Lv Yang, Li Xiu-Juan, Wang Hai-Ping, Liu Bo, Chen Wei, Zhang Lei
Department of Histology and Embryology, Hebei North University, Zhangjiakou, Hebei, China.
Department of Histology and Embryology, Hebei Medical University, Shijiazhuang, Hebei, China.
Iran J Basic Med Sci. 2020 Aug;23(8):1012-1019. doi: 10.22038/ijbms.2020.42396.10019.
To investigate and test the hypotheses that TGF-β1 enhanced myocardial differentiation through Wnt/β-catenin pathway with rat bone marrow mesenchymal stem cells (BMSCs).
Lentiviral vectors carrying the TGF-β1 gene were transduced into rat BMSCs firstly. Then several kinds of experimental methods were used to elucidate the related mechanisms by which TGF-β1 adjusts myocardial differentiation in rat BMSCs.
Immunocytochemistry revealed that cTnI and Cx43 expressed positively in the cells that were transduced with TGF-β1. The results of Western blot (WB) test showed that the levels of intranuclear β-catenin and total β-catenin were all significantly decreased. However, the cytoplasmic β-catenin level was largely unchanged. Moreover, the levels of GSK-3β were largely unchanged in BMSCs, whereas phosphorylated GSK-3β was significantly decreased in BMSCs. When given the activator of Wnt/β-catenin pathway (lithium chloride, LiCl) to BMSCs transducted with TGF-β1, β-catenin was increased, while phosphorylated β-catenin was decreased. In addition, cyclinD1, MMP-7, and c-Myc protein in BMSCs transducted with Lenti-TGF-β1-GFP were significantly lower.
These results indicate that TGF-β1 promotes BMSCs cardiomyogenic differentiation by promoting the phosphorylation of β-catenin and inhibiting cyclinD1, MMP-7, and c-Myc expression in Wnt/β-catenin signaling pathway.
研究并验证转化生长因子-β1（TGF-β1）通过Wnt/β-连环蛋白信号通路增强大鼠骨髓间充质干细胞（BMSCs）心肌分化的假说。
首先将携带TGF-β1基因的慢病毒载体转导至大鼠BMSCs中。然后采用多种实验方法阐明TGF-β1调节大鼠BMSCs心肌分化的相关机制。
免疫细胞化学显示，TGF-β1转导的细胞中肌钙蛋白I（cTnI）和连接蛋白43（Cx43）呈阳性表达。蛋白质免疫印迹（WB）检测结果显示，细胞核内β-连环蛋白和总β-连环蛋白水平均显著降低。然而，细胞质β-连环蛋白水平基本未变。此外，BMSCs中糖原合成酶激酶-3β（GSK-3β）水平基本未变，而磷酸化GSK-3β在BMSCs中显著降低。对转导了TGF-β1的BMSCs给予Wnt/β-连环蛋白信号通路激活剂（氯化锂，LiCl）后，β-连环蛋白增加，而磷酸化β-连环蛋白减少。此外，慢病毒-TGF-β1-绿色荧光蛋白（Lenti-TGF-β1-GFP）转导的BMSCs中细胞周期蛋白D1（cyclinD1）、基质金属蛋白酶-7（MMP-7）和原癌基因c-Myc蛋白显著降低。
这些结果表明，TGF-β1通过促进β-连环蛋白磷酸化并抑制Wnt/β-连环蛋白信号通路中cyclinD1、MMP-7和c-Myc的表达来促进BMSCs向心肌细胞分化。
