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.