Optimal Preparation Of Decellularized Tissue-Engineered Hyaline Cartilage Scaffolds And Its Use In The Repair Of Articular Cartilage Defects

Articular cartilage defect is one of main reasons that caused osteoarthritis.Due to the lack of blood vessels,lymphatic vessels,nerves,as well as the limited chondrocytes in the cartilage,it is difficult to heal itself once articular cartilage is damaged.Therefore,repairing cartilage defects has become an urgent issue in the treatment of osteoarthritis.Traditional clinical methods,such as microfracture,mosaicplasty,and autologous chondrocyte transplantation,cannot effectively solve the problem of cartilage repair.One issue is that the repaired tissue is not pure hyaline cartilage,and secondly,the repaired tissue cannot integrate well with the surrounding natural cartilage.Therefore,this article uses tissue engineering methods to prepare decellularized tissue-engineered hyaline cartilage scaffold.This scaffold has a high-purity hyaline cartilage phenotype and a soft,porous sponge-like structure,which is very beneficial to cartilage repair.Furthermore,further studies on recellularization and cross-linking modification of the scaffold have been conducted to increase its advantages as cartilage graft and provide a new feasible solution for achieving high-quality repair of joint cartilage defects in clinical applications.Based on the above design,this article carried out research in the following several parts.In the first part,different decellularization methods were designed to prepare dLhCG(decellularized Living hyaline Cartilage Graft)from LhCG(Living hyaline Cartilage Graft).The optimal decellularization method(Method Triton+NaOH)was selected to prepare dLhCG.Furthermore,human chondrocytes were seeded onto dLhCG for recellularization,and it was observed that human chondrocytes adhered and proliferated significantly without dedifferentiation in Recellularized dLhCG(RdLhCG).This confirmed the advantages of dLhCG as a cartilage scaffold and the potential of R-dLhCG as a new generation of cartilage grafts.In the second part,rabbit-derived R-dLhCG was constructed by repopulating autologous chondrocytes to explore the interactions between cells and scaffolds in RdLhCG,including cell proliferation,phenotype changes,and matrix secretion.Furthermore,by constructing a rabbit knee patellar cartilage full-layer defect model,autologous R-dLhCG were used as grafts to treat defects,and the actual results of cartilage repair were verified.The results demonstrated that R-dLhCG possess great advantages for cartilage repair,forming well-integrated hyaline cartilage and integrating well with surrounding natural cartilage,showing great potential for clinical applications.Finally,based on the first part,it was observed that dLhCG inevitably suffers from the loss of GAG(glycosaminoglycan)and decreased physical mechanical properties during decellularization.Hence,NHS/EDC crosslinking agents were used to crosslink dLhCG with chondroitin sulfate(CS)to optimize its mechanical properties and prevent the loss of glycosaminoglycan.By adjusting the concentration of the crosslinking agent,the degree of crosslinking,component content,mechanical properties,and tissue structure changes in dLhCG/CS scaffolds were investigated.The results showed that dLhCG/CS scaffolds could effectively restore the loss of glycosaminoglycan and improve the mechanical properties of the scaffold through crosslinking with CS.However,with increasing concentrations of the crosslinking agent(30mM and above),the scaffold also displayed adverse effects of crosslinking toxicity.Therefore,the concentration of crosslinking agent needs to be controlled within an appropriate range.