共培养诱导人脐血间充质干细胞向软骨细胞分化  

Chondrogenic differentiation of co-cultured human umbilical cord blood-derived mesenchymal stem cells

作　　者：郑朋飞[1] 陈雷[1] 董展[1] 蒋立[1] 鞠黎[1] 王儒法[1] 楼跃[1] 

机构地区：[1]南京医科大学附属南京儿童医院,江苏省南京市210008

出　　处：《中国组织工程研究》2013年第23期4196-4203,共8页Chinese Journal of Tissue Engineering Research

基　　金：2010南京市卫生局课题(QYK10163)~~

摘　　要：背景:研究发现共培养可诱导人源干细胞向特定细胞分化,但如何控制两种细胞间比例以最大效率获得目的细胞是研究中的难题。目的:观察人脐血间充质干细胞与兔软骨细胞按不同比例共培养后向软骨细胞分化的情况。方法:复苏人脐血间充质干细胞并进行传代,将经流式细胞仪鉴定的人脐血间充质干细胞与体外分离的兔软骨细胞按照2:1或3:1的比例共培养,并用100μg/L的胰岛素样生长因子1进行诱导。在共培养14d,提取细胞RNA和蛋白质,实时定量PCR检测聚集蛋白多糖和Ⅱ型胶原mRNA的表达,Western blot检测聚集蛋白多糖和Ⅱ型胶原蛋白的表达。结果与结论:人脐血间充质干细胞与兔软骨细胞共培养14d,共培养的细胞聚集蛋白多糖和Ⅱ型胶原mRNA及蛋白的表达高于单纯胰岛素样生长因子1诱导的细胞。在人脐血间充质干细胞与软骨细胞比例相同时,加入胰岛素样生长因子1可提高细胞聚集蛋白多糖和Ⅱ型胶原mRNA及蛋白的表达。同时人脐血间充质干细胞与兔软骨细胞按3:1共培养更能促进细胞Ⅱ型胶原mRNA和蛋白及聚集蛋白多糖蛋白的表达;而按2:1共培养时,细胞聚集蛋白多糖mRNA表达更多。可见人脐血间充质干细胞与兔软骨细胞共培养后可诱导人脐血间充质干细胞向软骨细胞分化,且按3:1的比例共培养可获得更多的软骨细胞。BACKGROUND： Human umbilical cord blood-derived mesenchymal stem cells can be induced through the co-culture to differentiate into other cells; however, it is difficult to control the proportions of two kinds of cells for maximizing the efficiency of obtaining target celts. OBJECTIVE： To investigate chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells by co-culture with rabbit chondrocytes at different ratios.METHODS： Human umbilical cord blood-derived mesenchymal stem cells were resuscitated and cultured, and identified by flow cytometry. Then human umbilical cord blood-derived mesenchymal stem cells were co-cultured with rabbit chondrocytes at 2：1 or 3：1 ratio, and induced with insulin-like growth factor-l. After 14 days in co-culture, the cell RNA and protein were extracted, aggrecan mRNA and collagen type II mRNA expression levels were detected by real-time quantitative-polymerase chain reaction, and aggrecan and collagen type I! protein expression levels were detected by western blot assay. RESULTS AND CONCLUSION： After human umbilical cord blood-derived mesenchymal stem cells were co-cultured with rabbit chondrocytes for 14 days, the aggrecan mRNA and collagen type 2 mRNA expression in the co-cultured cells was significantly higher than that in the cells only induced with insulin-like growth factor-l. Under the same ratio of human umbilical cord blood-derived mesenchymal stem ceils and rabbit chondrocytes, insulin-like growth factor-1 increased the aggrecan and collagen type I[ protein and mRNA expressions. A 3：1 co-culture ratio allowed the increment in the collagen type I! mRNA and protein expression, as well as aggrecan protein expression; while a 2：1 co-culture ratio upregulated aggrecan mRNA expression. Experimental findings indicate that, human chondrocytes can be successfully induced by co-culture of human umbilical cord blood-derived mesenchymal stem cells and rabbit chondrocytes, and 3：1 co-culture ratio can obtain more chondrocytes.

关 键 词：干细胞 脐带脐血来源干细胞 软骨细胞 共培养 胰岛素样生长因子 Ⅱ型胶原 聚集蛋白多糖 软骨缺损 软骨组织工程 种子细胞 其他基金 

分 类 号：R394.2[医药卫生—医学遗传学]