慢病毒介导TGF-β3/BMP-2联合基因转染诱导兔骨髓间充质干细胞成软骨细胞分化  

Combination gene transfection of TGF-β3 and BMP-2 mediated by lentivirus induce chondrogenic differentiation of rabbit bone marrow mesenchymal stem cells

作　　者：周瑜[1] 夏长所[1] 王昌耀[1] 王敏[2] 迟静薇[3] 王英振[1] 

机构地区：[1]青岛大学医学院附属医院东院区关节外科,山东省266100 [2]青岛大学医学院附属医院黄岛院区麻醉科 [3]青岛大学医学院附属医院黄岛院区中心实验室

出　　处：《中华临床医师杂志（电子版）》2014年第4期90-94,共5页Chinese Journal of Clinicians(Electronic Edition)

摘　　要：目的重组慢病毒介导TGF-β3/BMP-2联合基因转染兔骨髓间充质干细胞,诱导其软骨细胞分化,与TGF-β3单基因转染诱导其软骨细胞分化的效率作比较。方法全骨髓贴壁法分离、培养骨髓间充质干细胞,流式细胞术检测细胞表面抗原,培养后通过重组慢病毒分别转染阴性对照(N组)、TGF-β3单基因(T组)、TGF-β3/BMP-2联合基因(TB组),培养14 d后,荧光显微镜下观察各组细胞形态,RT-PCR、Western blot分别检测SOX-9、蛋白多糖、Ⅱ型胶原在基因水平与蛋白水平的表达量。结果流式细胞术检测造血干细胞标志物(CD34,2.07%)、白细胞表面抗原(HLA-DR,1.73%)均为阴性,透明质酸受体(CD44,82.79%)为阳性,荧光显微镜观察诱导后细胞形态多为多角形、方形或者类圆盘状,RT-PCR、Western blot结果均示T组、TB组SOX-9、蛋白多糖、Ⅱ型胶原表达量均比空白对照组(C组)、N组高(P<0.05),且TB组表达量高于T组(P<0.01)。结论 TGF-β3/BMP-2联合基因转染骨髓间充质干细胞能诱导其成软骨细胞分化,且联合基因转染效率高于TGF-β3单基因转染。Objective Combination gene transfection of TGF-β3/BMP-2 mediated by recombinant lentivirus induce chondrogenic differentiation of rabbit bone marrow mesenchymal stem cells（BMSCs）, and compared with TGF-β3 single gene transfecting. Methods BMSCs were isolated and cultured using a whole bone marrow adherent method, then detected cells＇ surface antigen, respectively transferred negative control（N group）, TGF-β3 single gene（T group）, TGF-β3/BMP-2 combination gene（TB group）, observed cell morphology of each group, the expression of SOX-9, type Ⅱcollagen（COL2）, aggrecan（ACAN）. Results The BMSCs revealed uniformly negative for the early hematopoietic stem cell marker（CD34, 2.07%） and the leucocytes surface antigen（HLA-DR, 1.73%）, positively for the hyaluronan receptor（CD44, 82.79%）. The differentiated cells took a shape of polygonal, square or disc-like, the expression of chondrogenic related markers such as sox-9, COL2 and ACAN were upregulated in T group and TB group than in C group and N group（P0.05）, furthermore, those markers were synthetized significantly more in TB group than in T group（P0.01）. Conclusion These findings indicated that BMSCs differentiated into chondrocytes induced by TGF-β3/BMP-2 combination gene, this method was superior to TGF-β3 single gene transfection.

关 键 词：间质干细胞 转化生长因子Β3 骨形态发生蛋白质类 软骨分化 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]