摘要
目的研究RNA干扰(RNAi)对SSP411进行基因沉默后胆管癌细胞体外增殖的变化。方法构建针对SSP411的RNAi质粒,将离体培养的胆管癌HuCCA-1细胞分为阴性对照(C)组、shRNA-SSP411-1转染(A1)组和shRNA-SSP411-2转染(A2)组。采用RT-PCR和Western blot法分别检测HuCCA-1细胞中SSP411mRNA和蛋白表达;通过MTT实验、平板克隆形成和软琼脂克隆球形成能力检测,观察目的基因被下调后细胞增殖能力的变化。结果 A1、A2组细胞的SSP411mRNA和蛋白表达较C组明显降低。与C组相比,A1、A2组胆管癌细胞生长被显著抑制(P<0.01);A1、A2组平板克隆形成数目低于C组[(66±18)个、(70±17)个vs.(115±16)个](P<0.01或P<0.05);与C组相比,A1、A2组软琼脂克隆球数目显著下降[(96±29)个vs.(53±9)个、(43±13)个](P<0.05)。结论 RNAi介导的SSP411基因沉默可抑制人胆管癌HuCCA-1细胞的增殖能力;SSP411可作为胆管癌治疗的新靶点。
Objective To investigate the effects of RNA interference（RNAi） targeting SSP411 gene on the proliferation of human cholangiocarcinoma cells in vitro. Methods Two RNAi vectors against SSP411 gene were constructed. HuCCA-1 cells were divided into three groups of C（negative control）, A1 （stably transfected with shRNA-SSP411-1 vector） and A2 （stably transfected with shRNA-SSP411-2 vector）. The expressions of SSP411 mRNA and protein were detected by RT-PCR and Western blot, respectively. The proliferation of HuCCA-1 cells after RNAi was assessed by MTT, colony forming ability and soft agar gel colony forming ability assays. Results The expressions of SSP411 mRNA and protein in groups of A1 and A2 were lower than those in group C. The cell growth was significantly inhibited in groups of A1 and A2, compared with group C（P〈0. 01）. The numbers of colony formation in groups of A1 and A2 were lower than those in group C[（66±18） pieces and （70±17） pieces vs. （115±16） pieces] （P〈0. 01 or P〈0. 05）. Similarly, compared with group C, the numbers of clone balls in groups of A1 and A2 were obviously decreased[（96±29） pieces vs. （53＋9） pieces and （43±13） pieces~]P〈0. 05）. Conclusion RNAi-mediated SSP411 gene silencing can inhibit the proliferation of HuCCA-1 cells. The SSP411 may be taken as a potential target for cholangiocarcinoma therapy.
出处
《江苏医药》
CAS
北大核心
2013年第8期888-891,共4页
Jiangsu Medical Journal
基金
南京医科大学校基金重点项目(2012NJMU087)
