关键词: 滑膜间质干细胞, 腺病毒, 骨形态发生蛋白, 绿色荧光蛋白

Abstract: Objective To study the feasibility of adenovirus-mediated BMP-4 transfected rabbit synovial mesenchymal stem cells (SMSCs) with green fluorescent protein (EGFP) gene into chondrocytes. Method The rabbit knee joint capsule was taken, and the type I collagenase was used to digest and separate the SMSCs. The surface markers were detected by flow cytometry. The recombinant adenovirus (Ad-BMP-4) containing BMP-4 gene and EGFP gene was transfected into the SMSCs by different multiplicity of infection (50,100,200,300,400,500 MOI respectively) Efficiency, find the most suitable transfection conditions, and test the cell growth curve under different transfection conditions. The EGFP-labeled cells were observed by ELISA to determine if the transfected SMSCs entered the chondrocyte differentiation lineage. Result The morphology of SMSCs obtained after isolation and purification showed the proper structure of mesenchymal stem cells. The surface markers highly expressed CD45 and CD90, and basically did not express CD34 and CD45, which was consistent with many features of SMSCs. The transfection efficiencies were 54.3%, 72.2%, 77.8%, 85%, 86.6%, 89.3% after transfection with MOI of 50,100,200,300,400,500, respectively, and the fluorescence microscopy was more and more High green fluorescence expression, MOI=300 for transfection suitable conditions. The result of ELISA showed that BMP-4 was highly expressed, and cartilage genes such as SOX9, COLⅡ and AGC protein were also highly expressed. Conclusion This study shows that recombinant adenovirus transfected rabbit SMSCs, MOI amount of 300 can quickly and efficiently express BMP-4 and activate other cartilage-related genes, prompting SMSCs to chondrocytes.

Key words: Synovial mesenchymal stem cells , Adenovirus； Bone morphogenetic protein, Green fluorescent protein