Zinc homeostasis is essential for maintaining redox balance, cell proliferation, and apoptosis. However, excessive zinc exposure is toxic and leads to mitochondrial dysfunction. In this study, we established a zinc overload model by treating rat cardiomyocyte H9c2 cells with ZnCl2 at different concentrations. Our results showed that zinc overload increased LDH and reactive oxygen species (ROS) levels, leading to cell death, mitochondrial membrane potential decrease and impaired mitochondrial function and dynamics. Furthermore, zinc overload activated the PINK1/Parkin signaling pathway and induced mitochondrial autophagy via ROS, while NAC inhibited mitophagy and weakened the activation of PINK1/Parkin pathway, thereby preserving mitochondrial biogenesis. In addition, our data also showed that Mfn2 deletion increased ROS production and exacerbated cytotoxicity induced by zinc overload. Our results therefore suggest that ZnCl2-induced ROS generation causes mitochondrial autophagy and mitochondrial dysfunction, damaging H9c2 cardiomyocytes. Additionally, Mfn2 may play a key role in zinc ion-mediated endoplasmic reticulum and mitochondrial interactions.
Resveratrol (Res) has cardioprotective, anti-inflammatory, anti-aging and antioxidant effects; however, its mechanism remains unclear. Here we explored the protective effects of resveratrol on cardiomyocytes, focusing on the role of Zn2+ and mitophagy. Using the MTT/LDH assay, we found that addition of a zinc chelator TPEN for 4h induced mitophagy and significantly reduced cell viability, increased cytotoxicity and apoptosis in H9c2, while Res significantly inhibited these effects of TPEN. Similarly, Res inhibited TPEN-induced expression of mitophagy-associated proteins P62, LC3, PINK1, Parkin and TOM20. The mitophagy inhibitor 3-MA blocked the function of Res. At the same time, we found that Mfn2 siRNA could reverse the inhibition of mitophagy by Res through AMPK (adenosine 5'-monophosphate (AMP)-activated protein kinase) and prevent the effect of the mitochondrial permeability transition pore (mPTP) opening. Thus, our data suggest that Res can exert mitochondrial protection by inhibiting mitophagy and preventing mPTP opening through AMPK-Mfn2 axis in myocardial cells.
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