靶向于雄激素受体的siRNA筛选及效能评价  

Screening and evaluation of a newly designed siRNA targeting androgen receptor

作　　者：张淼[1] 郭燕丽[1] 王洛夫[2] 李彦[3] 李锐[1] 

机构地区：[1]第三军医大学西南医院超声科,重庆400038 [2]第三军医大学大坪医院野战外科研究所泌尿外科,重庆400042 [3]第三军医大学西南医院中心实验室,重庆400038

出　　处：《第三军医大学学报》2011年第23期2459-2462,共4页Journal of Third Military Medical University

基　　金：国家自然科学基金面上项目(30970830)~~

摘　　要：目的设计并合成新的靶向于雄激素受体(AR)的siRNA序列,转染前列腺癌细胞筛选最佳干扰序列。方法设计并合成靶向于雄激素受体的siRNA,以不同浓度转染雄激素非依赖性前列腺癌细胞C4-2,确定最佳转染浓度。以最佳转染浓度转染细胞,分为siRNAⅠ组、siRNAⅡ组、siRNAⅢ组、阴性对照组、空白对照组和无关对照组。RT-PCR检测AR mRNA表达水平,Western blot检测AR蛋白表达水平。CCK-8分析绘制细胞生长曲线,观察C4-2细胞生长活性并筛选出抑制效果最佳的siRNA靶序列。结果以不同浓度的AR siRNA转染C4-2细胞后,细胞生长活性降低(P<0.05),并筛选出100 nmol/L的最佳转染浓度;RT-PCR和Western blot结果显示转染后AR的mRNA和蛋白表达水平显著下降(P<0.05);CCK-8法检测结果显示各组细胞增殖活性显著降低,其中siRNAⅠ的抑制效果最为明显(P<0.05)。结论新设计合成的AR siRNA能通过沉默AR表达高效抑制C4-2细胞的增殖。Objective To design and synthesize siRNA sequences targeting androgen receptor（AR）,and to screen effective sequences by transfecting C4-2 cells.Methods Three pairs of siRNA sequences targeting AR were designed and synthesized according to the cDNA of AR,and were transfected into androgen-independent prostate carcinoma C4-2 cells to determine an optimal concentration.Experimental groups comprised of siRNAⅠ,siRNAⅡ,and siRNAⅢ,negative control,blank control and unrelated control.RT-PCR and Western blotting were applied to detect mRNA and protein expression of AR,respectively.Cell growth activity and the most effective siRNA sequence were evaluated based on cell growth curves by CCK-8 analysis.Results Cell growth activities of C4-2 cells transfected with different concentrations of AR siRNA（20 nmol/L,50 nmol/L,100 nmol/L and 200 nmol/L,respectively） were suppressed,and 100 nmol/L was the optimum concentration of AR siRNA（P〈0.05）.Compared with those of the control groups,mRNA and protein levels of AR decreased significantly,and C4-2 cell growth activity was suppressed significantly in the groups of C4-2 cells transfected with AR siRNA（P〈0.05）.The inhibitory effect of AR siRNAⅠ was the most significant than that of the others.Conclusion The newly designed AR siRNA can inhibit C4-2 cell proliferation by silencing AR expression.

关 键 词：RNA干扰 雄激素受体 雄激素非依赖性前列腺癌 

分 类 号：R394.33[医药卫生—医学遗传学] R394.3[医药卫生—基础医学]