Effect and mechanism of METTL3 regulation of miR-126-5p on pyroptosis of human renal mesangial cells
Objective:To investigate the effect of methyltransferase-like protein 3(METTL3)on pyroptosis of human renal mesangial cells(HRMC)by regulating miR-126-5p and explore the underlying mechanisms.Meth-ods:HRMC was divided into 7 groups.Namely,control group,high-glucose treatment group,pcD-null group,pcD-METTL3 group,pcD-METTL3+miR-126-5p inhibitor(inhibitor)group,pcD-METTL3+inhibitor-NC gr-oup,and pcD-METTL3+inhibitor+AKT inhibition preparation(AZD5363)group.Cell pyroptosis was detected using flow cytometry,reverse transcription-quantitative PCR(RT-qPCR)was used to detect the expression of miR-126-5p,and western blotting was used to detect the expression of METTL3,Gasdermin D,NLRP3,phos-phorylated(p-)AKT,and p-NF-κB.RNA methylation immunoprecipitation(Me-RIP)was used to determine the N6-methyladenosine(m6A)modification level of miR-126-5p precursor.Results:Compared with the control group,cell pyroptosis,m6A modification of miR-126-5p precursor,as well as the expression levels of p-Akt and P-NF-κB were increased,while METTL3 and miR-126-5p expression levels were decreased in the high-glucose treatment group(all P＜0.05).After transfecting METTL3 overexpression vector,the above indicators were sig-nificantly reversed in the cells treated with high glucose(all P＜0.05).When the inhibitor of miR-126-5p was added after overexpressing METTL3,the expression of miR-126-5p was down-regulated,cell pyroptosis was in-creased,and the expression of p-AKT and p-NF-κB was up-regulated(all P＜0.05),but the expression levels of METTL3 and m6A modification of miR-126-5p precursor had no significant changes(P＞0.05).Adding AKT in-hibitor AZD5363 after overexpressing METTL3 resulted in reduced cell pyroptosis(P＜0.05),and down-regulat-ed expression of p-AKT and p-NF-κB,but the expression of METTL3 and miR-126-5p,as well as the m6A modi-fication of miR-126-5p precursor had no significant changes(P＞0.05).Conclusion:METTL3 promotes m6A methylation of miR-126-5p and reduces high glucose-induced pyroptosis of HRMC by inhibiting the AKT/NF-κB signaling pathway.
methyltransferase-like protein 3miR-126-5pN6-methyladenosinerenal mesangial cellscell py-roptosis
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