Collagen/silk Fibroin Composite Membrane Incorporating Growth Factor Was Prepared For Cartilage Tissue Engineering

The damage of articular cartilage caused by bone tumors and sports injury is increasing year by year,which is a great threat to human health.Emergence of tissue engineering has brought new technology for the treatment of bone defects.Study of biomaterials with excellent biological activity and biodegradablity makes disease tissue regeneration and repairing defects feasible.Especially,construction of temporospatial release control materials incorporating growth factors nanoparticles is a potential field.In this work,we developed a double layered collagen/silk fibroin(COL/SF)composite incorporating growth factors nanoparticles and implemented the spatiotemporal control release according topology difference,Then this composite was implanted into animal models to investigate the feasibility in repairing cartilage defects.The contents and results of this study are as follows:(1)Preparation of collagen/silk fibroin composite incorporating growth factors nanoparticles ingredient and collagen was extract from abandoned skin,SF was isolated from silkworm cocoons.A double-layered collagen/silk fibroin composite consisting of dense layer and loose layer was prepared by admixing and crosslinking.By determining its physical and chemical properties,optimal proportion of collagen/silk fibroin composite,optimal proportions of collagen and silk fibroin was 7:3 and 3:7 which possesses best hydrophilism,interval porosity and biocompatibility.(2)Preparation of collagen/silk fibroin composite incorporating growth factors and study on in vitro release kinetics.Nanoparticles were prepared,including PLGA microballoons,Polylysine-Heparin(PLL-Hep)and Chitosan-Heparin(Ch-Hep)nanoparticles.Microballoons or nanoparticles with optimal proportion was screeninged by determining size,Zate and encapsulation efficiency,then adsorbed into collagen/silk fibroin composite to prepare double-layered collagen/silk fibroin composite incorporating growth factors.Meanwhile,Flow cell technique was used to examine its release-dynamics in vitro.The results showed that PLL-Hep nanoparticles were best bearer for TGF-?1,what's more different release rates revealed that the release profiles of TGF-?1 obeyed a spatiotemporal model,which displayed a clear correlation with topology of composite membrane.(3)Adhesion and proliferation of BMSCs in collagen/silk fibroin composite incorporating growth factors.The BMSCs were isolated from bone marrow of rat tibias and femurs and the adhesion and proliferation behavior on incorporating TGF-?1 composite membranes was studied.The results confirmed that collagen/silk fibroin composite had favourable biocompatibility,BMSCs proliferated well on both dense and loose layers,while loose layer was better,These findings indicated that cells adhesion and proliferation were affected by TGF-?1 and the inhomogeneity of structure of double-layered collagen/silk fibroin composite,which is related to the spatiotemporal control release of growth factor.(4)A rabbit cartilage defect model was established.A double-layered composite with TGF-?1 was implanted into full-thickness articular cartilage defects in rabbits in accordance with its loose or dense layer down towards defects.The repair effect of cartilage defects were evaluated by general observation,histologic staining and histological scores.As a result,this composite did well in the repair of cartilage defects.While loose layer towards cartilage surface,the defects were almost completely repaired after 12 weeks,which indicated that the structure and spatiotemporal control release of growth factor of collagen/silk fibroin composite were favourable to articular cartilage repair.