WP1130通过抑制NLRP3炎症小体活化缓解小鼠的感染性休克

WP1130 relieves septic shock in mice by inhibiting NLRP3 inflammasome activation

目的探究小分子化合物WP1130抑制NLRP3炎症小体活化并缓解感染性休克的作用机制。方法细胞生物学水平:WP1130预处理小鼠骨髓来源巨噬细胞(BMDM)或人THP-1细胞后,利用多种NLRP3炎症小体活化剂(Nigericin、ATP、MSU、胞内LPS转染)活化NLRP3炎症小体,使用poly A:T活化AIM2炎症小体,通过Western blot和ELISA检测细胞培养上清中caspase-1和IL-1β的分泌水平,确定WP1130对NLRP3炎症小体的抑制效果及其特异性。利用激光共聚焦显微镜观察Nigericin诱导的线粒体损伤情况,探究WP1130是否影响NLRP3炎症小体组装活化的上游信号。动物水平:将SPF级雄性C57BL/6小鼠随机分为3组,即空白对照组(Control组)、感染性休克组(LPS组)、WP1130治疗组(WP1130+LPS组),ELISA检测小鼠血清和腹腔液中IL-1β、TNF-α的分泌水平。结果WP1130可剂量依赖性的抑制多种激动剂诱导的NLRP3炎症小体活化(P<0.05),但对非炎症小体相关炎性因子IL-6、TNF-α的分泌无显著影响(P>0.05)。此外,WP1130对AIM2炎症小体的活化无显著影响(P>0.05)。机制研究表明,WP1130并不影响NLRP3炎症小体组装活化的上游信号线粒体损伤。动物实验结果表明,相较感染性休克组(LPS组),WP1130治疗组(WP1130+LPS组)小鼠血清和腹腔液中IL-1β的水平显著降低(P<0.05),而TNF-α的分泌水平无统计学差异(P>0.05)。结论WP1130能够特异性抑制NLRP3炎症小体活化并缓解LPS诱导的小鼠感染性休克。

Objective To investigate the mechanism by which the small molecule compound WP1130 inhibits NLRP3 inflammasome activation and alleviates septic shock.Methods Mouse bone marrow-derived macrophages(BMDM)and human THP-1 cells were pre-treated with WP1130 before stimulation with different NLRP3 inflammasome agonists(Nigericin,ATP,MSU and intracellular LPS transfection),and AIM2 inflammasomes were activated with poly A:T.The levels of caspase-1and IL-1βin the cell culture supernatant were determined using Western blotting and ELISA,and mitochondrial damage in the cells was observed using confocal microscopy.In the animal experiment,male C57BL/6 mice were randomized into blank control group,septic shock group(LPS group)and WP1130 treatment group(WP1130+LPS group),and the levels of IL-1βand TNF-αin the serum and peritoneal cavity were detected using ELISA.Results In murine BMDM and human THP-1 cells,WP1130 significantly inhibited NLRP3 agonists-induced caspase-1 and IL-1βsecretion in a dose-dependent manner(P<0.05)but did not obviously affect the secretion of such inflammatory factors as IL-6 and TNF-αthat were not associated with inflammasomes(P>0.05).Treatment with WP1130 did not significantly affect poly A:T-induced activation of AIM2 inflammasomes(P>0.05)or induce obvious changes in mitochondrial damage,an upstream signal of NLRP3 inflammasome activation.In the mouse model of LPS-induced septic shock,WP1130 treatment significantly reduced the level of IL-1β(P<0.05)without obviously affecting TNF-αlevel either in the serum or in the peritoneal cavity(P>0.05).Conclusion WP1130 specifically inhibits NLRP3 inflammasome activation to alleviate LPS-induced septic shock in mice.