Preparation And Characterization Of Polypeptide-Modified Gradient 3D Composite Fibrous Scaffold For Osteochondral Repair

Injured articular cartilage can hardly be self-repaired,because it is an avascular,aneural and alymphatic connective tissue.At present,the commonly used methods for the treatment of articular cartilage defect include microfracture technology and autologous chondrocyte transplantation technology,and so on.These technologies can improve the joint function to a certain extent,but the long-term treatment effect is not ideal.As for large-scale cartilage defects,especially those cases having both cartilage and subchondral bone injuries,there is no an ideal clinical treatment to achieve satisfactory tissue regeneration.The strategy of scaffold-based osteochondral tissue engineering is deemed as a feasible and promsing way to face the challenges.As well known,articular cartilage demonstrates heterogeneous compositions perpendicularly,in brief,consisting of a superficial cartilaginous layer,a transitional calcified cartilage interlayer,and the subchondral bone layer,accordingly,biomimetic gradient scaffolds are essential for osteochondral regeneration,whereas,the current approaches are still not so satisfactory and more attempts are needed.In this dissertation,electrospun composite nanofibers were applied to fabricate the gradient 3D scaffold via freeze-drying fiber suspension and subsequent thermal crosslinking.The nanofibrous morphology would resemble the collagen fibrous network in natural extracellular matrix.Electrospun composite nanofibers were made of poly(L-lactide)(PLLA)and gelatin(Gel),combining the advantages of both natural and synthetic polymer.In producing the scaffold for the cartilage layer,glycosaminoglycans were introduced into the composite nanofibers.In producing the scaffolds for the intermediate calcified cartilage layer and the subchondral bone,the PLLA/Gel fibrous scaffolds were soaked in simulated body fluid for different periods to control the deposition amounts of hydroxyapatite.Then,the three layers of scaffolds were assembled into gradient scaffolds to match the heterogeneous transition in natural osteochondral tissue in terms of both nanofibrous structure and chemical compositions.To improve the cell recruitment ability of the scaffold in vivo,a kind of cell affinity polypeptide E7 was mounted onto the scaffold surface.By means of SEM observation,compression performance test,TGA measurement and degradation study,the physical and chemical properties of the prepared scaffolds were comparatively explored,and the optimized parameters for scaffold preparation were determined.Then,the biocompatibility of the scaffolds was confirmed from assays on cell viability(proliferation,dead/live staining,morphology)by incubation of bone marrow mesenchymal stromal cells(BMSCs)onto the scaffolds.The effects of different scaffolds on the chondrogenic and osteogenic differentiation of BMSCs were evaluated by ELISA assays.Finally,the gradient 3D fibrous scaffolds were implanted into the knee osteochondral defect model of New Zealand white rabbits to conduct in vivo evaluations on osteochondral regeneration.According to the findings obtained from biomechanical assessment,magnetic resonance imaging(MRI),micro computed tomography(Micro-CT)reconstruction,and so on,the gradient 3D scaffold modified with E7 polypeptide could effectively promote the regeneration of osteochondral defects compared to those scaffolds without E7 coating.In summary,a properly designed and fabricated gradient 3D nanofibrous scaffold can a promising substrate to induce the regeneration of osteochondral defect via the tissue engineering strategy.