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  • Title: Protection of H9c2 Myocardial Cells from Oxidative Stress by Crocetin via PINK1/Parkin Pathway-Mediated Mitophagy.
    Author: Chen J, Li YF, Zhang YS, Du TH, Lu Y, Li XY, Guo SW.
    Journal: J Vis Exp; 2023 May 26; (195):. PubMed ID: 37306415.
    Abstract:
    This study aimed to explore the oxidative stress-protective effect of crocetin on H2O2-mediated H9c2 myocardial cells through in vitro experiments, and further explore whether its mechanism is related to the impact of mitophagy. This study also aimed to demonstrate the therapeutic effect of safflower acid on oxidative stress in cardiomyocytes and explore whether its mechanism is related to the effect of mitophagy. Here, an H2O2-based oxidative stress model was constructed and assessed the degree of oxidative stress injury of cardiomyocytes by detecting the levels of lactate dehydrogenase (LDH), creatine kinase (CK), malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH Px). Reactive oxygen species (ROS)-detecting fluorescent dye DCFH-DA, JC-1 dye, and TUNEL dye were employed to assess mitochondrial damage and apoptosis. Autophagic flux was measured by transfecting Ad-mCherry-GFP-LC3B adenovirus. Mitophagy-related proteins were then detected via western blotting and immunofluorescence. However, crocetin (0.1-10 µM) could significantly improve cell viability and reduce apoptosis and oxidative stress damage caused by H2O2. In cells with excessive autophagic activation, crocetin could also reduce autophagy flow and the expression of mitophagy-related proteins PINK1 and Parkin, and reverse the transfer of Parkin to mitochondria. Crocetin could reduce H2O2-mediated oxidative stress damage and the apoptosis of H9c2 cells, and its mechanism was closely related to mitophagy.
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