转化生长因子-β1对骨髓干细胞分化过程中Wnt信号通路相关基因表达的影响  

Effects of transforming growth factor--β1 on the expression of Wnt signaling pathway related genes in the differentiation of bone marrow stromal cells

作　　者：曾晖[1] 肖德明[1] 陶可[1] 熊奡[1] 翁鉴[1] 辛风[1] 

机构地区：[1]北京大学深圳医院骨关节科,深圳518036

出　　处：《中华实验外科杂志》2013年第7期1347-1350,共4页Chinese Journal of Experimental Surgery

基　　金：深圳市科技计划资助项目（201002067）

摘　　要：目的观察转化生长因子（TGF）-β1对骨髓干细胞向软骨细胞分化过程中Wnt信号通路相关基因表达的影响。方法TGF-β1软骨诱导液培养骨髓干细胞（BMSCs），倒置相差显微镜进行形态学观察，甲苯胺蓝染色、鉴定。酶联免疫吸附试验（ELISA）法检测不同浓度TGF-β1对糖胺聚糖（GAG）表达量的影响，实时荧光定量聚合酶链反应（FQ—PCR）检测目的基因Wntl、Wnt3a、13-连环蛋白（13-eatenin）、Sox9和胶原蛋白II（CollagenII）mRNA表达。结果TGF-β1诱导培养第21天，甲苯胺蓝染色显示，95％以上细胞为软骨细胞；2μg/LTGF-β1诱导液组细胞培养液中GAG表达量为（69．16±1．18）mg／L，明显高于其余各组（P〈0．05）；FQ—PCR结果表明，与对照组比较，2μg/LTGF-β1诱导液组的Wntl、Wnt3amRNA的表达量分别为0．0266±0．0047、0．0154±0．0033，其表达受到了抑制（P〈0．05），13-eateninmRNA的表达量为0．0187±0．0021，明显低于对照组（P〈0．01）；相反，Sox9、CollagenIImRNA表达量分别为0．0682±0．0024、0．1134±0．0048，表达有所升高（P〈0．05）。结论TGF-β1促进BMSCs向软骨细胞分化，Wnt／βcatenin信号通路相关基因表达受到抑制，而软骨细胞特异基因表达增加。Objective To observe the transforming growth factor-β1 （TGF-β1） on the expression of Writ signaling pathway related genes during the chondrogenesis of bone marrow stromal cells （BMSCs） in vitro. Methods BMSCs cultured in TGF-β1 medium were observed by inverted phase contrast microscope and identified by toluidine blue staining. The expression levels of glycosaminoglycan （GAG） and target genes including Wntl, Wnt3a, β-catenin, Sox9 and collagen II mRNA under different concentrations of TGF-β1 were detected by using enzyme linked immunosorbent assay （ELISA） and real-time fluorescence quantitative polymerase chain reaction （FQ-PCR） respectively. Results At day 21 after TGF-β1 induc- tion, toluidine blue staining showed that more than 95% cells differentiated into chondrocytes ; the amount of GAG in the cell cultured medium in 2 ~g/L TGF-[31-induced group was （69. 16 ± 1.18） rag/L, signifi- cantly higher than other groups （ P 〈 0. 05 ）. FQ-PCR results showed that Wntl, Wnt3a and 13-catenin mRNA expression in 2 μg/L TGF-β1-induced group was inhibited [ 0. 0266 ± 0. 0047, 0. 0154 ±-0. 0033, 0. 0187±0. 0021, （P〈0. 05）], but in contrast Sox9 and collagen II mRNA expression levels were in- creased [ 0. 0682± 0. 0024, 0. 1134± 0. 0048 （ P 〈 0. 05 ） ] as compared with the control group. Conclu- sion TGF-β1 could promote BMSCs chondrogenie differentiation, and the expression of Wnt/β-eatenin signaling pathway related genes Wntl, Wnt3a and β-catenin mRNA was inhibited and the expression of chondrocyte specific genes Sox9 and collagen II mRNA was increased.

关 键 词：转化生长因子-Β1 骨髓干细胞 WNT信号通路 糖胺聚糖 

分 类 号：R3[医药卫生—基础医学]