siRNA沉默KLF4表达促进HeLa细胞上皮间质化

Silencing the expression of KLF4 by siRNA improve the epithelial mesenchymal transition of Hela cells

目的:探讨Klf4表达水平对Hela细胞上皮间质转化的影响。方法:设计合成针对Klf4的siRNA和阴性对照siRNA,并转染至Hela细胞。用含10ng/ml TGF-β1和10%胎牛血清的DMEM培养液诱导Hela细胞发生上皮间质转化,对照组使用不含TGF-β1的培养液。倒置显微镜下观察记录细胞的形态学改变。Western blot法和细胞免疫荧光法检测Klf4、上皮标志分子E-cadherin、ZO-1及间质标志分子N-cadherin的表达变化。结果:在TGFβ1的诱导作用下,Hela细胞呈现出梭形、类成纤维细胞的形态,发生了上皮间质转化。Western blot和细胞免疫荧光检测结果显示,Hela细胞在TGFβ1诱导作用下,Klf4、上皮标志分子E-cadherin、ZO-1表达减少,间质标志分子N-cadherin表达增多。转染了si Klf4的Hela细胞经TGFβ1诱导后,与转染了阴性对照siRNA的细胞相比,上皮标志分子E-cadherin、ZO-1表达略有减少,而间质标志分子N-cadherin的表达则明显增多。结论:抑制Klf4表达可以促进Hela细胞的上皮间质转化。

Objective： To investigate the effect of Klf4 on the EMT of Hela. Method： siRNA targeted to Klf4 and control siRNA was designed and transfected into Hela cells. DMEM with 10ng/ml TGF-β1 and 10% fetal calf serum（FBS）were used to induce the epithelial mesenchymal transition （EMT）of Hela. Culture medium without TGF-β1 was taken as control. The morphology of Hela cells were observed and recorded by phrase contrast microscope. The expressions of Klf4,E-cadherin,ZO-I and N-cadherin were detected by Western blot and Immunofluorescence. Results： Hela cells acquired mesenchymal features, showed spindle-shape, fibroblast-like morphology and EMT aroused under the induction of TGFβ1. The expressions of Klf4,epithelial marker E-cadherin, ZO-1 were down-regulated while mesenchymal marker N-cadherin was up-regulated in Hela cells induced by TGFβ1 both in Western blot and Immuno-fluorescence. Compared with cells transfected by control siRNA, the expression of epithelial marker E-cadherin, ZO-1 were down-regulated slightly while the mesenchymal marker N-cadherin was up-regulated significantly after Hela cells were transfected by siKLF4 and induced by TGFβ1 both in Western blot and Immunofluorescence. Conclusion： The EMT of Hela is improved after inhibiting the expression of Klf4 by siRNA.