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Metformin ameliorates ferroptosis in cardiac ischemia and reperfusion by reducing NOX4 expression via promoting AMPKα
Zhenhua Wu1,2, Yunpeng Bai3, Yujuan Qi2
1Academy of Medical Engineering and Translational Medicine, Tianjin University, Tianjin, China.
Context:
Metformin (Met) has a protective effect against cardiac ischemia and reperfusion (I/R) injury.
Objective:
This study uncovered the Met effect on ferroptosis in cardiac I/R.
Materials And Methods:
Sprague-Dawley rats underwent cardiac I/R treatment (ischaemia 30 min; reperfusion 24 h) (I/R group), and administered intravenously with Met (200 mg/kg) (I/R + Met group). Haematoxylin-eosin staining, Prussian blue staining, immunohistochemistry and transmission electron microscope were conducted on cardiac tissues. H9c2 cells underwent oxygen-glucose deprivation/reoxygenation (OGD/R group) and treated by Met (0.1 mM) (OGD/R + Met group). Adenosine monophosphate-activated protein kinase α (AMPKα) siRNA was transfected into OGD/R-induced H9c2 cells. Cell counting kit-8 (CCK-8) assay, dichloro-dihydro-fluorescein diacetate (DCFH-DA) and JC-1 staining were conducted on H9c2 cells. Ferroptosis-related indicators and gene expression were detected by enzyme-linked immunosorbent assay (ELISA), quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and Western blot.
Results:
In cardiac I/R rat, Met decreased heart and serum MDA, cardiac and serum non-heme iron, and serum CK-MB and LDH (inhibition rate: 50.0%, 48.8%, 47.6%, 29.5%, 30.6% and 34.7%, respectively), relieved cardiac tissue ferroptosis and mitochondria damage, increased fraction shortening and ejection fraction (157.5% and 146.2% on day 28, respectively), up-regulated AMPKα and down-regulated NOX4 in cardiac tissues. In OGD/R-induced H9c2 cells, Met (0.1 mM) increased cell viability (promotion rate: 170.0%), decreased non-heme iron and MDA (inhibition rate: 30.1% and 47.9%, respectively), relieved ferroptosis, up-regulated AMPKα and down-regulated NOX4. AMPKα silencing abrogated these effects of Met on the OGD/R-induced H9c2 cells.
Discussion And Conclusions:
Met shows effectiveness in relieving ferroptosis in cardiac I/R. In the future, Met may be an effective drug for relieving ferroptosis in cardiac I/R patients clinically.
Insights
Metformin protects against cardiac ischemia-reperfusion injury by reducing ferroptosis. This study shows Metformin alleviates ferroptosis in cardiac I/R models, suggesting its clinical potential.
Area of Science:
- Cardiovascular Research
- Cellular Biology
- Pharmacology
Background:
- Metformin (Met) is known to protect against cardiac ischemia and reperfusion (I/R) injury.
- The specific mechanisms by which Metformin exerts its protective effects, particularly concerning ferroptosis in cardiac I/R, require further elucidation.
Purpose of the Study:
- To investigate the effect of Metformin on ferroptosis in the context of cardiac I/R.
- To explore the role of Adenosine monophosphate-activated protein kinase α (AMPKα) in Metformin's protective actions against ferroptosis in cardiac I/R.
Main Methods:
- Cardiac I/R models in Sprague-Dawley rats and oxygen-glucose deprivation/reoxygenation (OGD/R) models in H9c2 cells were utilized.
- Histological, biochemical, and molecular analyses including Haematoxylin-eosin staining, Prussian blue staining, immunohistochemistry, transmission electron microscopy, CCK-8 assay, DCFH-DA, JC-1 staining, ELISA, qRT-PCR, and Western blot were performed.
- AMPKα silencing was employed in H9c2 cells to assess its role in Metformin's effects.
Main Results:
- Metformin administration significantly reduced markers of ferroptosis, oxidative stress (MDA), and iron overload (non-heme iron) in both rat cardiac I/R and H9c2 cell OGD/R models.
- Metformin treatment improved cardiac function (fraction shortening, ejection fraction) in rats and cell viability in H9c2 cells.
- Metformin upregulated AMPKα and downregulated NOX4 expression, effects that were abrogated by AMPKα silencing.
Conclusions:
- Metformin demonstrates significant effectiveness in alleviating ferroptosis associated with cardiac I/R.
- The findings suggest that Metformin's protective effects are mediated, at least in part, through the AMPKα signaling pathway.
- Metformin holds promise as a potential therapeutic agent for managing ferroptosis in clinical scenarios of cardiac I/R.
