PDE4D Mediates the Cardiac Ischemia Reperfusion Injury and Is m6A Modified by METTL14.

BACKGROUND AND

OBJECTIVES

Cardiac ischemia-reperfusion (I/R) injury is a major contributor to morbidity following myocardial infarction. While PDE4D has been implicated in cerebral I/R injury, its role and regulatory mechanisms in the cardiac context remain unclear.

METHODS

PDE4D expression was analyzed using quantitative polymerase chain reaction and Western blot. Apoptosis was assessed by TUNEL staining, flow cytometry, and detection of caspase-3, Bax, and Bcl-2 levels. N6-methyladenosine (m6A) modification of PDE4D was examined via methylated RNA immunoprecipitation, luciferase reporter assays, and RNA stability measurements. The interaction between IGF2BP1 and PDE4D was confirmed by RNA immunoprecipitation.

RESULTS

PDE4D was significantly upregulated in I/R-injured myocardium and in H9c2 cardiomyocytes subjected to hypoxia/reoxygenation. Knockdown of PDE4D reduced infarct size, suppressed apoptosis, and improved cell viability. METTL14 mediated m6A methylation at 2 conserved sites within PDE4D mRNA, thereby enhancing its RNA stability. Silencing METTL14 decreased PDE4D expression and reduced its transcript stability. IGF2BP1 recognized m6A-modified PDE4D and promoted its RNA stability; ablation of IGF2BP1 abolished the effects of METTL14. Overexpression of PDE4D reversed the cardioprotective effects resulting from METTL14 silencing.

CONCLUSIONS

METTL14 stabilizes PDE4D mRNA through m6A methylation, which is recognized by IGF2BP1, thereby exacerbating cardiac I/R injury.