姜黄素通过AMPK/FUNDC1缓解LPS诱导的心肌细胞损伤

Protective Role of Curcumin on AMPK/FUNDC1-dependent Mitophagy Against LPS-induced Cardiotoxicity

目的 阐明AMPK/FUNDC1在姜黄素(curcumin, Cur)减轻脂多糖诱导的心肌细胞毒性中的作用及机制。方法将分离的乳鼠心肌原代细胞(neonatal mouse cardiomyocytes, NMCMS)构建脂多糖损伤模型。以CCK-8细胞活力检测试剂盒检测细胞活力、ROS检测试剂盒检测ROS生成量、ATP检测试剂盒检测线粒体ATP生成、JC-1荧光染色观察线粒体膜电位、Western blot法检测自噬标志蛋白的表达情况。结果 与Con组比较,LPS处理后显著降低心肌细胞活力且ROS水平显著增加,同时AMPK的磷酸化水平及线粒体自噬标志蛋白FUNDC1的表达显著降低。Cur处理后这种改变被显著逆转,从而保护NMCMS免受LPS诱导的心肌细胞毒性损伤。而siAMPK的加入则部分抵消Cur对心肌细胞的保护作用,此外线粒体自噬抑制剂3-MA及线粒体自噬激活剂rapamycin处理后表明,FUNDC1作为AMPK的下游的关键调控分子,参与Cur对LPS诱导的心肌损伤的保护作用。结论 Cur通过激活AMPK/FUNDC1信号通路,缓解LPS诱导的心肌细胞毒性。

Objective To elucidate the mechanism of AMPK/FUNDC1 signaling pathway in the protective role of curcumin(Cur) on LPS-induced cytotoxicity. Methods Neonatal mouse cardiac myocytes were isolated. Cell viability was detected by CCK-8 kit, ATP content and ROS production were detected simultaneously, and mitochondrial membrane potential was performed with JC-1 probe. Besides, Western blot was used to detect the expression of mitophagy marker proteins. Results Compared with Con group, LPS significantly decreased cell viability and increased ROS level, while the phosphorylation level of AMPK and the expression of mitochondrial autophagy marker protein FUNDC1 decreased significantly, which was significantly reversed by Cur treatment, and protect NMCMS against LPS-induced cytotoxicity. While siAMPK treatment partially counteracted the protective effect of Cur on LPS-induced cytotoxicity. In addition, the treatment of mitophagy inhibitor 3-MA and mitophagy activator rapamycin showed that FUNDC1, as a downstream molecule of AMPK, was the key regulatory factor of Cur on LPS-induced cytotoxicity. Conclusion Cur attenuates LPS-induced cytotoxicity by up regulating AMPK phosphorylation and activating FUNDC1-dependent mitophagy.