Study On The Mechanism Of Treponema Pallidum By P2X7 Receptor Activation Of NLRP3 Inflammasome And Promote Macrophages Secretion IL-1β Production

Objectives:Syphilis is the sexually transmitted disease caused by Treponema pallidum,the pathogenic mechanism of it is still unclear.IL-1β plays an important role in the body to fight infection,injury and immune response,the P2X7 receptor can modulate the NLRP3 inflammasome activation to affect the secretion of IL-1β.The aim of this study was to investigate the molecular mechanism of NLRP3 inflammasome activation and IL-1β maturation by Tp induced THP-1 macrophages,clarify the P2X7 receptor activation is an important factor in Tp induced NLRP3 inflammasome activation.Methods:Tp was cocultured with the THP-1-derived macrophages according to different MOI(multiplicity of infection)of 200:1,100:1,50:1,20:1.The mRNA expression of P2X7R,NLRP3,caspase-1 and IL-1β were investigated using Realtime-PCR.The protein expressions were quantified by Western blotting,and the mature IL-1β level in the supernatants were measured by ELISA.Then select the MOI 20:1 cocultured in Oh,2h,4h,6h,8h,12h,24h and 48h,collecting of the cells and the supernatant respectively.At last,THP-1 macrophages were treated with P2X7R agonist ATP(5mM),antagonist A438079(100μM),KCI(100mM)or using siRNA interference the expression of P2X7R/NLRP3,Realtime-PCR,Western-Blotting and ELISA were used to detect the expression of related signal molecule and observe the affect in Tp induced IL-1β secretion.Results:Tp could induce IL-1β production from the macrophages in a concentration dependent and time-dependent response manner within the tested range.When macrophages were systematically stimulated with Tp at MOI 20:1,the secretion of IL-1β were continues to increase with time,reached the highest level at 24h.The Realtime-PCR and Western blot results found that Tp could increase the expression of NLRP3,caspase-1 and IL-1β in mRNA and protein.Treatment with P2X7R agonist ATP significantly increased the secretion of IL-1β.After A438079,KCl processing macrophages,found that the production of IL-1β induced by Tp is significant inhibition.Furthermore,the secretion level of IL-1β were significantly decreased after Tp induced macrophages were treated with P2X7R and NLRP3 siRNA interference.Conclusions:Tp induces macrophages to secrete IL-1β,and Tp induced the secretion of the mature IL-1β is associated with the activation of the P2X7 receptor and the activation of NLRP3 inflammasome.