转化生长因子β3基因转染兔骨髓间充质干细胞诱导其向软骨表型的分化

背景：内源性诱导软骨分为就是通过一定的载体将目的基因整合入干细胞内,使其自行分泌诱导因子诱导自身进行分化。目的：观察将转化生长因子β3通过腺相关病毒载体转染诱导兔骨髓间充质干细胞向软骨表型转化的能力。方法：取体外培养的第3代骨髓间充质干细胞进行重组腺相关病毒转染,将转染后3,6,9,12d细胞裂解提取蛋白进行酶联免疫检测目的蛋白转化生长因子β3的体外表达。RT-PCR,免疫印迹westernblot分别从基因和蛋白水平上检测1,2周Ⅱ型胶原的表达,甲苯胺蓝染色检测1,2周蛋白多糖的表达。结果与结论：重组腺相关病毒转染后,骨髓间充质干细胞可以较稳定的表达目的蛋白转化生长因子β3,并且转染成功的骨髓间充质干细胞较阴性对照组能够更好的向软骨表型转化。证实转化生长因子β3可以腺相关病毒为载体转染骨髓间充质干细胞并诱导其向软骨表型分化。

Differentiation of rabbit mesenchymal stem cells into cartilage phenotype induced by transforming growth factor beta 3

BACKGROUND：Endogenous induction of cartilage differentiation is to use a certain carrier to integrate the target genes into stem cells to induce self-differentiation by self-induced secretion of cytokines.OBJECTIVE：To observe the ability of rabbit bone marrow mesenchymal stem cells（BMSCs） transfected with transforming growth factor beta 3（TGF-β3） to differentiate into cartilage phenotype through adeno-associated virus vector.METHODS：The third passage BMSCs in logarithmic growth phase was transfected by rAAV2-TGF-β3.Total protein was extracted from cells at 3,6,9,12 days after transfection,and TGF-β3 was detected through ELISA.After 2 weeks of culturing,mRNA expressions of type Ⅱ collagen were determined by RT-PCR and the collagen Ⅱ protein was detected by western blot.The expression of proteoglycan was shown by toluidine blue staining.RESULTS AND CONCLUSION：After transfection,BMSCs can express TGF-β3 stably,and make a better cartilage transformation compared to negative control group.TGF-β3 can be transfected into BMSCs through adeno-associated virus vectors and induce cell differentiation into cartilage phenotype.