Cell-Responsive Synthetic Biomaterials Formed in situ

Citation

Pratt, Alison Beth (2001) Cell-Responsive Synthetic Biomaterials Formed in situ. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/HFEF-7F61. https://resolver.caltech.edu/CaltechETD:etd-08132007-143014

Abstract

Two-way communication between cells and their extracellular matrices plays an important role in establishing and maintaining cell and tissue morphology. Synthetic materials for tissue engineering have so far not been able to mimic these interactions. This work describes a new class of synthetic biomaterials that display the essential characteristics of naturally occurring matrices. The materials can be formed in contact with cells and tissues to create covalently cross-linked hydrogels which are effectively nonporous on the length scale of a cell process. The otherwise nonadhesive gels can be rendered specifically cell-adhesive by incorporation of ligands for cell surface adhesion molecules, such as integrins. The materials are degradable by plasmin, a proteolytic enzyme used by cells during proliferation and migration in natural matrices. Furthermore, the materials can be designed to sequester heparin-binding growth factors.

The materials consist primarily of poly(ethylene glycol) which is cross-linked in situ by selective conjugate addition reactions of its termini, activated with conjugate acceptors such as vinylsulfones, with peptides containing three or more cysteine residues in the form of reduced thiols. The cross-linking peptides are designed such that they are hydrolyzed by plasmin. A simple cell migration assay was developed to evaluate the material design parameters. Human fibroblasts were observed to migrate out of microliter fibrin clots into the synthetic materials in a manner dependent on the type of material and the culture conditions. Inhibiting fibrinolysis with a plasmin-insensitive substrate or with a plasmin inhibitor, [epsilon]-amino-n-caproic acid, inhibited migration. Adding growth factors, FGF-2 or PDGF-BB, that upregulate plasminogen activation increased the rate of cell migration. Decreasing the amount of RGD within the materials decreased the rate and extent of cell migration in a concentration-dependent manner. Replacing the RGD peptide with an RDG peptide decreased the amount of outgrowth.

In a critical size calvarial defect in the rat, the synthetic materials were formed in the presence of an osteoinductive protein, rhBMP-2, and supported de novo bone formation. Materials containing a heparin-based growth factor delivery system promoted significantly more bone than materials without the delivery system. After three weeks, 94% of the defect area was covered with new bone, and the opacity of the new bone was 84% of that of neighboring, uninjured bone.

Thesis Files