Peroxiredoxin III safeguards cardiac function against doxorubicin by regulating mitochondrial quality control via H2O2 detoxification

Abstract

Doxorubicin (DOX) is a widely used anticancer drug associated with severe cardiotoxicity primarily driven by mitochondrial dysfunction and reactive oxygen species (ROS) production. Hydrogen peroxide (H2O2), a major mitochondrial ROS, significantly contributes to cardiotoxicity, yet its precise mechanistic role in DOX-induced cardiotoxicity remains incompletely understood. Here, we investigated the critical role of mitochondrial H2O2 in DOX-induced cardiotoxicity using rat cardiomyocyte H9c2 cells with stable knockdown or reconstitution of mitochondrial antioxidant enzyme peroxiredoxin III (PrxIII), which specifically regulates mitochondrial H2O2 levels. Our results demonstrated that severe mitochondrial H2O2 accumulation (>10-fold compared to control) in PrxIII-depleted cells exacerbated mitochondrial oxidative stress, evidenced by increased cardiolipin oxidation and mitochondrial membrane potential dissipation. Furthermore, excessive mitochondrial H2O2 impaired mitochondrial fusion by reducing fusion-related protein expression, disrupted autophagic flux via lysosomal dysfunction, and significantly attenuated mitophagy, ultimately leading to enhanced apoptosis. Conversely, moderate mitochondrial H2O2 levels (5- to 8-fold increase compared to control) observed in PrxIII-expressing cells promoted mitochondrial elongation, enhanced mitophagy, and preserved autophagic flux, suggesting a protective adaptation against DOX-induced oxidative stress. In vivo experiments using PrxIII knockout mice confirmed that loss of PrxIII aggravated DOX-induced cardiac dysfunction. Bioinformatic analysis of independent public transcriptome datasets and targeted qPCR validation in PrxIII-deficient cardiac tissues further confirmed that the mitochondrial quality control pathways identified in vitro are robustly dysregulated in vivo. Additionally, PrxIII deficiency markedly increased mitochondrial structural damage without significantly affecting cardiac fibrosis or hypertrophy. Notably, mitigating the mitochondrial H2O2 burden and protecting the mitochondrial inner membrane using the mitochondria-targeted antioxidant peptide SS-31 successfully rescued this exacerbated cardiac dysfunction. In conclusion, our findings establish mitochondrial H2O2 as a pivotal determinant in DOX-induced mitochondrial dysfunction and cardiotoxicity, highlighting PrxIII as a promising therapeutic target for mitigating oxidative cardiac injury.