Effect and mechanism of dihydromyricetin on LPS/ATP-induced activation of NLRP3 inflammasome in vascular endothelial cells
Objective To investigate the effect of dihydromyricetin(DHM)on the activation of NLRP3 inflamma-some induced by lipopolysaccharides(LPS)/adenosine triphosphate(ATP)in vascular endothelial cells.Methods Hu-man umbilical vein endothelial cells(HUVECs)were cultured and randomly divided into the control group,LPS/ATP group,LPS/ATP+25 µmol/L DHM group,LPS/ATP+50 µmol/L DHM group and LPS/ATP+100 µmol/L DHM group,respectively.Except the control group,HUVECs in the other groups were treated with LPS(0.5 µg/mL)for 3.5 h and ATP(5 mmol/L)for 30 min.HUVECs in the DHM groups were pretreated with DHM(25,50,and 100 µmol/L)for 1 h before stimulation with LPS/ATP.The expression levels of NLRP3,ASC,pro-caspase-1,cleaved caspase-1 and silent in-formation regulator-1(SIRT1)were determined by Western blotting.Pathway enrichment analysis of SIRT1 was performed using Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG),target genes regulated by SIRT1 were analyzed and a network diagram was constructed using transcriptional regulatory network analysis;SIRT1 localization was detected by immunofluorescence;and the interaction mode between DHM and SIRT1 was analyzed by molecular dock-ing.Results Compared with the blank control group,the expression levels of NLRP3,ASC,and cleaved caspase-1 pro-tein were higher in the LPS/ATP group(all P<0.05);compared with the LPS/ATP group,the expression levels of NL-RP3,ASC,and cleaved caspase-1 protein were lower in all DHM groups(all P<0.05).SIRT1 was mainly distributed in the nucleus of cells.Compared with the blank control group,the relative fluorescence intensity was weaker in the LPS/ATP group(P<0.05);compared with the LPS/ATP group,the relative fluorescence intensity was stronger in the LPS/ATP+100 µmol/L DHM group(P<0.05).The LPS/ATP group had significantly lower expression of SIRT1 protein than the blank control group(P<0.05);compared with the LPS/ATP group and the LPS/ATP+25 µmol/L DHM group,SIRT1 protein expression levels were higher in the LPS/ATP+50 µmol/L DHM group and LPS/ATP+100 µmol/L DHM group(all P<0.05).GO enrichment analysis revealed that the bioprocesses enriched were closely related to nutrient level re-sponse,while the cellular component enriched were closely related to the chromatin silencing complex.The molecular function enrichment analysis had a relation to molecular functions such as chromatin-DNA binding.KEGG pathway enrich-ment analysis showed that SIRT1 was involved in longevity regulation,lipid and atherosclerosis signaling pathways.Fur-ther study indicated that DHM could be embedded into the binding pocket of SIRT1.Conclusion DHM inhibits LPS/ATP-induced activation of NLRP3 inflammasome probably by up-regulating the SIRT1 expression in HUVECs.
万方数据
维普
