摘要
Objective:To determine the effects of berberine(BBR) on the activation of toll-like receptor 4(TLR4),nuclear factor(NF) κB(NF-κB) signaling and NLRP3 inflammasome in patients with gout.Methods:Peripheral blood mononuclear cells(PBMCs) were obtained from 24 acute(AP) and 41 non-acute(NAP) phases of primary gout patients,respectively,as well as 30 healthy controls(HC).TLR4,NF-κB(p65),NLRP3,apoptosis-associated specklike protein containing a CARD(PYCARD),cysteinyl aspartate specific proteinase-1(CASP1),and interleukin-1 β(IL-1 β) mRNA expression levels in PBMCs were measured by quantitative reverse transcriptase polymerase chain reaction.The protein levels of TLR4,myeloid differentiation factor 88(MyD88),NF-κB(p50/65),inhibitor of kappa B kinase α/β(IKKα/β),NF-κB inhibitor α(IKB α),phospho-IKKα/β(p-IKKα/β),NLRP3,PYCARD,and CASP1were monitored by Western blotting.Serum IL-1 β protein level was measured using enzyme-linked immunosorbent assay(ELISA).In addition,PBMCs from HC and macrophages derived from a spontaneously immortalized monocyte-like cell line(THP-1) were stimulated using monosodium urate(MSU,100 μ g/mL),0.1% dimethyl sulfoxide,25 μmol/L BBR,and 10,25,and 50 μmol/L BBR+100 μg/mL MSU for different time periods.The protein levels of IL-1β and IL-18 in cell culture supernatants was measured by ELISA,and the protein expressions of TLR4,MyD88,NF-κB(p50/p65),IKK α/β,IκBα,p-IKKα/β,NLRP3, PYCARD,and CASP1 in macrophages were analyzed by Western blotting.Results:(1) TLR4,NF-κB(p65),PYCARD,CASP1,and IL-1β mRNA levels in PBMCs were significantly higher in the AP group than in the HC group(P<0.05).The NLRP3 mRNA expression levels in PBMCs were found to be significantly lower in the AP and NAP groups than in the HC group(P<0.05,P<0.01).(2) The protein levels of TLR4,IKKβ,MyD88,NF-κB,p-IKKα/β,PYCARD,and CASP1 in PBMCs were significantly higher,and those of IκBα,IKKα,and NLRP3 were found to be significantly lower in the AP group than in the HC group(P<0.05 or P<0.01).(3) The serum IL-1β protein levels were significantly higher in the AP and NAP groups than in the HC group(P<0.01).(4)The IL-1β protein level was significantly lower in the culture supernatants of the PBMCs stimulated with MSU for 3 and 6 h in the 25 and 50 μmoL/L BBR groups compared with that in the MSU group(P<0.01).(5) The protein levels of IL-1β and IL-18 were also significantly lower in the culture supernatants of macrophages stimulated with MSU for 3 and 6 h in BBR groups compared with those in the MSU group(P<0.01).(6) The protein levels of TLR4, MyD88,NF-κB(p50,p65,p105),IKKα/β,p-IκBα,p-IKKα/β,PYCARD,and CASP1 were significantly differed between the macrophages stimulated with MSU for 0.5 and 6 h in BBR groups compared with those in the MSU group(P<0.05 or P<0.01).Conclusions:Activation of TLR4-NF κB signaling and NLRP3 inflammasome by MSU crystals drives the progression of gout inflammation.BBR ameliorates gouty inflammation,which is mechanistically associated with its regulation of TLR4-NF-κB signaling and NLRP3inflammasome expression.
基金
Supported by the Innovative Team Building Project of Chengdu Medical College(No.CYTD18-03)
the Key Projects of the First Affiliated Hospital of Chengdu Medical College(No.CYFY2019ZD01)
Project of Sichuan Provincial Administration of Traditional Chinese Medicine(No.2020JC0023)。
作者简介
Correspondence to:XU Dan,Prof.E-mail:447926416@qq.com。
