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目的:探讨siRNA对胃癌BGC823和MGC803细胞中Cullin1(Cul1)基因表达、细胞增殖的影响及其分子作用机制。方法:体外化学合成Cul1siRNA序列,在siLentFect Lipid Reagent介导下转染2种胃癌细胞。蛋白质印迹法检测Cul1、Cyclins和CDK抑制剂蛋白表达,CCK8法检测细胞增殖活性,流式细胞术检测细胞周期。结果:转染Cul1siR-NA后胃癌BGC823和MGC803细胞中Cul1蛋白表达分别下调39%和84%,Cyclin D1下调59%和62%,Cyclin E下调47%和54%,而p27蛋白表达上调52%和23%,但p21蛋白表达无明显变化。Cul1干涉后24h2种胃癌细胞的增殖能力下降(P<0.01),48和72h下降更明显,P<0.01。Cul1干涉后3和6hG1期细胞比例较对照组下降缓慢(P<0.01),而Sub-G1期Cul1干涉组与对照组差异无统计学意义。结论:Cul1siRNA可有效的干涉胃癌BGC823和MGC803细胞中Cul1基因的表达,Cul1干涉后可能通过影响Cyclins和CDK抑制剂蛋白表达从而使细胞周期停滞在G1期,最终能明显抑制胃癌细胞的增殖。
Objective: To investigate the effect of siRNA on Cullin1 (Cul1) gene expression and cell proliferation in gastric cancer BGC823 and MGC803 cells and its molecular mechanism. Methods: Cul1 siRNA was synthesized in vitro and transfected into two kinds of gastric cancer cells under the guidance of siLentFect Lipid Reagent. The protein expression of Cul1, Cyclins and CDK inhibitor was detected by Western blotting. The cell proliferation was detected by CCK8 and the cell cycle was detected by flow cytometry. RESULTS: Cul1 protein expression was down-regulated by 39% and 84% in GCB823 and MGC803 cells transfected with Cul1siRNA, 59% and 62% in Cyclin D1, 47% and 54% in Cyclin E, and 52% And 23% respectively, but p21 protein expression did not change significantly. The proliferation ability of 24h2 gastric cancer cells decreased after Cul1 intervention (P <0.01), and decreased more significantly at 48 and 72h (P <0.01). The proportion of cells in 3 and 6 hG1 phase after Cul1 intervention decreased more slowly than that in control group (P <0.01), but there was no significant difference between Cul1 intervention group and control group in Sub-G1 phase. CONCLUSION: Cul1 siRNA can effectively interfere with the expression of Cul1 gene in gastric cancer BGC823 and MGC803 cells. Cul1 interference may affect the proliferation of gastric cancer cells by affecting the expression of Cyclins and CDK inhibitors in G1 phase.