Disulfide-crosslinked hyaluronan-gelatin hydrogel films: a covalent mimic of the extracellular matrix for in vitro cell growth

• Published in: Biomaterials 2003-09, Vol.24 (21), p.3825-3834
• Main Authors: Shu, Xiao Zheng, Liu, Yanchun, Palumbo, Fabio, Prestwich, Glenn D
• Format: Article
• Language: English
• Subjects: 3T3 Cells; Animals; Biocompatible Materials - chemistry; Biomaterial; Cell Adhesion; Cell Division; Cell growth; Cell Survival; Cross-Linking Reagents - pharmacology; Disulfide; Disulfides - chemistry; Dithiothreitol - pharmacology; Extracellular Matrix - metabolism; Fibroblasts - metabolism; Gelatin; Gelatin - chemistry; Hyaluronic acid; Hyaluronic Acid - chemistry; Hyaluronoglucosaminidase - chemistry; Hydrogel, Polyethylene Glycol Dimethacrylate - chemistry; Hydrogels; In Vitro Techniques; Magnetic Resonance Spectroscopy; Methylgalactosides - chemistry; Mice; Mice, Inbred BALB C; Models, Chemical; Polystyrenes - chemistry; Sodium Chloride - chemistry; Sulfhydryl Compounds; Time Factors
• ISSN: 0142-9612; 1878-5905
• DOI: 10.1016/S0142-9612(03)00267-9

A new disulfide crosslinking method was developed for the preparation of blended hyaluronan (HA)–gelatin hydrogels to form a synthetic, covalently linked mimic of the extracellular matrix (ECM). The HA and gelatin were chemically modified using 3,3′-dithiobis(propionic hydrazide) (DTP). After reduction with dithiothreitol (DTT), the thiol derivatives of HA (HA-DTPH) and gelatin (gelatin-DTPH) were obtained and characterized. To minimize interference with biological function, the degree of substitution of HA-DTPH and gelatin-DTPH was kept below 50%. Solutions of HA-DTPH and gelatin-DTPH in varying blends (20%, 40%, 60%, 80% gelatin) were prepared in 1% w/v NaCl and crosslinked by disulfide bond formation in air. Hydrogel films were dried and further crosslinked with dilute hydrogen peroxide. Disulfide crosslinked HA-DTPH, gelatin-DTPH, and blends thereof, were degradable enzymatically by collagenase and by hyaluronidase (HAse). The rapid digestion of the crosslinked 100% gelatin-DTPH film by collagenase was significantly retarded by the presence of 20% or 40% HA-DTPH. Addition of at least 40% w/v gelatin into the 100% HA-DTPH films significantly improved the attachment and spreading of Balb/c 3T3 murine fibroblasts seeded on the surface of the hydrogel. These results demonstrate that disulfide-crosslinked HA–gelatin hydrogels, a new type of covalent synthetic ECM, constitute biocompatible and biodegradable substrata for cell culture in vitro.