Zhang Zhuhua, Zhou Mingxue, Liu Hongxu, Liu Wei, Chen Jiaping
Department of Cardiology, Beijing Hospital of Traditional Chinese Medicine, Affiliated to the Capital Medical University, Beijing, China.
Cardiovasc Diagn Ther. 2023 Apr 28;13(2):395-407. doi: 10.21037/cdt-23-86. Epub 2023 Apr 24.
Myocardial cell death resulting from ischemia-reperfusion (I/R) injury has been a predominant contributor to morbidity and mortality globally. The mitochondria-centered mechanism plays an important role in the formation of I/R injury. This study intended to discuss the protective mechanism of Shen Yuan Dan (SYD) on cardiomyocytes hypoxia-reoxygenation (H/R) injury via the regulation of mitochondrial quality control (MQC). Additionally, this study clarified the mechanism by which SYD suppressed mitophagy activity through the suppression of the PTEN-induced kinase 1 (PINK1)/Parkin pathway.
To induce cellular injury, H9c2 cardiomyocytes were exposed to H/R stimulation. Following the pretreatment with SYD, cardiomyocytes were subjected to H/R stimulation. Mitochondrial membrane potential (MMP), adenosine triphosphate (ATP), superoxide dismutase (SOD), and methane dicarboxylic aldehyde (MDA) were detected to evaluate the degree of cardiomyocyte mitochondrial damage. Laser confocal microscopy was applied to observe the mitochondrial quality, and the messenger (mRNA) levels of mitofusin 1 (Mfn1), mitofusin 2 (Mfn2), optic atrophy protein 1 (Opa1), dynamin-related protein 1 (Drp1), fission 1 (Fis1), and peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) in cardiomyocytes were assessed using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Western blotting was employed for the estimation of light chain 3 (LC3)-I, LC3-II, PINK1, and Parkin in cardiomyocytes.
It was discovered that SYD pretreatment elevated MMP in H/R injury cardiomyocytes, enhanced ATP content, activated SOD activity, and reduced MDA level. SYD treatment increased the mRNA levels of Mfn1, Mfn2, Opa1 and PGC-1α decreased the mRNA levels of Drp1 and Fis1, and reduced the protein levels of LC3, PINK1, and Parkin.
SYD plays a protective role in H/R injury to cardiomyocytes by regulating mitochondrial quality. Meanwhile, SYD may inhibit mitophagy activity through inhibiting the PINK1/Parkin pathway. This study provides insights into the underlying mechanism of SYD in alleviating myocardial I/R injury.
缺血再灌注（I/R）损伤导致的心肌细胞死亡一直是全球发病率和死亡率的主要原因。以线粒体为中心的机制在I/R损伤的形成中起重要作用。本研究旨在探讨参元丹（SYD）通过调节线粒体质量控制（MQC）对心肌细胞缺氧复氧（H/R）损伤的保护机制。此外，本研究阐明了SYD通过抑制磷酸酶和张力蛋白同源物诱导激酶1（PINK1）/帕金蛋白（Parkin）途径抑制线粒体自噬活性的机制。
为诱导细胞损伤，将H9c2心肌细胞暴露于H/R刺激。在用SYD预处理后，使心肌细胞接受H/R刺激。检测线粒体膜电位（MMP）、三磷酸腺苷（ATP）、超氧化物歧化酶（SOD）和丙二醛（MDA），以评估心肌细胞线粒体损伤程度。应用激光共聚焦显微镜观察线粒体质量，并用逆转录定量聚合酶链反应（RT-qPCR）评估心肌细胞中线粒体融合蛋白1（Mfn1）、线粒体融合蛋白2（Mfn2）、视神经萎缩蛋白1（Opa1）、动力相关蛋白1（Drp1）、分裂蛋白1（Fis1）和过氧化物酶体增殖物激活受体γ共激活因子-1α（PGC-1α）的信使核糖核酸（mRNA）水平。采用蛋白质免疫印迹法检测心肌细胞中轻链3（LC3）-I、LC3-II、PINK1和Parkin的表达。
发现SYD预处理可提高H/R损伤心肌细胞的MMP，增加ATP含量，激活SOD活性，并降低MDA水平。SYD处理增加了Mfn1、Mfn2、Opa1和PGC-1α的mRNA水平，降低了Drp1和Fis1的mRNA水平，并降低了LC3、PINK1和Parkin的蛋白水平。
SYD通过调节线粒体质量对心肌细胞H/R损伤起保护作用。同时，SYD可能通过抑制PINK1/Parkin途径抑制线粒体自噬活性。本研究为SYD减轻心肌I/R损伤的潜在机制提供了见解。
