A1 and A2b adenosine receptors regulate GPX4 against ferroptosis of cardiomyocytes in myocardial infarction rat model and in vitro

Aim: The regulation of GPX4 by A(1)AR and A(2b)AR was investigated, and whether the inhibition of A(1)AR and A(2b)AR on ferroptosis of myocardial cell is related to GPX4 was also discussed. Methods: we constructed a rat model of myocardial ischemia and reperfusion (MIR) model and hypoxia/reoxygenation (H/R) model of H9C2 cells, and MIR rats were intraperitoneally injected with A(1)AR and A(2b)AR agonists and antagonists. TTC staining, DHE, TUNEL, western blot experiments, immunohistochemistry assay were implemented to analyze the influence of A(1)AR and A(2b)AR on ferroptosis and potential role of GPX4. To further authenticate the result of non-specific agonists and antagonists, we transfected siRNA interference or over expression vectors into cells. CCK8, flow cytometry and western blot were performed to evaluate cell proliferation and apoptosis, and the expression of GPX4 and ferroptosis-related proteins. Results: The experimental results showed that reduced expression of A(1)AR, A(2b)AR and GPX4 was found after MIR. A(1)AR and A(2b)AR activation by agonists increased GPX4 expression and decreased production of lipid ROS, further inhibiting apoptosis of cardiomyocytes. In addition, we also analyzed the effect of A(1)AR and A(2b)AR on ferroptosis-related proteins. We found that expression of FIH1 protein increased and expression of ACSL4 and NOX1 proteins decreased. Consistent with results in vivo, cellular data also indicated that A(1)AR and A(2b)AR overexpression could increase proliferation ability of H9C2, and inhibit apoptosis and ROS production, upregulate GPX4 and FIH1, and downregulate ACSL4 and NOX1. Conclusion: A(1)AR and A(2b)AR could regulate GPX4, thereby affecting ferroptosis of cardiomyocytes in a rat model of MIR.

Automated summary

This study found that A(1)AR and A(2b)AR can affect ferroptosis of myocardial cells in a rat model of MIR by regulating the expression of GPX4. Activation of the receptors increases the expression of GPX4 and reduces the production of lipid ROS, thereby inhibiting apoptosis of cardiomyocytes.