负载SOX-9的大鼠脂肪干细胞成软骨分化的实验研究

目的 SOX-9基因转染大鼠的ADSCs(Adipose stromal cells,ADSCs),诱导其向软骨细胞分化,为软骨组织工程学种子细胞提供新方法.方法 分离、培养大鼠的ADSCs,免疫荧光法进行鉴定;SOX-9基因转染ADSCs,对转染后细胞进行筛选;MTT法测定筛选后细胞的增殖活性;RT-PCR、Western blot对筛选后细胞的表达进行检测.结果 成功分离、培养大鼠的ADSCs;细胞表面标志物CD29和CD44表达阳性,CD106和CD34表达阴性;基因转染后细胞的增殖力变强;转染SOX-9的ADSCs在目的基因SOX9、Aggrecan、Collagen Ⅱ mRNA的表达和软骨特异性基质-Collagen Ⅱ的分泌上明显高于转染空载体组和对照组;结论 SOX-9基因转染后ADSCs具有了软骨细胞的表型特征,可以用作软骨组织工程的种子细胞.

An experimental study on differentiation into chondrocytes of rat's ADSCs transfected by SOX-9 gene

Objective To transfect rats ADSCs with SOX-9 gene and induce their differentiate into chondrocytes, providing a new method for the tissue-engineering cartilage seed cells. Methods Isolated and cultured the rats ADSCs, then identificated the cells by immune fluorescence assay. Transfected the cells with SOX-9, and selected the cells with transgene. MTT was used to detect the proliferation of the selected cells. Meanwhile, RT-PCR, and Western blot were used to analysis the expression level of the selected cells. Results We succeed in separating and culturing rats ADSCs. The cell surface markers CD29 andCD44 were positive, whereas CD106 and CD34 were negative. The prolifetation ability became stronger after transfectd with SOX-9 gene. The transfected group had higher destined gene SOX9、Aggrecan、Collagen Ⅱ mRNA expression and cartilage specific matrix-Collagen Ⅱ secretion than the vector group and control group. Conclusions Transfected ADSCs by SOX-9 gene have the pheno-types of chondrocytes,and can be used as the cartilage tissue-engineering seed sells.