Activation of Ferroptosis by miRNA-188-3p Mediated Downregulation of GPX4 Contributes to Ischemia/Reperfusion-Induced Renal Injury

Abstract

Background: Ferroptosis, a regulatory mode of cell death, has been extensively recognized as a crucial mechanism of ischemia/reperfusion (IR) that leads to acute kidney injury (AKI). However, the underlying mechanisms are not totally understood. This study aimed to explore the role of ferroptosis in IR-related acute kidney injury (AKI). Methods: Renal IR rat model and hypoxia/reoxygenation (HR)-induced HK-2 (human kidney 2) cell model were established. HK-2 cell viability was measured by CCK-8 (cell counting kit 8) assay. Ferroptosis activation was evaluated through Fe²⁺ and reactive oxygen species levels, and lipid peroxidization through glutathione level and malondialdehyde level. We used Western blot analysis to detect glutathione peroxidase 4 (GPX4) expression, and qRT-PCR (quantitative real-time polymerase chain reaction) to evaluate GPX4 mRNA and microRNA level. Luciferase reporter assay was applied to verify if miR-188-3p bound GPX4 directly. Results: Ferroptosis was observed in the renal tissue of IR rats. miR-188-3p expression correlated negatively with GPX4 expression in renal injury (R² = 0.4477). miR-188-3p knockdown and GPX4 overexpression inhibited HR-induced cell injury and ferroptosis. We further found that miR-188-3p could directly bind to the 3′UTR (untranslated region) of GPX4 mRNA, thus negatively regulating GPX4 expression. Conclusions: We demonstrated that miR-188-3p promoted ferroptosis in IR-associated AKI through GPX4 downregulation.