siRNA靶向沉默PcG蛋白EZH2对人胃癌细胞株MKN45增殖和侵袭力的影响

Effects of PcG Protein EZH2-targeting siRNA on Proliferation and Invasiveness of Human Gastric Cancer Cell Line MKN45

背景：PcG蛋白是一组转录抑制因子，其核心成分EZH2在胃癌组织中表达增高，并与胃癌的进展相关。目的：应用RNA干扰（RNAi）技术研究EZH2对人胃癌细胞株MKN45增殖和侵袭力的影响，从细胞水平探讨EZH2参与胃癌发生的机制。方法：使用EZH2小干扰RNA（siRNA）转染MKN45细胞，以实时逆转录聚合酶链反应（RT-PCR）和蛋白质印迹法检测EZH2mRNA和蛋白表达，CCK-8（Cell Counting Kit-8）实验检测细胞增殖情况，流式细胞仪分析细胞周期．MatrigelTM侵袭实验检测细胞侵袭力。结果：EZH2siRNA能有效抑制MKN45细胞的EZH2mRNA和蛋白表达。与阴性对照siRNA组相比，EZH2siRNA组MKN45细胞增殖抑制率为14．7％（P＜0．05），侵袭能力显著降低[穿过Transwell小室膜细胞数：（38．5±3．4）个／HPF对（92．7±9．7）个／HPF，P＜0．05），细胞周期阻滞于G0／G1和G2／M期。结论：EZH2siRNA可抑制人胃癌细胞的增殖和侵袭力，证实EZH2通过促进细胞增殖和侵袭在胃癌发生中发挥作用。

Background：Polycomb group （PcG） protein is a group of transcriptional repressor. EZH2, the crucial component of PeG protein is overexpressed in gastric cancer, and positively correlated with the progression of this malignancy. Aims： To study the effects of EZH2 on proliferation and invasiveness of human gastric cancer cell line MKN45, and to clarify the molecular mechanism of EZH2 in gastric tumorigenesis by using RNA interference （RNAi） technique. Methods： Small interfering RNA （siRNA） targeting to EZH2 was transfected into MKN45 cells, and then the expression levels of EZH2 mRNA and protein were detected by real-time reverse transcriptase polymerase chain reaction （RT-PCR） and Western blotting. The cell proliferation was determined by Cell Counting Kit-8 （CCK-8） assay, the cell cycle distribution was assessed by flow cytometry, and the invasive ability was examined by Matrigel^TM invasive assay. Results： EZH2 siRNA could effectively reduce the expression levels of EZH2 mRNA and protein in MKN45 cells. Compared with the siRNA negative controls, the proliferation of MKN45 cells in EZH2 siRNA group was obviously inhibited with an inhibition rate of 14.7% （P〈0.05）, the invasive ability was significantly decreased （ceils crossing the membrane of Transwell Chamber, 38.5±3.4 vs. 92.7±9.7 per high power field, P〈0.05）, and the cell cycle was arrested at G0/G1 and G2/M phases. Conclusions： EZH2 siRNA can reduce the proliferation and invasiveness of human gastric cancer cells, which suggests that EZH2 plays a role in gastric tumorigenesis by promoting cell proliferation and invasiveness.