Engeletin Inhibits Inflammation and Ferroptosis and Attenuates Cardiomyocyte Injury Induced by Hypoxia-Reoxygenation.

Background: Investigate the function and mechanism of engeletin in myocardial ischemia reperfusion injury (MIRI).

Methods: Hypoxia-reoxygenation (HR) was achieved by subjecting H9c2 cells to 2 hours of hypoxia followed by 4 hours of reoxygenation. The viability of the H9c2 cells was measured by cell counting kit-8 assay. The expressions of interleukin-1 beta (IL-1β), interleukin-6 (IL)-6 and tumor necrosis factor-alpha (TNF-α) were detected by reverse transcription polymerase chain reaction. Reactive oxygen species (ROS) generation was detected by cell-permeable fluorogenic probe Dichloro-dihydro-fluorescein diacetate. Malondialdehyde, superoxide dismutase (SOD) and glutathione (GSH) levels were measured by corresponding kits. The accumulation of intracellular iron ions was accurately measured by the Iron Assay kit. Cell apoptosis was detected by Annexin V-FITC/Propidium Iodide staining. Protein expression was detected by Western blotting to investigate the activation of nuclear factor erythroid 2-related factor 2 (Nrf2) and nuclear factor-kappa B (NF-κB) signaling pathways.

Results: Engeletin treatment reversed the cell viability induced by HR, and also alleviated cell inflammation by inhibiting the expression of inflammatory cytokines, specifically IL-1β, IL-6 and TNF-α. Furthermore, engeletin treatment significantly inhibited the ROS generation induced by HR, inhibited MDA expression, and promoted SOD and GSH expressions. In addition, engeletin treatment decreased the intracellular concentration of ferrous iron, and promoted both glutathione peroxidase 4 and solute carrier family 7 member 11 expressions. The cell apoptosis results illustrated that engeletin significantly inhibited the apoptosis induced by HR. The Western blotting results showed that engeletin could activate the Nrf2 pathway and downregulate the NF-κB pathway. Engeletin alleviated MIRI in a left anterior descending artery mouse myocardial infarction model.

Conclusions: Engeletin functioned as a dual regulator both on NF-κB and Nrf2 pathways to alleviate the cell inflammation and ferroptosis induced by HR.