NADPH氧化酶在Ang II介导H9C2心肌细胞凋亡中的作用

摘要

Objective To analyze the apoptosis of H9C2 cardiomyocytes induced by angiotensin II (Ang II) through NADPH oxidase/P38MAPK pathway.Methods H9C2 cardiomyocytes were cultured in vitro, and then divided into control group (added only nutrient fluid), Ang II group (added Ang II besides nutrient fluid), apocynin group (added apocynin besides nutrient fluid) and Ang II+apocynin group (added Ang II and apocynin besides nutrient fluid, eachn=6). The activity of NADPH oxidase was detected after 24 h, apoptosis was detected by using terminal-deoxynucleoitidyl transferase mediated nick end labeling (TUNEL), and expressions of P38MARK and relevant apoptotic protein were detected by using Western Blot method.Results The apoptotic rate was (12.20± 1.18)% in control group, (14.71±3.88)% in apocynin group, (62.33±4.79)% in Ang II group and (13.67±2.59)%in Ang II+apocynin group. Compared with control group, the apoptotic rate increased significantly in Ang II group, and compared with Ang II group it decreased in Ang II+apocynin group (allP<0.05). Compared with control group, the activity of NADPH oxidase decreased in apocynin group and increased significantly in Ang II group (P<0.05). The activity of NADPH oxidase decreased in Ang II+apocynin group compared with Ang II group (P<0.05). Compared with normal cells, the expressions of P38MAPK and apoptotic protein Bax increased, and expression of anti-apoptotic protein Bcl-2 decreased after Ang II effecting (allP<0.05). The expressions of P38MAPK and apoptotic protein Bax decreased, and expression of anti-apoptotic protein Bcl-2 increased in Ang II+apocynin group compared with Ang II group (allP<0.05).Conclusion The apoptosis of H9C2 cardiomyocytes is induced by Ang II through NADPH oxidase/P38MAPK pathway.%目的：分析血管紧张素Ⅱ（Ang Ⅱ）介导的H9C2心肌细胞凋亡是否通过NADPH氧化酶/P38MAPK途径。方法体外培养H9C2心肌细胞，分组干预：①Control组：仅加细胞培养液；②AngⅡ组：在Control组的基础上加入AngⅡ；③apocynin组：在Control组的基础上加apocynin；④AngⅡ+apocynin组：在Control组的基础上加入AngⅡ以及apocynin（n=6）。干预24 h后测定NADPH氧化酶活性，TUNEL法检测细胞凋亡，Western Blot 检测P38MARK及相关凋亡蛋白的表达水平。结果 Control组、apocynin组、AngⅡ组、Ang II+apocynin组凋亡比例分别为（12.20±1.18）%、（14.71±3.88）%、（62.33±4.79）%、（13.67±2.59）%。与Control组比较，AngⅡ组凋亡比例明显增加，而AngⅡ+apocynin组凋亡比例较AngⅡ组下降，差异有统计学意义（P均＜0.05）。与Control组比较，加入NADPH氧化酶抑制剂apocynin后细胞NADPH氧化酶活性降低；而仅加入AngⅡ的细胞NADPH氧化酶活性明显升高，差异有统计学意义（P＜0.05）。AngⅡ+apocynin组的NADPH氧化酶活性较AngⅡ组降低，差异有统计学意义（P＜0.05）。与正常细胞比较，AngⅡ作用后，P38MAPK表达增加，凋亡蛋白Bax表达增加，抗凋亡蛋白Bcl-2表达减少，差异有统计学意义（P均＜0.05）。Ang II+apocynin组较AngⅡ组P38MAPK表达降低，凋亡蛋白Bax表达减少，抗凋亡蛋白Bcl-2表达增加，差异有统计学意义（P均＜0.05）。结论血管紧张素II通过NADPH氧化酶/P38MAPK途径介导H9C2心肌细胞凋亡。