RNAi沉默STAT3基因对人卵巢癌细胞生长的抑制作用

摘要

Objective: To study the effects of STAT3 knockdown by siRNA on the growth of an ovarian cancer cell line (SKOV3 cells).Methods: The siRNA knockdown plasmid against STAT3 (pSilencerl. 0-U6-siRNA-STAT3) was successfully constructed and transfected into SKOV3 cells. The STAT3 expression in SKOV3 cells transfected with pSilencerl. 0-U6-siRNA-STAT3 was detected by Western blot and Northern blot. MTT assay and flow cytometer (FCM) were used to observe the growth ratio and apoptosis in SKOV3 cells.Results.. The Western blot and RT-PCR analysis demonstrated that pSilencer 1.0-U6-siRNA-STAT3 could significantly inhibit the expression of STAT3 in SKOV3 cells. MTT and FCM results showed that it could suppress the growth of SKOV3 cells and induce apoptosis of SKOV3 cells in vitro.Conclusions.. pSilencer 1.0-U6-siRNA-STAT3 could significantly inhibit STAT3 expression and suppress the growth of SKOV3 cells and induce the apoptosis of SKOV3 ceils.%目的 探讨RNA干扰沉默STAT3基因表达对人卵巢癌细胞SKOV3的生长抑制作用.方法针对STAT3 mRNA序列设计合成3对编码小干扰RNA(siRNA)的DNA模板,构建重组质粒(pSilencer 1.0-U6-siRNA-STAT3),转染SKOV3细胞进行体外研究;采用Western blot、实时聚合酶链反应(RT-PCR)和免疫细胞化学技术检测重组质粒对STAT3基因表达的作用;MTT法检测重组质粒对SKOV3细胞增殖的影响;流式细胞术(FCM)及吖啶橙染色检测重组质粒诱导的细胞凋亡.结果 (1)免疫组织化学技术证实卵巢癌细胞株SKOV3及癌组织中STAT3呈高表达;(2)成功构建pSilencer 1.0-U6-siRNA-STAT3重组质粒,并成功转染SKOV3细胞;(3)MTT法证实重组质粒抑制SKOV3细胞的增殖,流式细胞技术证明重组质粒诱导细胞凋亡;(4)RT-PCR与Western blot分析表明,重组质粒在mRNA及蛋白质水平特异的抑制STAT3的表达;(5)STAT3基因表达的下调抑制抗凋亡蛋白Bcl-2的表达.结论 pSilencer 1.0-U6-siRNA-STAT3重组质粒可抑制STAT3在人卵巢癌细胞SKOV3中的表达,并抑制肿瘤细胞的生长,促进其凋亡.