The demethylase FTO regulates the m6A modification and inflammation pathway in patients with cardiomyocytes af-ter hypoxia-reoxygenation

ZHAO Hai-mei; CHENG Hong; JIAO Qi-hui

摘要：Background Myocardial ischemia-reperfusion injury is an important clinical problem.Hypoxia-reperfusion(HR)triggers a series of complex cellular reactions and pathological changes.In recent years,the role of N-6-methyladenosine(m6A)modification in gene expression regulation has received extensive attention.New evi-dence suggests that m6A modifications may play a key role in the regulation of cellular responses to HR-dam-aged cardiomyocytes.Inflammation is an important part of the pathogenesis of myocardial ischemia-reperfu-sion injury.In the HR response,inflammatory factors such as interleukin(IL),tumor necrosis factor(TNF-a)and other cytokines are released and activated,which can trigger a series of intracellular signaling pathways leading to myocardial cell damage,apoptosis,and inflammation.In the context of cardiomyocyte H/R injury,it is important to study the relationship between m6A modification and the regulation of inflammatory cyto-kines expression.It is of great significance to fully understand the underlying mechanism of myocardial isch-emia-reperfusion injury and develop new therapeutic strategies.Methods The hypoxic-reoxygenation model of H9C2 cells was established,and the overexpression and knockout model of fat mass and obesity associated gene(FTO)was established.The cells were divided into 6 groups:control group(CG),hypoxic-reperfusion group(HRG),FTO overexpression group(OG),FTO overexpression hypoxic-reperfusion group(OHG),FTO knockout group(KG)and FTO knockout hypoxic-reperfusion group(KHG).The changes of M6A were detect-ed by colorimetry,FTO expression was detected by Western Blot,and the changes of FTO and phosphorylated nuclear factor kappa B(P-NF-κB)were observed by immunofluorescence.IL-6 was detected by enzyme-linked immunosorbent assay(ELISA).Results The level of m6A in HRG was significantly higher than that in CG(P＜0.05).The same pattern was observed in OG and KG(P＜0.05).The level of m6A in KHG were higher than that in HRG,while lower in OHG than that in HRG(P＜0.05).The expression of FTO decreased during hypoxia and reoxygenation(P＜0.05).The expression of FTO in KHG was lower than that in KG(P＜0.05).There was no significant difference in FTO expression between OG and OHG(P＞0.05).Compared with HRG,FTO in OHG was decreased(P＜0.01).Compared with HRG,FTO in KHG was decreased(P＜0.01).The changes of P-NF-κB and IL-6 were consistent before and after hypoxia reoxygenation.The expression of these two inflammatory factors in HRG was significantly higher than that in CG(P＜0.05).OHG compared with HRG,the two groups of inflammatory factors decreased;KHG was significantly higher than HRG(P＜0.05).Conclusions Overexpression of FTO can significantly inhibit the increase of m6A,reduce inflammato-ry factors,and protect damaged cells during hypoxia and re-oxidation processes.[S Chin J Cardiol 2024;25(3):169-178]