背景:心力衰竭(Heartfailure,HF)是一种严重危害人类健康和生活质量的临床综合征,是心血管疾病的终末阶段。铁凋亡作为一种新的铁依赖性程序性细胞死亡模式,与心血管疾病的发生发展密切相关。已发现二氢乳清酸脱氢酶(DHODH)在抑制铁性凋亡和改善线粒体功能中起关键作用。其表达可被雌二醇(E2)上调。最近的研究发现DHODH可以通过将辅酶Q(CoQ)还原为CoQH2来抑制铁凋亡。因此,本研究旨在探讨DHODH上调对心力衰竭病理性肥大和纤维化的影响及其机制。
方法:采用主动脉横缩(TAC)法建立小鼠心力衰竭模型,老鼠的手术。手术后两天,皮下注射E2或相同体积的芝麻油8周.然后,超声心动图检测小鼠左心室收缩功能相关指标,组织学分析检测小鼠心肌纤维化程度;定量聚合酶链反应(q-PCR)和westernblot(WB)分析检测心力衰竭标志物的表达水平;透射电镜观察小鼠心肌细胞线粒体的形态学变化。用去氧肾上腺素刺激96小时建立细胞模型。通过试剂盒和WB分析检测铁凋亡标志物。通过JC-1荧光探针验证线粒体功能,和2\',7'-二氯二氢荧光素二乙酸酯(DCFH-DA)染色。转染后的小干扰RNA(siRNA)对CoQ的敲除成果用WB剖析检测。添加Fer-1作为阳性对照,以验证心肌细胞的铁凋亡相关变化。
结果:在动物模型中,我们发现,E2治疗通过促进小鼠心肌细胞DHODH上调,减轻了TAC诱导的心肌肥大和纤维化,并抑制了心肌细胞铁性凋亡.在细胞模型中,DHODH上调可防止去氧肾上腺素诱导的心肌细胞衰竭。然而,通过转染下调CoQ表达来抑制对上调DHODH的作用。
结论:DHODH的上调可有效改善TAC术后小鼠心肌肥厚、纤维化等心力衰竭的表现。抑制心肌细胞的铁性凋亡,并改善线粒体功能。该机制涉及CoQ相关的生物过程。
BACKGROUND: Heart failure (HF) is a clinical syndrome that seriously endangers human health and quality of life as the terminal stage of cardiovascular diseases. Ferroptosis as a new iron-dependent programmed cell death mode that is closely related to the occurrence and development of cardiovascular diseases. Dihydroorotate dehydrogenase (DHODH) has been found to play a crucial role in inhibiting ferroptosis and improving mitochondrial function, and its expression can be upregulated by estradiol (E2). Recent studies have found that DHODH can inhibit ferroptosis by reducing coenzyme Q (CoQ) to CoQH2. Therefore, this study aims to explore the effect of up-regulation of DHODH on the pathological hypertrophy and fibrosis of heart failure and its mechanisms.
METHODS: The mouse heart failure model was established by transverse aortic constriction (TAC), surgery in mice. Two days after the operation, a subcutaneous injection of E2 or the same volume of sesame oil was given for 8 weeks. Then, the left ventricular systolic function related indicators of mice were measured by echocardiography, and the degree of myocardial fibrosis of mice was detected by histological analysis; the expression levels of heart failure markers were detected by quantitative polymerase chain reaction (q-PCR) and western blot (WB) analysis; the morphological changes of mitochondria in cardiac cells of mice were observed by transmission electron microscopy. Cell model were established by stimulating with phenylephrine for 96 hours. Ferroptosis markers were detected by kits and WB analysis. Mitochondrial function was verified by a JC-1 fluorescent probe, and 2\',7\'-Dichlorodihydrofluorescein diacetate (DCFH-DA) staining. The knockdown results were detected by WB analysis after transfection of small interfering RNA (siRNA) of CoQ. Fer-1 was added as a positive control to verify the ferroptosis-related changes of myocardial cells.
RESULTS: In the animal model, we found that E2 treatment alleviates TAC-induced cardiac hypertrophy and fibrosis and suppresses cardiomyocyte ferroptosis by promotes DHODH upregulation in murine cardiomyocytes. In the cell model, DHODH upregulation protects against phenylephrine-induced cardiomyocytes with failure. However, the effect on up-regulating DHODH was inhibited by transfection to down-regulate CoQ expression.
CONCLUSIONS: The up-regulation of DHODH could effectively ameliorate the manifestations of heart failure such as myocardial hypertrophy and fibrosis in mice after TAC surgery, inhibit ferroptosis of cardiac myocytes, and ameliorate mitochondrial function. The mechanism involves CoQ-related biological processes.