIC ENVIRONMENTS WITH A SYNTHETIC PRPTIDE ANALOGUE OF COLLAGEN. Raj Bhatnagar, Jing Jing Qian, Anna Wedrychowska, and Nancy Smith, University of California, San Francisco, CA. 

The flow of chemical and mechanical signals among cells, and between cells and their environment plays a crucial role in cell differentiation and morphogenesis. Biomimetic environments must take into account the need of cells to receive both biochemical and mechanical stimuli. Collagen mediates the flux of many of these signals via highly specific receptors on cells and acts synergistically with growth factors. We identified a cell binding domain (766GTPGPQGIAGQRGVV780) in -1(I) chain of collagen.