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ALKBH5 Regulates SPHK1-Dependent Endothelial Cell Angiogenesis Following Ischemic Stress
Rajesh Kumari1, Roshan Dutta1, Prabhat Ranjan1
1Division of Cardiovascular Disease, Department of Medicine, The University of Alabama at Birmingham, Birmingham, AL, United States.
Insights
Endothelial cells dysfunction in heart disease is clarified by studying ALKBH5. This RNA demethylase maintains angiogenesis during ischemic injury by regulating SPHK1 methylation and eNOS signaling.
Area of Science:
- Cardiovascular Biology
- Molecular Biology
- Epigenetics
Background:
- Endothelial cell dysfunction is implicated in heart diseases like myocardial infarction and atherosclerosis.
- The precise molecular mechanisms underlying cardiac endothelial dysfunction remain unclear.
- Investigated the role of the m6A RNA demethylase ALKBH5 in endothelial cells (ECs) during ischemic injury.
Purpose of the Study:
- To elucidate the function of ALKBH5 in ECs angiogenesis following ischemic stress.
- To understand the molecular pathways regulated by ALKBH5 in the context of cardiac ischemia.
Main Methods:
- ECs were subjected to ischemic conditions (lipopolysaccharide and hypoxia).
- ALKBH5 function was assessed using siRNA-mediated gene silencing (loss-of-function).
- Investigated the impact of ALKBH5 on eNOS phosphorylation and SPHK1 protein levels and mRNA methylation.
Main Results:
- ALKBH5 expression is upregulated post-ischemia and supports ischemia-induced ECs angiogenesis.
- ALKBH5 is essential for maintaining eNOS phosphorylation and SPHK1 protein levels.
- ALKBH5 silencing increased SPHK1 m6A methylation; METTL3 overexpression reduced SPHK1 expression.
Conclusions:
- ALKBH5 plays a crucial role in maintaining angiogenesis in ECs after acute ischemic stress.
- This maintenance occurs through decreased SPHK1 m6A methylation.
- The downstream signaling pathway involves eNOS-AKT signaling.
Background:
Endothelial cells dysfunction has been reported in many heart diseases including acute myocardial infarction, and atherosclerosis. The molecular mechanism for endothelial dysfunction in the heart is still not clearly understood. We aimed to study the role of m6A RNA demethylase alkB homolog 5 (ALKBH5) in ECs angiogenesis during ischemic injury.
Methods And Results:
ECs were treated with ischemic insults (lipopolysaccharide and 1% hypoxia) to determine the role of ALKBH5 in ECs angiogenesis. siRNA mediated ALKBH5 gene silencing was used for examining the loss of function. In this study, we report that ALKBH5 levels are upregulated following ischemia and are associated with maintaining ischemia-induced ECs angiogenesis. To decipher the mechanism of action, we found that ALKBH5 is required to maintain eNOS phosphorylation and SPHK1 protein levels. ALKBH5 silencing alone or with ischemic stress significantly increased SPHK1 m6A mRNA methylation. In contrast, METTL3 (RNA methyltransferase) overexpression resulted in the reduced expression of SPHK1.
Conclusion:
We reported that ALKBH5 helps in the maintenance of angiogenesis in endothelial cells following acute ischemic stress via reduced SPHK1 m6A methylation and downstream eNOS-AKT signaling.
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