泛素特异性蛋白酶15对结肠癌细胞上皮间质转化的影响

Effects of ubiquitin specific peptidase 15 on epithelial-mesenchymal transition in colon cancer cells

目的:研究泛素特异性蛋白酶15(ubiquitin specific peptidase 15,USP15)对结肠癌HCT116细胞上皮间质转化的影响。方法:TGF-β_1可诱导结肠癌细胞发生上皮间质转化。采用TGF-β_1处理HCT116细胞,同时采用人USP15特异性小干扰RNA(TGF-β_1+siUSP15)转染HCT116细胞,阴性对照序列(TGF-β_1+siScramble)作为对照。采用siUSP15转染结肠癌奥沙利铂耐药细胞株(HCT116/L-OHP),然后采用奥沙利铂处理48 h。阴性对照序列(siScramble)作为对照,实时荧光定量聚合酶链反应检测USP15、E钙黏蛋白和波形蛋白mRNA表达。Western印迹检测USP15、E-钙黏蛋白、波形蛋白的蛋白表达。细胞划痕实验检测HCT116的迁移能力。CCK8法检测细胞抑制率。结果:siUSP15能有效抑制USP15在HCT116中的表达。与TGF-β_1+siScramble组相比,TGF-β_1+siUSP15组细胞中E钙黏蛋白表达上调(P<0.05),而波形蛋白表达下降(P<0.05)。细胞划痕实验显示,与TGF-β_1+siScramble组相比,TGF-β_1+siUSP15细胞的迁移能力显著下降(P<0.05)。CCK8检测结果显示,USP15的表达下调能增加HCT116/L-OHP对奥沙利铂的敏感性(P<0.05)。结论:USP15在结肠癌细胞上皮间质转化过程中发挥重要作用。

Objective To investigate the effects of ubiquitin specific peptidase 15（USP15） on epithelial-mesenchymal transition（EMT） of HCT116 colon cancer cells. Methods TGF-β1 which could induce EMT of colon cancer cell was used to treat HCT116 cells. USP15 cells was transfected with small interfering RNA against human USP15（TGF-β1＋siUSP15）and negative control cells with TGF-β1＋siScramble. Oxaliplatin-resistant colon cancer cells（HCT116/L-OHP） were transfected with small interfering RNA against human USP15（siUSP15） and negative control cells with siScramble sequence.Then the cells were treated with oxaliplatin for 48 hours. The m RNA expressions and protein levels of USP15, E-cadherin and vimentin were detected by quantitative real-time polymerase chain reaction and Western blot respectively. The migration ability of HCT116 cells was measured by scratch wound assay. Inhibition of the cells was analyzed by CCK8. Results The expression of USP15 in HCT116 cells was inhibited by siUSP15 remarkably. Compared with TGF-β1＋siScramble group, the expression of E-cadherin in TGF-β1＋siUSP15 group increased（P〈0.05） and the expression of vimentin decreased（P〈0.05）. The migration ability of the cells in TGF-β1＋siUSP15 group was shown by scratch wound assay to be attenuated significantly compared with TGF-β1＋siScramble group（P〈0.05）. CCK8 assay showed the downregulation of USP15 expression to promotes the chemosensitivity of HCT116/L-OHP to oxaliplatin. Conclusions It was shown that USP15 could play an important role in the EMT of colon cancer cells.