作者：Xin; LIU; Yong; ZHANG; Bao-feng; YANG
摘要：OBJECTIVE To investigate the anti-pyroptotic effects of melatonin in atherosclerotic endothelium and to elucidate the potential mechanisms.METHODS ApoE-/-mice were randomly divid.ed into four groups（n=8）:the normal-diet group（ND）,the normal-diet group treated with melatonin(10 mg·kg-1)（ND+MLT）,the high-fat-diet group（HFD）,and the high-fat-diet group treated with melatonin（HFD+MLT）.After 12 weeks,the expression levels of pyroptosis related genes including NLRP3,ASC,cleaved-caspase 1,GSDMD-N,IL-1β and IL-18 were examined in aortic endothelium by Western blotting,qRT-PCR and immunofluorescent staining.Besides,levels of MEG3 and miR-223 were also tested by qRT-PCR.The interaction between MEG3 and miR-223 was detected by luciferase assay.For in vitro study,human aortic endothelial cells（HAECs） were transiently transfected with miR-223 mimic,miR-223 inhibitor（AMO-223）,MEG3-overexpressing plasmid or negative controls.After 6 h of transfection,the medium was replaced by fresh medium with or without ox-LDL（25 μg·mL-1） for 24 h and then treated with or without melatonin(10 μmol·L-1) for 48 h.Cell pyroptosis was evaluated by Hoechst 33342/PI staining and differentially expressed pyroptosis related genes.RESULTS Melatonin markedly reduced the atherosclerotic plaque in aorta of ApoE-/-mice.Meanwhile,melatonin also attenuated the expression NLRP3,ASC,cleaved-caspase1,NF-κB/GSDMD,GSDMD-N termini,IL-1β,and IL-18 in aortic endo.thelium.Consistent anti-pyroptotic effects were also observed in ox-LDL-treated HAECs.We found that lncRNAMEG3 enhanced pyroptosis in HAECs.Moreover,MEG3 acted as an endogenous sponge by sequence complementarity to suppress the function of miR-223 and to increase NLRP3 expression and enhance endothelial cell pyroptosis.Furthermore,knockdown of miR-223 blocked the anti-pyroptotic actions of melatonin in ox-LDL-treated HAECs.CONCLUSION Melatonin prevents endothelial cell pyroptosis via MEG3/miR-223/NLRP3 axis in atherosclerosis and therefore melatonin replacement might
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