Repair Of Rabbit Articular Cartilage With Synovial Mesenchymal Stem Cells Combined With Allogeneic Bone

Objective:Designed to establish a model of knee articular cartilage defects in New Zealand rabbits,allogeneic bone scaffold materials prepared by deep cryogenic technology are combined with BMP-4 and SMSCs to achieve effective repair of articular cartilage injury,thereby revealing the law of cartilage repair,in order to provide new ideas for cartilage damage repair.Methods:1.Rabbit SMSCs obtained and prepared: New Zealand rabbit knee joint cavity was taken,and the SMSCs were obtained by digestion with type I collagenase,and the markers were detected by flow cytometry.The gradient MOI virus(Ad-BMP-4)was used to infect the SMSCs,the transfection efficiency and cell growth curve were detected,and the expression of SMSCs-related proteins was detected by ELISA.2.Construction and treatment of allogeneic bone grafts: Using a transverse nail sleeve to drill 36 cartilage columns in the medial malleolus of 18 adult New Zealand rabbits,the cartilage column was gradually cooled down and placed in a liquid nitrogen tank for 3 weeks.3.Model building and processing: 24 adult New Zealand rabbits were randomly divided into 4 groups,a full thickness articular cartilage defect model with a diameter of 4 mm and a depth greater than 4 mm was created on the femoral condyle surface using a transverse nail sleeve.Group A is a simple cartilage defect model,group B uses simple allogeneic bone repair,group C used SMSCs to repair allogeneic bone repair,group D was treated with BMP-4 transfected SMSCs in combination with allogeneic bone repair.4.Evaluation of the repair effect: Four groups of animals were taken at 4,8,and 16 weeks postoperatively.Pathological staining was performed after the specimen was fixedly sliced.The wakitani histology scoring system was used to comprehensively assess the defect repair status from the repaired tissue cell types,matrix staining,surface smoothing,cartilage thickness,and integration with surrounding normal cartilage tissue.Results:1.The obtained morphology of SMSCs showed the proper structure of mesenchymal stem cells.The surface markers of SMMSCs were highly expressed on CD44 and CD90,and CD34 was not expressed at all,which was consistent with the characteristics of SMSCs.The infection efficiency depends on the MOI level,and 300 MOI is chosen as the appropriate infection titer.ELISA detection of transfected SMSCs cells can maintain the normal proliferation of cells,and can rapidly and efficiently express BMP-4,activate other cartilage-related genes Sox9,COL II and other expression,and promote the differentiation of SMSCs into chondrocytes.2.Transfected SMSCs combined with allogeneic bone were used to repair rabbit cartilage injury.The experimental group repaired the flatness of the tissue at each time point and integrated with the surrounding cartilage better than other control groups.In the experimental group,HE and Safranin O showed increased extracellular matrix and chondrocyte depression.Immunohistochemistry showed strong expression of type II collagen.Conclusion:Adenovirus carrying BMP-4 gene plays an important role in the differentiation of SMSCs into chondrocytes after successful infection of SMSCs,and has the ability to promote the differentiation of SMSCs into cartilage.The combination of BMSCs transfected with SMSC gene can promote the repair of rabbit knee articular cartilage injury,and provide a new idea for the experimental study of cartilage tissue engineering repair of SMSCs composite carrier in the future.