RNA干扰技术干涉后核糖体蛋白S15a在胃癌中的表达及意义

Knock—down of RPS15a by siRNA inhibits proliferation of gastric cancer BGC823 cells

目的探讨RNA干扰(RNAi)技术干涉后核糖体蛋白S15a(RPS15a)在胃癌中的表达及意义。方法选取12例胃癌患者手术切除的胃癌组织,利用RNAi干涉RPS15a基因,将小片段克隆到质粒中,分别提取质粒后转染胃癌细胞BGC823。将未转染、随机小片段转染非编码序列、siRPS15a-1转染、siRPS15a-2转染的BGC823细胞分为BGC823-WT组、siNC组、siRPS15a-1组、siRPS15a-2组,RT-PCR法及Western blot分别检测各组RPS15a RNA及蛋白的表达情况,流式细胞仪检测细胞增殖的变化。结果 RNAi后细胞中出现了明显的绿色荧光,提示此转染方式是可行的。转染检测结果表明,本实验提取的RNA未受到质粒DNA的污染,可以进行表达的比较。siRPS15a-1组、siRPS15a-2组RNA及蛋白表达相对灰度比值与BGC-WT组、siNC组比较,差异均有统计学意义(均P<0.05),BGC-WT组与siNC组比较差异无统计学意义(P>0.05)。RNAi后细胞增殖速度显著下降,多数细胞集中在G_0/G_1期。结论 RNAi技术不仅可以下调RPS15a在胃癌中表达,而且还可以抑制胃癌细胞的增殖活性。

Objective To investigate the effects of ribosomal protein S15a （RPS15a） in proliferation of gastric cancer cellBGC823 by RNA interference technique. Methods BGC823 cells were transfected with random non- coding sequences （siNC group）, siRPS15a- 1 （siRPS15a- 1 group） or siRPS15a- 2 （siRPS15a- 2 group）, respectively. RPS15a RNA and protein expression was detected by RT- PCR and Western Blot test, respectively;and the cellproliferation was determined by flow cytometry. Re-sults The expression levels of RPS15 in SiRPS15a- 1 and siRPS15a- 2 groups were significantly lower than those in non- trans-fected BGC823 cells （WT group） and siNC group （al P〈0.05）; no significant differences was found between WT and siNC groups （P〉0.05）. cellproliferation rate were decreased significantly in SiRPS15a- 1 and siRPS15a- 2 groups, and most cells were in G0/G1 phase. Conclusion RPS15a RNA interference technique inhibits cells growth, which is associated with the knock- down of RPS15a expression in BGC823 cells.