背景:线粒体未折叠蛋白反应(UPRmt)是几种疾病中针对线粒体功能障碍的第一道防线。黄芩素,这是黄芩根的提取物,对代谢紊乱和心血管疾病具有丝裂性保护作用。然而,目前尚不清楚黄芩素是否通过UPRmt缓解肥胖引起的心脏损伤.
目的:本研究旨在阐明黄芩素在脂毒性诱导的心肌细胞凋亡中的作用,并探讨UPRmt相关机制。
方法:在体外实验中,建立棕榈酸(PA)处理的AC16心肌细胞以模拟肥胖诱导的心肌损伤。用黄芩素预处理AC16细胞后,细胞活力的水平,凋亡,线粒体膜电位,线粒体氧化应激,和UPRmt相关蛋白测定。此外,用ML385或siRNA处理AC16细胞以探索NRF2信号传导对UPRmt的调节。在体内实验中,用黄芩素给药8周的雄性db/db小鼠验证黄芩素对肥胖引起的心脏损伤的影响,UPRmt,和NRF2相关途径。
结果:在AC16心肌细胞中,PA剂量依赖性地增加UPRmt标志物(HSP60、LONP1、ATF4和ATF5)的表达。这种增加伴随着线粒体ROS的产生增加,线粒体膜电位降低,并提高了细胞色素c的表达水平,caspase-3和Bax/Bcl2被切割,最终导致细胞凋亡。黄芩素治疗可逆转UPRmt活化和线粒体损伤,并阻止线粒体介导的细胞凋亡。此外,NRF2通过其抑制剂ML385或siRNA下调黄芩素介导的NRF2信号激活和UPRmt抑制并引发线粒体功能障碍。此外,NRF2缺乏更强烈地激活UPRmt,导致线粒体氧化应激和PA诱导的心肌细胞凋亡,因此表明NRF2在线粒体稳态调节中起着至关重要的作用。在db/db小鼠的体内研究中,黄芩素抑制UPRmt,增强了抗氧化能力,并通过NRF2激活途径减轻心脏功能障碍。
结论:据我们所知,这些结果首次揭示了黄芩素通过激活NRF2信号传导抑制UPRmt,从而诱导针对脂毒性诱导的线粒体损伤和心肌细胞凋亡的保护作用,并提示NRF2在UPRmt调控中的新作用.
BACKGROUND: The mitochondrial unfolded protein response (UPRmt) is the first line of defense against mitochondrial dysfunction in several diseases. Baicalein, which is an extract of Scutellaria baicalensis Georgi roots, exerts mitoprotective effects on metabolic disorders and cardiovascular diseases. However, it remains unclear whether baicalein alleviates obesity-induced cardiac damage through the UPRmt.
OBJECTIVE: The present research designed to clarify the role of baicalein in lipotoxicity-induced myocardial apoptosis and investigated the UPRmt-related mechanism.
METHODS: In the in vitro experiment, palmitic acid (PA)-treated AC16 cardiomyocytes were established to mimic obesity-induced myocardial injury. After pretreatment of AC16 cells with baicalein, the levels of cell vitality, apoptosis, mitochondrial membrane potential, mitochondrial oxidative stress, and UPRmt-related proteins were determined. Additionally, AC16 cells were treated with ML385 or siRNA to explore the regulation of the UPRmt by NRF2 signaling. In the in vivo experiment, male db/db mice administered with baicalein for 8 weeks were used to validate the effects of baicalein on cardiac damage induced by obesity, the UPRmt, and the NRF2-related pathway.
RESULTS: In AC16 cardiomyocytes, PA dose-dependently increased the expression of UPRmt markers (HSP60, LONP1, ATF4, and ATF5). This increase was accompanied by enhanced production of mitochondrial ROS, reduced mitochondrial membrane potential, and elevated the expression levels of cytochrome c, cleaved caspase-3, and Bax/Bcl2, eventually leading to cell apoptosis. Baicalein treatment reversed UPRmt activation and mitochondrial damage and impeded mitochondrial-mediated cell apoptosis. Moreover, NRF2 downregulation by its inhibitor ML385 or siRNA diminished baicalein-mediated NRF2 signaling activation and UPRmt inhibition and triggered mitochondrial dysfunction. Additionally, NRF2 deficiency more intensely activated the UPRmt, resulting in mitochondrial oxidative stress and apoptosis of PA-induced cardiomyocytes, thus indicating that NRF2 plays a vital role in mitochondrial homeostasis regulation. In the in vivo study in db/db mice, baicalein inhibited the UPRmt, enhanced the antioxidant capacity, and attenuated cardiac dysfunction through a NRF2-activated pathway.
CONCLUSIONS: To our best knowledge, these results provide the first insight that baicalein inhibits the UPRmt to induce a protective effect against lipotoxicity-induced mitochondrial damage and cardiomyocyte apoptosis via activating NRF2 signaling and suggest a new role of NRF2 in UPRmt regulation.