MR1 siRNA抑制人肝癌细胞BEL-7402的增殖  

MR1 siRNA suppresses proliferation of human hepatoma BEL-7402 cells

作　　者：王金华[1] 边春景[1] 赵春华[1] 

机构地区：[1]中国医学科学院北京协和医学院基础医学研究所组织工程研究中心,北京100005

出　　处：《基础医学与临床》2009年第1期1-5,共5页Basic and Clinical Medicine

基　　金：国家自然科学基金(30570771)

摘　　要：目的研究MR1基因对肝癌细胞增殖的影响及机制。方法化学合成MR1 siRNA,用脂质体lipofectamine2000转染肝癌细胞系BEL-7402细胞,RT-PCR检测MR1 siRNA基因沉默效果。用SRB法、集落形成法和生长曲线法检测MR1 siRNA对BEL-7402细胞增殖的影响。流式细胞术检测BEL-7402细胞周期,用细胞同步化的方法,即与G2期阻滞药物噻氨酯哒唑(nocodazole)联合应用,以间接反映MR1 siRNA对肿瘤细胞周期的影响。Western blot法检测Cyclin D1蛋白。结果(1)300 nmol/LMR1 siRNA处理BEL-7402细胞24 h后,MR1基因的mRNA水平明显降低。(2)经siRNA处理后,BEL-7402细胞存活细胞数明显下降,细胞生长速度明显减慢,集落形成率显著下降,Cyclin D1表达水平明显降低。(3)MR1 siRNA与nocodazole联合处理BEL-7402细胞后,G2期细胞比例显著减少,而G1期细胞明显增多。结论MR1基因与人肝癌BEL-7402细胞增殖相关,抑制MR1表达,能够引起细胞的增殖抑制,其作用机制与诱导细胞的G1期阻滞有关。Objective To identify the function and mechanism of MR1 in proliferation of BEL-7402 cells. Methods siRNA targeting MR1 and negative control siRNA were synthesized and transfected into BEL-7402 cells using Lipofectamine 2000. The silencing effect of MR1 siRNA was determined by semi-RT-PCR. SRB assay, colony formation assay and growth curve assay were used to investigate whether MR1 siRNA regulated cellular proliferation. Cell cycles were assessed by flow cytometry. G2 arrest reagent nocodazole was used to show the potential effect of MR1 siRNA on G1 arrest. The expression of Cyclin D1 was determined by Western blotting. Results （1）MR1 mRNA significantly decreased in BEL-7402 cells 24 h after MR1 siRNA transfection. （2）MR1 siRNA induced the down-regulation of cell growth. The expression of Cyclin D1 in MR1 siRNA tranfected BEL-7402 cells decreased significantly. （3）Flow cytometry results showed that MR1 siRNA markedly decreased G2 phase population with nocodazole treatment, and distinctly increased G1 phase population. Conclusion The gene MR1 is involved in the proliferation of BEL-7402 cells. MR1 siRNA causes inhibition of the proliferation of BEL-7402 cells. One of the mechanisms of MR1 siRNA on the proliferation of BEL-7402 cells is the induction of G1 arrest.

关 键 词：肌纤基因调节因子 SIRNA 细胞周期 细胞增殖 BEL-7402细胞 

分 类 号：R735.7[医药卫生—肿瘤]