葡萄糖/葡萄糖氧化酶诱导H9c2心肌细胞凋亡模型的建立  

Establishment of models of cardiomyocyte apoptosis in H9c2 cells induced by glucose / glucose oxidase

作　　者：蔡诗婷[1,2,3] 刘晓颖[1,2] 谭虹虹[1,2] 陈少贤[1,2] 杨轶[1,2,3] 杨敏[1,2,4,2] 

机构地区：[1]广东省人民医院医学研究中心 [2]广东省医学科学院,广州510080 [3]汕头大学医学院,广东汕头515041 [4]广东省人民医院药学部

出　　处：《岭南心血管病杂志》2013年第5期611-615,共5页South China Journal of Cardiovascular Diseases

基　　金：广东省科技计划项目(项目编号:2009B080701054);广东省中医药管理局项目(项目编号:20122178)

摘　　要：目的研究不同浓度高糖(glucose,G)/葡萄糖氧化酶(glucose oxidase,GO)对H9c2细胞凋亡的影响,以确定建立H9c2细胞因糖自氧化所致凋亡模型的合适浓度。方法在含33 mmol/L葡萄糖的高糖培养液中以不同浓度的GO干预H9c2细胞12 h,MTT比色法检测H9c2细胞的存活率;光学显微镜下观察H9c2细胞形态学;AnnexinV/PI结合流式细胞仪检测凋亡率;2′,7′-二氯荧光黄双乙酸盐(DCFH-DA)探针检测细胞内活性氧簇(reactive oxygen species,ROS)水平。结果 (1)随G/GO浓度增加,H9c2细胞的存活率逐渐降低,其中1.6 mU/mL、2 mU/mL、2.4 mU/mL和2.8 mU/mL组存活率显著低于对照组,差异有统计学意义(P<0.05);其中G(33 mmol/L)/GO(2 mU/mL)的存活率为49.3%±6.7%。(2)随G/GO浓度增加,H9c2细胞的细胞凋亡率逐渐升高,细胞呈现典型的凋亡特征,其中1.6 m U/mL、2 mU/mL、2.4 mU/mL和2.8 mU/mL凋亡率显著高于对照组,差异有统计学意义(P<0.05);其中G(33 mmol/L)/GO(2 mU/mL)的细胞凋亡率为44.56%±3.02%。(3)G(33 mmol/L)/GO(2 mU/mL)作用H9c2细胞12 h时,ROS含量显著高于对照组,差异有统计学意义(荧光强度:47.70%±2.09%vs.17.59%±0.75%,P<0.05)。结论 G/GO引起的H9c2细胞氧化应激所致的凋亡有剂量依赖性,G(33 mmol/L)/GO(2 mU/mL)是建立H9c2心肌细胞凋亡模型的合适浓度。Objectives To investigate the effects of different concentrations of glucose （G）/glucose oxidase （GO） on the apoptosis of H9c2 cells,and to establish a model for oxidative stress-induced apoptosis in H9c2 cells.Methods The H9c2 cells were treated with 33 mmol/L glucose and serial concentrations of GO for 12 h.The viability of H9c2 cells was evaluated by MTT assay ; the morphology of H9c2 cells was observed under microscope ; the apoptosis was measured by flow cytometry and Annexin V/PI ; the generation of reactive oxygen species （ROS-superoxide anion and hydrogen peroxide） was measured by DCFH-DA assay.Results （1） The viability of H9c2 cells decreased with the increase of GO concentration,and those in 1.6 mU/mL group,2 mU/mL group,2.4 mU/mL group and 2.8 mU/mL group were significantly lower than that in control group （P〈0.05）,among which,that of G （33 mmol/L） / GO （2 mU/mL） was 49.3%±6.7%.（2） The apoptotic rate increased with the increase of G/GO concentration,and those in 1.6 mU/mL group,2 mU/mL group,2.4 mU/mL group and 2.8 mU/mL group were significantly higher than that in control group （P〈0.05）,among which,that of G （33 mmol/L） / GO （2 mU/mL） was 44.56%±3.02%.（3） As compared with that in control group,the level of ROS significantly increased 12 h after being treated with G （33 mmol/L）/GO （2 mU/mL） （fluorescence intensity：47.70%±2.09% vs.17.59% ± 0.75%,P〈0.05）.Conclusions The oxidative stress induced apoptosis caused by G/GO in H9c2 cells is dose-dependent,and G （33 mmol/L） / GO （2 mU/mL） may be the ideal concentration to establish the apoptosis model in H9c2 cells.

关 键 词：糖尿病 葡萄糖氧化酶 H9C2细胞 凋亡 

分 类 号：R587.1[医药卫生—内分泌]