Aldo-Keto reductase 1C3 reduces myocardial cell damage after acute myocardial infarction by activating the Kelch-like ECH-associated protein 1-nuclear factor erythroid 2-related factor 2-antioxidant response element pathway to inhibit ferroptosis.

Abstract

Acute myocardial infarction (AMI) is a high-risk cardiovascular condition associated with increased cellular damage and oxidative stress. Aldo-Keto Reductase 1C3 (AKR1C3) is a stress-regulating gene. Nevertheless, its specific role and mechanisms regarding AMI remain unclear. We assessed cardiac function through echocardiography; tissue damage was evaluated using Hematoxylin and Eosin (HE) and Masson trichrome staining. AKR1C3 expression levels were measured through Reverse transcription-quantitative polymerase chain reaction and western blot. Assessed cell viability using Cell Counting Kit-8 and lactate dehydrogenase (LDH) assays. The extent of ferroptosis was determined by measuring the levels of Fe2+, boron-dipyrromethane (BODIPY) and malondialdehyde (MDA), the glutathione/glutathione disulfide (GSH/GSSG) ratio, and the expression of Glutathione Peroxidase 4 (GPX4) and Solute carrier 7A11 (SLC7A11). Kelch-like ECH-associated protein 1-Nuclear factor erythroid 2-related factor 2-Antioxidant response element (Keap1-Nrf2-ARE) pathway activation was analyzed through western blotting. Nrf2 was inhibited with ML385 and activated with (R)-Sulforaphane to investigate the Keap1-Nrf2-ARE pathway. The rats in the AMI group displayed reduced heart function, more tissue damage, and lower AKR1C3 expression compared to the Sham group. Similarly, hypoxia-treated H9C2 cells showed reduced viability, and decreased AKR1C3 expression. Overexpressing AKR1C3 in H9C2 cells enhanced viability. Knocking down AKR1C3 exhibited the opposite effect. Of the inhibitors tested, Ferrostatin-1 most effectively restored cell viability in hypoxia-treated H9C2 cells. Moreover, H9C2 cells subjected to hypoxia suggested Keap1-Nrf2-ARE pathway inhibition. Overexpressing AKR1C3 reduced ferroptosis and activated the Keap1-Nrf2-ARE pathway in hypoxia-treated cells, knocking down AKR1C3 exhibited the opposite effect. Further experiments using ML385 in hypoxia-treated H9C2 cells with overexpressed AKR1C3 showed decreased viability and increased ferroptosis compared to the control. Using (R)-Sulforaphane in hypoxia-treated H9C2 cells with knocked-down AKR1C3 exhibited the opposite effect. This study's findings indicate that AKR1C3 plays a role in regulating ferroptosis in myocardial cells, with the Keap1-Nrf2-ARE pathway likely being a key mechanism behind it.