N-乙酰半胱氨酸抑制亚砷酸钠诱导心肌细胞凋亡和自噬作用的研究  

Protective effects of N-acetylcysteine on apoptosis and autophagy induced by sodium arsenite in H9C2 cardiac cells

作　　者：焦延娜 李建辉[1,2] 高小博[1,2] 骆海燕 马旭[1,2] 陆彩玲[1,2] 

机构地区：[1]国家卫生计生委科学技术研究所遗传优生中心,北京100081 [2]北京协和医学院研究生院

出　　处：《中国计划生育学杂志》2015年第5期299-304,共6页Chinese Journal of Family Planning

基　　金：国家自然科学基金(No.81300185.No.81470525);国家科技重大专项(2014ZX09304307-002)

摘　　要：目的:探讨N-乙酰半胱氨酸(NAC)抑制亚砷酸钠诱导H9C2细胞凋亡和自噬作用及机制。方法:用亚砷酸钠诱导H9C2细胞构建凋亡和自噬模型。采用CCK-8比色法检测细胞的存活率,Hoechst 33258核染色法观察凋亡细胞的形态和数量改变.双氯荧光素染色法检测细胞内活性氧(ROS)水平,Western Blot法测定蛋白P53、Bcl-2、Bax及LC3的表达水平,以及磷酸化ERK1/2蛋白及ERK1/2蛋白总量的表达水平。结果:与对照组相比,15μM亚砷酸钠处理的H9C2细胞存活率显著降低.细胞内ROS水平明显增加.凋亡细胞数量明显增多,自噬标志蛋白LC3-Ⅱ与LC3-Ⅰ比值明显增大(P<0.01)。1 mM NAC预处理后显著减少H9C2细胞内ROS的生成和促凋亡蛋白P53和Bax的表达,诱导抗凋亡蛋白Bcl-2表达升高,同时降低自噬相关蛋白LC3-Ⅱ与LC3-Ⅰ的比值。亚砷酸钠处理诱导ERK1/2蛋白磷酸化(P<0.01),而NAC和亚砷酸钠共处理抑制了ERK1/2蛋白的磷酸化。结论:亚砷酸钠降低H9C2细胞的存活率,增加细胞内ROS的生成,诱导细胞凋亡和自噬。NAC通过降低H9C2细胞内的ROS水平和抑制ERK1/2蛋白的磷酸化拮抗亚砷酸钠诱导的细胞凋亡和自噬。Objective.. To study effects of N-acetylcysteine （NAC） on apoptosis and autophagy induced by sodium arsenite in H9C2 rat myocardial cells and the underlying mechanism. Methods： H9C2 cells were divided as four groups： control group, sodium arsenite group, NAC group, arsenic and NAC group. Cell viability was measured by CCK-8 colorimetry. Changes in morphology and the amount of apoptotic cells were detected by Hoechst 33258 staining. Intracellular level of reactive oxygen species （ROS） was measured by DCFH-DA staining. Expressions of P53, Bcl-2, Bax and LC3 were evaluated by Western Blot, as well as the level of phosphorylation and total of ERK1/2 in H9C2 cells. Results： Compared with the control group, 15 btM sodium arsenite decreased cell viability, while increased the level of intracellular ROS and induced apoptosis and autoph- agy （P ~ 0.01）. And pre-treatment with 1 mM NAC decreased the expression of proapoptotic proteins, such as P53 and Bax, while increased the antiapoptotic protein, Bcl-2. And the ratio of LC3-II/LC3-I was also reduced. Further research showed that sodium arsenite induced the expression of phosphorylation of ERK1/2（P〈0.01）, while co-treatment with NAC and arsenic inhibited it. Conclusion： NAC could inhibit apoptosis and autophagy of H9C2 cells induced by sodium arsenite via reducing the ROS production and the level of phosphorylation of ERK1/2.

关 键 词：N-乙酰半胱氨酸 亚砷酸钠 H9C2心肌细胞 细胞凋亡 细胞自噬 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]