HSP27在卵巢癌顺铂耐药细胞系中的作用

Roles of HSP27 in Cisplatin Resistance in Ovarian Cancer Cells

目的探讨HSP27在卵巢癌耐药细胞系及敏感细胞系中的表达及其在卵巢癌顺铂耐药中的作用。方法通过RT-PCR及Western blot检测卵巢癌细胞株OV2008和CI3K中HSP27 mRNA和蛋白的表达水平;将合成的针对HSP27基因的特异性siRNA及正义真核表达质粒pEGFP-C1-HSP27(命为p-HSP27),分别转染CI3K和OV2008细胞。通过RT-PCR及Western blot检测转染前后CI3K及OV2008细胞中HSP27 mRNA和蛋白表达的变化;MTT及流式细胞仪分别测定转染前后CI3K及OV2008细胞对顺铂敏感度的变化。结果 (1)HSP27在CI3K细胞中的mRNA和蛋白表达水平显著高于OV2008细胞,差异有统计学意义(P<0.05);(2)转染siRNA48 h,CI3K细胞对顺铂的半数抑制浓度(IC50)明显低于转染空载体和未转染的CI3K细胞(P<0.05);转染正义质粒p-HSP27 48 h,OV2008细胞对顺铂的IC50明显高于转染空载体和未转染的OV2008细胞(P<0.05);(3)转染siRNA及正义质粒48 h,再用顺铂作用CI3K细胞24 h。转染siRNA的CI3K细胞的凋亡率比转染空载体和未转染的CI3K细胞的凋亡率明显增加,前者明显高于后两者(P<0.05);转染正义质粒的OV2008细胞的凋亡率比转染空载体和未转染的细胞的凋亡率明显下降,前者明显低于后两者(P<0.05)。结论 CI3K及OV2008细胞内HSP27基因表达的强弱可影响其对顺铂的敏感度,提示HSP27可能在卵巢癌顺铂耐药中起重要作用。

Objective To investigate the expression and role of HSP27 in Cisplatin-sensitive ovarian cancer cell OV2008 and Cisplatin-resistant ovarian cancer cell CI3K.Methods To examine expression of HSP27 gene in OV2008 cells and CI3K cells;siRNA and eukaryotic expression plasmid pEGFP-C1-HSP27were synthesized and transfected into human ovarian cancer cell by lipo2000,mRNA and protein expressions of HSP27 at different times after transfection were measured by real-time RT-PCR and Western blot,and the drug sensitivity to Cisplatin was detected by MTT and flow cytometry.Results（1） The expression of protein and mRNA on HSP27 in CI3K cells were significantly higher than those in OV2008 cells,the differences are significant（P〈0.05）.（2） Transfection of siRNA 48 h later,the 50% inhibition concentration（IC50） to Cisplatin of CI3K transfected with siRNA was obviously lower than that of empty-vector transfected cells and untransfected cells（P〈0.05）.Transfection with HSP27 full length plasmid p-HSP27 48 h later,the IC50 of OV2008 to Cisplatin was obviously higher than that of empty-vector transfected cells and untransfected cells（P〈0.05）.（3） Transfection of plasmid siRNA and p-HSP27 48 h,and then treated by Cisplatin 24 h later,the apoptosis rate of CI3K was increased[（41.42±0.68）%],compared with empty vector（28.87±2.65）% and non-transfected CI3K cells（26.96±3.79）%（P〈0.05）;The apoptosis rate of OV2008 transfected with p-HSP27（27.68±1.64）% was significantly decreased compared with empty vector transfected（47.52±3.12）% and non-transfected（49.32±2.31%） OV2008 cells（P〈0.05）.Conclusion HSP27 gene expression in CI3K and OV2008 cells could affect sensitivity to Cisplatin,suggesting that HSP27 gene might play an important role on Cisplatin-resistant in ovarian cancer.