Abstract 358: Hypoxia and Hyperglycemia Induces M 6 A Mrna Methylation in Major Heart Cells

Abstract

Background: According to American heart association, over 70 % diabetic patients die from heart and stroke related diseases. The term “diabetic heart diseases” includes coronary heart disease, heart failure and diabetic cardiomyopathy in diabetic patients. Epigenetic and epitranscriptomic modifications play critical roles in progress of diabetic heart disease. Recent evidences indicated that m 6 A methylation involved in ischemic cardiomyopathy. But, the role of m 6 A mRNA methylation in cardiovascular diseases along with diabetic co-morbidity factors has not been studied in details. Thus, here we hypothesize that alterations in m 6 A mRNA methylation under hypoxic and hyperglycemic conditions contributes to severity of ischemic heart disease. Method and Results: To address our hypothesis, we have determined the levels of m 6 A mRNA methylation in NRVM, NRVF and HUVEC, HMVE and mouse primary endothelial cells under hypoxic and hyperglycemic conditions. We also examined the m 6 A levels in mice hearts post 5 days of MI. m 6 A mRNA methylation was significantly upregulated both in human and mouse ischemic hearts. Furthermore, hypoxia and hyperglycemia significantly induced m 6 A methylation in neonatal rat cardiomyocytes, fibroblasts, and mouse primary endothelial cells (isolated from WT and db/db mice). Next, we measured the methylation machinery both at RNA and protein levels. Interestingly, in corroboration with our methylation data, the expression of both m 6 A writers (Mettl3 and WTAP) and Readers (YTHDF2) was significantly increased. To determine the target transcripts which were highly methylated post-ischemia, we performed deep sequencing of methylated RNA after their immunoprecipitation using MeRIP-sequencing protocol. Our MeRIP-seq data has suggested a differentially m 6 A methylated targets both in-vitro and in-vivo ischemic sample. Conclusion: Over all, for the first time our data showed that hypoxia and hyperglycemia alters m 6 A mRNA methylation which may contribute to enhance the severity of cardiovascular diseases under hyperglycemic conditions. Further understanding of the mechanisms, may present a novel approach to potentially regulate m 6 A methylation, which may help in preventing/reducing heart failure in diabetic patients.