CAJ | 학술논문

背景：M2型丙酮酸激酶（ PKM2）在肿瘤组织中呈特异性高表达，与肿瘤的生长和糖酵解水平密切相关，是潜在的治疗靶点。目的：探讨siRNA沉默PKM2对胃癌SGC-7901细胞增殖、凋亡、迁移、侵袭以及糖酵解水平的影响。方法：选择未转染SGC-7901细胞、转染空质粒pU6 SGC-7901细胞以及转染PKM2 siRNA的SGC-7901细胞，采用蛋白质印迹法和Real-time PCR检测PKM2 mRNA和蛋白表达；免疫荧光法检测PKM2在细胞内的表达和分布；CCK-8法、流式细胞分析、细胞迁移和细胞侵袭实验检测细胞增殖、凋亡、迁移以及侵袭能力；蛋白质印迹法、分光光度法分别检测葡萄糖转运蛋白-1（Glut-1）、乳酸脱氢酶A（LDHA）蛋白表达以及葡萄糖、乳酸浓度、乳酸脱氢酶（LDH）活性。结果：与其余两组相比，PKM2 siRNA转染组细胞PKM2 mRNA和蛋白表达明显下降（ P<0.05），胞内表达明显减弱，细胞增殖、迁移和侵袭能力明显下降，细胞凋亡明显增加（ P<0.05），Glut-1、LDHA蛋白表达明显下降（ P<0.05），葡萄糖摄取率、乳酸产量、LDH活性明显减低（P<0.05）。结论：RNA干扰能抑制胃癌SGC-7901细胞PKM2 mRNA和蛋白表达，进而抑制胃癌细胞的增殖能力和糖酵解水平。
배경：M2형병동산격매（ PKM2）재종류조직중정특이성고표체，여종류적생장화당효해수평밀절상관，시잠재적치료파점。목적：탐토siRNA침묵PKM2대위암SGC-7901세포증식、조망、천이、침습이급당효해수평적영향。방법：선택미전염SGC-7901세포、전염공질립pU6 SGC-7901세포이급전염PKM2 siRNA적SGC-7901세포，채용단백질인적법화Real-time PCR검측PKM2 mRNA화단백표체；면역형광법검측PKM2재세포내적표체화분포；CCK-8법、류식세포분석、세포천이화세포침습실험검측세포증식、조망、천이이급침습능력；단백질인적법、분광광도법분별검측포도당전운단백-1（Glut-1）、유산탈경매A（LDHA）단백표체이급포도당、유산농도、유산탈경매（LDH）활성。결과：여기여량조상비，PKM2 siRNA전염조세포PKM2 mRNA화단백표체명현하강（ P<0.05），포내표체명현감약，세포증식、천이화침습능력명현하강，세포조망명현증가（ P<0.05），Glut-1、LDHA단백표체명현하강（ P<0.05），포도당섭취솔、유산산량、LDH활성명현감저（P<0.05）。결론：RNA간우능억제위암SGC-7901세포PKM2 mRNA화단백표체，진이억제위암세포적증식능력화당효해수평。
Background:Pyruvate kinase M2( PKM2 ),which is highly expressed in cancer cells,plays an important role in cancer cell growth and aerobic glycolysis and is a promising target for cancer therapy. Aims:To investigate the effect of silencing PKM2 by siRNA on proliferation,apoptosis,migration,invasion and glycolysis of gastric cancer cell line SGC-7901 cells. Methods:SGC-7901 cells were divided into three groups:non-transfectd SGC-7901 cells,SGC-7901 cells transfected with empty plasmid and SCG-7901 cellls transfected with PKM2 siRNA. Expression of PKM2 was detected by fluorescent quantitative PCR and Western blotting at mRNA and protein levels. Intracellular expression and distribution of PKM2 was determined by immunofluorescence. CCK-8 assay,flow cytometry,migration and invasion experiment were used to assess cell proliferation,apoptosis,migration and invasion,respectively. Western blotting was used to determine the expression of glucose transporter-1( Glut-1)and lactate dehydrogenase A( LDHA). Spectrophotometry was used to detect levels of extracellular glucose and lactic acid and intracellular lactate dehydrogenase( LDH). Results:Compared with non-transfected group and negative control( empty plasmid )group,PKM2 mRNA and protein expressions were significantly decreased in PKM2 siRNA plasmid transfected group(P<0. 05). Intracellular expression of PKM2 was markedly decreased in PKM2 siRNA cells. After transfected with PKM2 siRNA plasmid,cell proliferation,migration and invasion were significantly inhibited,and cell apoptosis was increased significantly( P<0. 05 ). Expressions of Glut-1 and LDHA were down-regulated in PKM2 silenced cells( P <0. 05 ). Glucose uptake,lactic acid production and LDH activities were significantly decreased in PKM2 silenced cells(P<0. 05). Conclusions:RNA interference targeting PKM2 gene inhibits PKM2 expression in human gastric adenocarcinoma cell line SGC-7901 cells,thereby inhibits cell proliferation and glycolysis.