TGF-β1 induces activation of HSC-T6 cells and epithelial-mesenchymal transition in rats
- 医学院－已发表论文 
目的探讨转化生长因子-β1（TGF-β1）在大鼠肝星状细胞系（HSC-T6）活化及上皮间质转换（EMT）中的作用。方法体外培养HSC-T6，用MTT法筛选TGF-β1对HSC-T6作用的最佳浓度；用10μg／L TGF-β1处理HSC-T624h，相差倒置显微镜下观察细胞形态改变，免疫荧光染色法检测细胞骨架结构F-actin蛋白的表达，RT-qPCR法检测肌动蛋白α-SMA及代表上皮间质转换的神经黏附素（N-cadherin）、波形蛋白（vimentin）和上皮黏附素（E-cadherin）基因表达；用不同浓度（0、5和10μg/L）的TGF-β1处理HSC-T624h，Western blot检测α-SMA、N-cadherin、vimentin和E-cadherin蛋白表达。结果10μg/L TGF-β1干预HSC-T624h有最好的细胞存活率；TGF-β1刺激HSC-T6后，细胞拉伸，伪足增多呈星形，细胞间连接疏松，呈显著活化状态；F-actin聚集形成应力纤维丝，沿细胞长轴分布；实验组α-SMA mRNA及vimentin mRNA的表达量明显高于对照组（P〈0．05），而E-cadherin mRNA的表达量明显降低（P〈0．05）；在不同浓度的TGF-β1呈剂量依赖性致α-SMA及N-cadherin和vimentin的蛋白表达量增多，而E-cadherin的蛋白表达量减少。结论TGF-β1可诱导HSC-T6活化及上皮间质转换。Objective To observe the effect of TGF-β1 on activation and epithelial mesenchymal transition （EMT） in rat hepatic stellate cell-T6. Methods Adopt the MTT method to screen the optimum concentration of TGF-β1 in vitro HSC-T6 cultured. After the HSC-T6 stimulation by TGF-β1 of 10 μg/L for 24 hours, the morphology of the cells was observed under inverted phase contrast microscope, the expression of F-actin which on behalf of cotoskeletal structure was detected by immunofluorescence staining; the expression of α-SMA and N-cadherin, vimentin, E-eadherin was measured by RT-qPCR ; The changes of α-SMA, N-cadherin, vimentin and E-eadherin were assessed by Western blot after different concentrations （0,5 and 10 μg/L） of TGF-β1 interventing HSC-T6 for 24 h. Results The optimal cell survival rate was recorded when 10 μg/L TGF-β1 dealt with cells for 24 h. After HSC-T6 were treated with TGF-β1 ,cells stretched, pseudopodia increased and turn into stellate, ceils connections were looser, so that represented a significantly activated state. F-actin filaments gathered to form stress and distributed along the long axis of the cells ; The expression of α-SMA mRNA and vimentin mRNA in experimental group was significantly higher while E-cadherin mRNA was obviously lower than the control group （P〈 0. 05）. TGF-β1 made the protein expression of α-SMA and N-cadherin, vimentin in dose-dependent increased while E-cadherin was decreased.Conclusions TGF-β1 may induce activation and epithelial-mesenchymal transition of HSC-T6.