Aerobic Exercise Improves Cardiac Pathological Remodeling in High-fat Diet Mice by Regulating m6A Methylation Modified circRNAs
Objective:To explore the role of N6-methyladenosine(m6A)methylation modified circular RNAs(circRNAs)in aerobic exercise-induced improvement of cardiac pathological remodeling in high-fat diet mice.Methods:C57BL/6J wild-type mice were randomly divided into four groups:Normal diet-sedentary(ND-SED)group,normal diet-exercise(ND-EX)group,high-fat diet-sedentary(HFD-SED)group and high-fat diet-exercise(HFD-EX)group.The ND-SED and ND-EX groups were fed with normal diet,while the HFD-SED and HFD-EX groups were fed with 60%fat-enriched high-fat diet for 12 weeks,the ND-EX and HFD-EX groups were subjected to 8-week aerobic exercise.After the exercise intervention,the body weight,body fat content,cardiac function,serum TC and TG levelsand heart tissue TG content were measured.m6A regulatory factors were detected by using RT-qPCR.Protein expressions of METTL3 and FTO were determined by using Western blot.m6A RNA methylation content in cardiac tissue was quantified using colorimetric methods.MeRIP-seq was used to detect circRNAs with m6A methylation modification in the heart.SRAMP was employed to predict circRNA methylation sites with m6A methylation difference and expression difference;and MeRIP-qPCR was performed to verify its m6A level.Results:8-week aerobic exercise significantly reduced body weight/fat content of high-fat diet-fed mice,and increased left ventricular ejection fraction(EF)and fractional shortening(FS),inhibited cardiac fibrosis,lipid deposition,and pathological remodeling.Aerobic exercise also decreased m6A methylation level in the heart of high-fat diet-fed mice,while the methyltransferase METTL3 with played an important role in this process.MeRIP-seq analysis revealed that the majority of m6A-modified circRNAs in the mice heart of HFD-SED and HFD-EX groups had only one m6A modification site,with a significant enrichment in overlapping regions and exons.Compared with the HFD-SED group,there were 13 upregulated and 19 downregulated m6A-modified circRNAs in that of HFD-EX group.The differential methylation of these circRNAs was mainly observed in overlapping regions,with most of them ranging from 1 to 10 000 bps and being located on chromosomes 1,7,and 13.Compared with the HFD-SED group,there were 40 circRNAs differentially expressed in the HFD-EX group,among which 22 circRNAs were significantly up-regulated and 18 circRNAs were significantly down-regulated.Combined analysis of MeRIP-seq and RNA-seq data revealed that only circRNA_4614 exhibited both differential m6A methylation levels and expression levels,showing a significant increase in both aspects.SRAMP prediction indicated that circRNA_4614 harbored eight potential m6A sites,among which one had high confidence while two had moderate confidence.Compared to the HFD-SED group,there was a significant increase in m6A methylation level for circRNA_4614 in the HFD-EX group,consistent with MeRIP-seq sequencing results.Conclusions:Aerobic exercise could significantly reduce m6A methylation levels in high-fat diet-fed mice,and regulate the expression profiles and m6A modification patterns of cardiac circRNAs.The involvement of circRNA_4614 mediated by m6A modification may play an important role in improving obesity-induced cardiac remodeling through aerobic exercise.
NETL
NSTL
万方数据
