RNAi沉默STAT3基因对人宫颈癌SiHa细胞定植和侵袭的影响

Inhibition of STAT3 Expression by RNA Interference on Location and Metastasis in Human Cervical Carcinoma SiHa Cells

目的:探讨信号转导因子与转录激活因子3(STAT3)短发夹RNA(shRNA)真核表达载体对宫颈癌SiHa细胞定植和侵袭能力的影响。方法:针对人STAT3的基因设计并合成编码小干扰RNA(siRNA)的寡核苷酸,克隆入pSilencer2.1-U6-neo质粒中,构建STAT3基因shRNA真核表达质粒,脂质体法将重组质粒转染人宫颈癌SiHa细胞。逆转录-聚合酶链反应(RT-PCR)和蛋白质印迹(Western blot)法分别检测STAT3基因的mRNA及蛋白表达水平;软琼脂克隆形成实验和体外侵袭实验检测SiHa细胞的定植和侵袭能力。结果:构建的STAT3基因shRNA真核表达载体成功地转染人宫颈癌SiHa细胞,细胞STAT3蛋白及mRNA表达均下降(P<0.05);SiHa细胞在软琼脂中形成的克隆数和穿透Matrigel膜的细胞数均明显减少(P<0.05)。结论:STAT3基因shRNA真核表达载体通过下调STAT3基因表达,抑制宫颈癌细胞的定植和侵袭能力。

Objective：To explore the effects of eukaryotic vector expressing short hairpin RNA （shRNA） targeting signal transducers and activators of transcription 3 （STAT3） on the location and metastasis of human cervical carcinoma SiHa cells.Methods：The shRNA templates was designed based on STAT 3 gene sequence and was cloned into pSilencer2.1-U6-neo vector.The resultant plasmid was transfected into SiHa cells with Lipofectamine 2000.The Western blot and RT-PCR were used to detect STAT3 protein and mRNA respectively.The colony formation assay and Transwell cabin assay were performed to measure the location and metastasis of SiHa cells.Results：The plasmid pSTAT3-siRNA was successfully constructed and transfected into SiHa cells.The expressions of STAT3 in SiHa cells decreased.At the same time,the number of colony formation in soft agar and cells penetrating matrigel also decreased.Conclusions：The STAT 3 specific shRNA expression vector could inhibit the cellular location and metastasis of cervical cancer cells through suppressing the STAT3 expression.