BMAL1减轻H2O2诱导的心肌细胞损伤机制研究
Mechanism of BMAL1 attenuating H2O2-induced cardiomyocyte injury
易娜 1肖雯 2田源 3袁李礼3
作者信息
- 1. 长沙市第四医院心内科(邮编 410007)
- 2. 湖南师范大学生命科学学院
- 3. 湖南省脑科医院心内科
- 折叠
摘要
目的 探讨脑和肌肉组织芳香烃受体核转运蛋白的类似蛋白1(BMAL1)通过核因子E2相关因子2(NRF2)调节活性氧(ROS)/NOD样受体热蛋白结构域相关蛋白3(NLRP3)炎症小体通路对过氧化氢(H2O2)诱导的心肌细胞损伤的影响.方法 体外培养H9c2细胞和BMAL1稳定过表达的H9c2细胞,建立H2O2诱导的H9c2细胞损伤模型,并将细胞分为对照(Control)组、H2O2组、BMAL1过表达(BMAL1-OE)组、BMAL1过表达+H2O2(BMAL1-OE+ H2O2)组、BMAL1过表达+NRF2抑制剂(BMAL1-OE+ML385)组、BMAL1过表达+NRF2抑制剂+H2O2(BMAL1-OE+ ML385+H2O2)组.采用CCK-8法检测细胞活力,荧光探针2′,7′-二氯荧光素二乙酸酯检测ROS生成,Western blot检测BMAL1、NRF2和NLRP3蛋白表达,酶联免疫吸附试验法检测白细胞介素(IL)-1β释放.结果 与Control组相比,H2O2组H9c2心肌细胞活力减弱,ROS生成增多,BMAL1和NRF2蛋白表达水平降低,NLRP3蛋白表达水平升高,IL-1β释放增多(P＜0.05);与H2O2组相比,BMAL1-OE+H2O2组H9c2心肌细胞活力升高,ROS生成减少,BMAL1和NRF2蛋白表达水平升高,NLRP3蛋白表达水平降低,IL-1β释放减少(P＜0.05).与BMAL1-OE+H2O2组相比,BMAL1-OE+ML385+H2O2组H9c2心肌细胞活力减弱,ROS生成增多,NLRP3蛋白表达水平升高,IL-1β释放增多(P＜0.05).结论 BMAL1可减轻H2O2诱导的H9c2心肌细胞损伤,其机制可能与NRF2调节ROS/NLRP3炎症小体通路有关.
Abstract
Objective To investigate the effect of BMAL1 on H2O2-induced cardiomyocyte injury through NRF2-regulated ROS/NLRP3 inflammasome pathway.Methods H9c2 cells and H9c2 cells with stable over-expressed BMAL1 were cultured and divided into the control group,the H2O2 group,the BMAL1-OE group,the BMAL1-OE+H2O2 group,the BMAL1-OE+ML385 group and the BMAL1-OE+ML385+H2O2 group.All groups were pre-intervened with corresponding inhibitors,and then treated with 0.2 mmol/L H2O2,except for the control group and the BMAL1-OE group.After the intervention,CCK-8 assay was used to measure cell viability,fluorescent probe DCFH-DA was used to measure ROS generation and Western blot assay was used to detect BMAL1,NRF2 and NLRP3 protein expressions.ELISA was used to determine IL-1β release.Results Compared with the control group,the cell viability was decreased,ROS generation was increased,BMAL1 and NRF2 protein expressions were decreased,NLRP3 expression and IL-1β release were increased in the H2O2 group(P＜0.05).Compared with the H2O2 group,the cell viability was increased,ROS generation was decreased,BMAL1-OE and NRF2 protein expressions were increased,NLRP3 expression and IL-1β release were decreased in the BMAL1-OE+H2O2 group(P＜0.05).Compared with the BMAL1-OE+H2O2 group,the cell viability was decreased,ROS generation was increased,NLRP3 expression and IL-1β release were increased in the BMAL1-OE+ML385+H2O2 group(P＜0.05).Conclusion BMAL1 attenuates H2O2-induced H9c2 cardiomyocyte injury,and its mechanism may be related to the regulation of ROS/NLRP3 inflammasome pathway through NRF2.
关键词
ARNTL转录因子类/NF-E2相关因子2/活性氧/NLR家族,热蛋白结构域包含蛋白3/脑和肌肉组织芳香烃受体核转运蛋白的类似蛋白1/炎症小体Key words
ARNTL transcription factors/NF-E2-related factor 2/reactive oxygen species/NLR family,pyrin domain-containing 3 protein/BMAL1/inflammasome引用本文复制引用
基金项目
湖南省教育厅科研项目(21B0088)
湖南省卫生健康委科研项目(202203012960)
湖南省卫生健康委科研项目(202103010510)
湖南省自然科学基金(2022JJ80062)
湖南省自然科学基金(2022JJ80101)
湖南省中医药科技项目(D2022024)
出版年
2024
NETL
NSTL
万方数据