The Molecular Mechanism Of Atorvastatin In Regulating NLRP1 Inflammasome

Objectives Atherosclerosis(As)is a chronic inflammatory disease.NLRP1 inflammasome can promote the secretion of inflammatory cytokines IL-1? and IL-18,which may play an important role in the inflammatory response of As.Atorvastatin is a HMG-Co A reductase inhibitor,which is widely used in clinical studies owing to its significant effect on lowering lipid.Recently,the strong antiinflammatory effect atorvastatin has been found,but the specific mechanisms are still unclear.In this study,the effects of atorvastatin on the expression of SREBP-1,NLRP1 and its related inflammatory cytokines were observed in mice.The roles of atorvastatin in regulating the mechanism of NLRP1 inflammasome were also investigated.Our studies provide new experimental basises for understanding the anti-inflammatory effects of atorvastatin and the clinical medication of Atherosclerosis.Methods Twenty-four Kunming male mice were randomly divided into four groups.Same volume of PBS coeliac injected in control group at the same time.The LPS group was injected i.p.with LPS(2mg/kg).The atorvastatin group was gavaged with atorvastatin(10mg/kg).InLPS+atorvastatin group,the mice challenged intraperitoneally(i.p.)with LPS(2mg/kg)and gavaged with atorvastatin(10mg/kg).All mice were treated for a week and executed,then collected blood and related tissues for further analysis.Triglyceride,total cholesterol,HDL-C,and LDL-C were determined by oxidase method.The levels of IL-1? and IL-18 were determined by ELISA.The m RNA expression of SREBP-1,NLRP1,Caspase-1 and ASC were detected respectively by RT-q PCR.The protein expression of SREBP-1,NLRP1,Caspase-1 and ASC in liver tissue were quantified by Western blot and immunohistochemistry,to clarify the effect of atorvastatin on the expression of SREBP-1 and NLRP1 in mice.Chromatin immunoprecipitation(Ch IP)was performed to detect if SREBP-1 was directly bound to the NLRP1 promoter.After treated with LPS and atorvastatin,we odserved whether the binding of SREBP-1 to NLRP1 was changed,to infer the role of atorvastatin in this process.The NLRP1 promoter mutant plasmids were constructed and then transfected into THP1 cells,after that the activation of NLRP1 promoter induced by SREBP-1 was analyzed by dual-luciferase reporter assay,to explore the molecular mechanism of atorvastatin mediated by SREBP-1 in the regulation of NLRP1.Results Compared with the control group,the levels of IL-1? and IL-18 in mice serum were up-regulated and the m RNA and protein expression of SREBP-1,NLRP1 and Caspase-1 were significantly increased in LPS group.Atorvastatin decreased the mature of IL-1? and IL-18,and inhibited the m RNA and protein expression of SREBP-1,NLRP1 and Caspase-1.The results of Ch IP confirmed that SREBP-1 could bind to the NLRP1 promoter directly.LPS elevated the activity of NLRP1 promoter,but the effect of atorvastatin was opposite.Dual luciferase reporter assay showed that atorvastatin inhibited the activity of NLRP1 promoter induced by LPS and/or SREBP-1 overexpression.After mutation the binding sites(-2041 to-2035)and(-424 to-418)of SREBP-1 in the NLRP1 promoter,the increased activity induced by LPS and/or SREBP-1 overexpression was inhibited,and its activation was further decreased after treatment with atorvastatin.These results indicated that atorvastatin could downregulate the activity of the NLRP1 promoter by suppressing the binding of SREBP-1 to NLRP1 promoter.Conclusion Atorvastatin decreases the levels of of IL-1? and IL-18 by inhibiting the expression of SREBP-1 and NLRP1 in mice,then exerts anti-inflammatiory effects.Atorvastatin regulates the activation and transcription of NLRP1 by inhibiting the combination between NLRP1 and SREBP-1 in THP-1 cells,then reduces the secretion of IL-1? and IL-18.