| PurposeIn the first part of the study,the autologous seed cells most suitable for cartilage regeneration and repair were selected by comparing the proliferation ability and cartilage ability of mesenchymal stem cells derived from the tissues around the knee joint.In the second part,the articular chondrocytes were co-cultured with the most suitable mesenchymal stem cells to determine the effect of chondrocytes on mesenchymal stem cells and the specific mechanism of influence.In the third part,the first two parts of the res?lts were combined to determine the most suitable tissue engineering program for articular cartilage regeneration and repair.MethodIn the first part of the study,the infrapatellar fat pad mesenchymal stem cells,subcutaneous adipose-derived mesenchymal stem cells,bone marrow mesenchymal stem cells,synovial mesenchymal stem cells were obtained,and the proliferation ability,three-way differentiation ability and induction three-way were compared.Differentiation ability,the optimal mesenchymal stem cells with stronger ability to differentiate into cartilage were used as seed cells for tissue engineering of articular cartilage regeneration.In the second part,the articular chondrocytes were co-cultured with the most suitable mesenchymal stem cells to identify the effect of co-culture on the phenotype of mesenchymal stem cells,and the main mechanism(exosome or soluble secretory factor)that affects the mesenchymal stem cell phenotype by screening chondrocytes.In the third part,New Zealand white rabbits were selected as experimental animals to establish an animal model of knee joint cartilage injury.Combined with the res?lts of the first two parts,based on the PLGA-HA gradient composite scaffold,combined with different treatment combinations,the general recovery and pathological res?lts were clearly promoted.The best tissue-general treatment for articular cartilage regeneration was clewered.Res?ltsIn the first part of the study,the infrapatellar fat pad mesenchymal stem cells,the subcutaneous adipose-derived mesenchymal stem cells,the bone marrow mesenchymal stem cells,the synovial mesenchymal stem cells were obtained successf?lly,and the proliferation experiments showed that the proliferative capacity of the infrapatellar fat pad mesenchymal stem cells and synovial mesenchymal stem cells were superior to that of the other two mesenchymal stem cells(P<0.05),and the genes of cartilage differentiation-specific markers and the expression of protein of the infrapatellar fat pad mesenchymal stem cells and the synovial mesenchymal stem cells were significantly higher than that of the other two mesenchymal stem cells(P<0.05).Therefore,the infrapatellar fat pad mesenchymal stem cells and the synovial mesenchymal stem cells have the potential to be the excellent seed cells in repairing of articular cartilage,considering in the clinical the infrapatellar fat pad was easier to obtain than the synovial tissue,so the infrapatellar fat pad mesenchymal stem cells were selected as the key cells for subsequent research.In the second part,the chondrocytes was co-cultured with the infrapatellar fat pad mesenchymal stem cells,and co-cultured with the blank group.The comparisons showed that the chondrocytes co?ld significantly change the phenotype of the infrapatellar fat pad mesenchymal stem cells.Although the proliferative capacity of the infrapatellar fat pad mesenchymal stem cells did not change significantly(P>0.05),the expression of specific markers of cartilage differentiation of Col-2 and Sox-9 was significantly increased(P<0.05).Moreover,the gene expression levels of Oct-4 and Nucleostemin were significantly decreased(P<0.05),indicating that chondrocytes significantly promoted the differentiation of subgingival fat pad mesenchymal stem cells into cartilage.In order to clarify the mechanism of chondrocytes reg?lating the subgingival fat pad mesenchymal stem cells,this study obtained exosomes secreted by chondrocytes and secreted soluble cytokines,which act on subgingival fat pad mesenchymal stem cells,respectively,and then pass RT-PCR and Western Blot were used to detect the phenotype of subgingival fat pad mesenchymal stem cells.The res?lts showed that the phenotype of subgingival fat pad mesenchymal stem cells in the chondrocyte exosome intervention group was associated with chondrocyte/axillary fat pad mesenchymal stem cells.The res?lts after the c?ltivation were similar.Therefore,it can be determined that chondrocytes can reg?late cartilage differentiation of the infrapatellar fat pad mesenchymal stem cells through exosomes.In the third part,based on the res?lts of the first two parts,based on the PLGA-HA gradient composite scaffold,set up four treatment combination group: group A-simple stent group,group B-stent + chondrocyte exosomes group,group C-stent + The infrapatellar fat pad mesenchymal stem cells group,group D-scaffold + the infrapatellar fat pad mesenchymal stem cells + chondrocyte exosomes group.Tissue samples were obtained at 4 weeks,8 weeks,and 12 weeks after surgery,and the effects were repaired by gross view,H&E staining,and toluidine blue staining.The res?lts showed that the repair effect of articular cartilage injury in group D was the best.The effect of group C was slightly worse than that of group D,but it was better than that of group A and group B.The repair effect of group A and group B was similar,and the repair effects on articular cartilage injury were similar but both no ideal.Therefore,PLGA-HA gradient composite scaffold combined with articular cartilage exosomes and the infrapatellar fat pad mesenchymal stem cells combined with repair of articular cartilage injury is the best tissue process repair program in this study.ConclusionThrough the research of the three parts of this subject,the excellent cartilage ability of the infrapatellar fat pad mesenchymal stem cells is cleared,which is the ideal seed cells for the regeneration of articular cartilage in the future;and the articular cartilage can promote the infrapatellar fat pad by secreting exosomes.The mesenchymal stem cells differentiate into cartilage,which may be one of the possible reasons for the simple joint cavity injection of mesenchymal stem cells to promote the repair of articular cartilage injury.In addition,the final res?lts of this study suggest that PLGA-HA gradient composite scaffold combined with articular cartilage excretion body and the infrapatellar fat pad mesenchymal stem cells combined with repair of articular cartilage injury can obtain better curative effect,and further explore its clinical transformation value in the future,and provide a new treatment idea for clinical treatment of articular cartilage injury. |
PDF Full Text Request