Sdha基因敲低对小鼠肝细胞BNL CL.2细胞增殖、细胞周期和凋亡的影响  

Effect of succinate dehydrogenase complex subunit A gene knockdown on cell proliferation,cell cycle and apoptosis of mouse hepatic cell line BNL CL.2 cells

作　　者：李欣[1] 曹焕玲 赵亚伟[2] 郭银汉[1] 王庆阳[2] 

机构地区：[1]北京中医药大学中药学院,北京100029 [2]军事医学科学院基础医学研究所免疫学研究室,北京100850

出　　处：《中国药理学与毒理学杂志》2016年第2期107-112,共6页Chinese Journal of Pharmacology and Toxicology

基　　金：国家科技重大专项(2013ZX09103003)~~

摘　　要：目的探讨琥珀酸脱氢酶复合物亚单位A(SDHA)对小鼠肝细胞BNL CL.2细胞增殖、细胞周期和凋亡的影响。方法用慢病毒载体介导的sh RNA敲低BNL CL.2细胞中sdha基因的表达。流式细胞仪检测慢病毒的感染效率;实时荧光定量PCR和Western蛋白印迹法分别检测sdha m RNA和SDHA蛋白表达水平;细胞计数法检测细胞增殖;流式细胞术检测细胞凋亡和细胞周期。结果无相关sh RNA对照组和sdha sh RNA组慢病毒的感染效率均>80%。与无相关sh RNA对照组相比,感染sdha sh RNA慢病毒载体的BNL CL.2细胞sdha m RNA水平降低20倍左右(P<0.01),蛋白表达水平降低10倍左右(P<0.01);细胞增殖速度减慢,为无相关sh RNA对照组的70%左右(P<0.05);细胞周期发生改变,G0/G1期细胞百分率是无相关sh RNA对照组的1.17倍(P<0.01),G2/M期细胞百分率为1.37倍(P<0.01),S期细胞百分率为73.8%(P<0.01);但细胞凋亡率无明显差异。结论降低SDHA表达对小鼠肝细胞BNL CL.2细胞增殖有明显的抑制作用,其抑制作用与细胞周期阻滞相关,与细胞凋亡无明显关系。OBJECTIVE To investigate the effect of succinate dehydrogenase complex subunit A （sdha） gene on cell proliferation,cell cycle and apoptosis of mouse hepatic cell line BNL CL.2 cells.METHODS The BNL CL.2 cells were transfected by two kinds of sdha-shRNA lentivirus to knockdown sdha gene.The infection efficiency of BNL CL.2 cells infected with lentiviral vectors was analyzed by flow cytometry.The expression of sdha gene and SDHA protein was detected by real-time PCR and Western blotting,respectively.The effect of sdha gene on cell proliferation of BNL CL.2 cells was examined by growth curve,while cell cycle and apoptosis were analyzed by flow cytometry.RESULTS The infection efficiency of BNL CL.2 cells in sh-control group and in sdha-shRNA group was above 80%.Compared with sh-control group,the expression of sdha gene in BNL CL.2 cells infected with sdha-shRNA lentivirus was decreased by about 20 times （P〈0.01）,the expression of SDHA protein was decreased by about 10 times （P〈0.01）,and the growth rate was about 70% that of sh-control group （P〈0.05）.The cells were arrested in S phase,and the percentage of cells in S phase was 0.74 times that of sh-control group （P〈0.01）.The percentage of cells in G0/GI phase was 1.17 times that of sh-control group （P〈0.01）.The percentage of cells in G2/M was 1.37 times that of sh-control group （P〈0.01）.But there was no obvious difference in the apoptosis rate.CONCLUSION The reduced expression of SDHA protein can inhibit the proliferation of mouse hepatic cells,and the inhibitory mechanism may be cell cycle arrest.There is possibly no relationship between inhibition and cell apoptosis.

关 键 词：琥珀酸脱氢酶复合物亚单位A BNL CL.2细胞 细胞增殖 细胞周期 细胞凋亡 

分 类 号：R575[医药卫生—消化系统]