应用RNA干扰技术阻断LNCap细胞SRC-1基因的表达及意义

Inhibition of SRC-1expression in prostate cancer cells by RNAi and its significance

目的：利用RNA干扰（RNA i）技术,阻断前列腺癌LNCap细胞中类固醇受体共激活分子（SRC-1）的表达,并研究SRC-1基因沉默后对LNCap细胞的影响。方法：将设计好的siRNA,用脂质体法转染LNCap细胞,通过Q-PCR、蛋白免疫印迹检测SRC-1的表达变化,并用CCK-8法检测细胞生长情况的变化。结果：转染SRC-1siRNA 24 h后的前列腺癌LNCap细胞中SRC-1 mRNA的量与空白对照组相比较下降了约35%,转染48 h后下降了约77%,差异具有统计学意义（P〈0.05）。转染24~48 h后LNCap细胞中SRC-1蛋白的表达量（24 h为0.359±0.034;48 h为0.257±0.065）与阴性对照组（24 h为0.782±0.078;48 h为0.766±0.043）、转染试剂组（24 h为0.840±0.013;48 h为0.786±0.051）和空白对照组（24 h为0.816±0.065;48 h为0.805±0.107）相比较明显减少（P〈0.05）。转染24、48、72、96 h后LNCap细胞的生长抑制率分别为25%、52%、55%和60%。结论：SRC-1与LNCap细胞的生长相关。SRC-1在激素非依赖性前列腺癌中的高表达可能参与了前列腺癌雄激素非依赖性的转变过程。抑制SRC-1的表达可能成为临床治疗激素依赖性前列腺癌的方法之一。

Objective： To investigate the inhibition of the expression of steroid receptor coactivator-1（SRC-1） in the LNCap cell line through RNA interference（RNAi） and the effect of the silenced SRC-1 gene on LNCap cells.Methods： The experiment included four groups： siRNA transfection,siRNA negative control,bland vehicle（with Lipofectamine2000 but no siRNA）,and blank control（with neither Lipofectamine2000 nor siRNA）.LNCap cells were transfected with designed siRNA using the liposomes method,the expressions of SRC-1 determined by Q-PCR and Western blot,and the proliferation of the LNCap cells detected by the CCK-8 method.Results： The expression of SRC-1 mRNA in the transfected LNCap cells was decreased by 35% at 24 hours and 77% at 48 hours,with statistically significant differences from the blank control group（P0.05）.The SRC-1 protein expression of the transfected group was 0.359±0.034 at 24 hours and 0.257±0.065 at 48 hours,markedly decreased as compared with that of the negative control（0.782±0.078 and 0.766±0.043）,bland vehicle（0.840±0.013 and 0.786±0.051）,and blank control group（0.816±0.065 and 0.805±0.107）（P0.05）.The LNCap cell growth inhibition rates were 25%,52%,55% and 60% at 24,48,72 and 96 hours,respectively.Conclusion： The expression of SRC-1 is correlated with the growth of LNCap cells;its high expression in androgen-independent prostate cancer cells may be involved in the progression to androgen-independence.Inhibiting the expression of SRC-1 may be an option for the treatment of androgen-dependent prostate cancer.