NLRP3炎症小体抑制剂MCC950改善非酒精性脂肪性肝炎形成的机制研究

Mechanism of NLRP3 inflammasome inhibitor MCC950 in improving the formation of nonalcoholic steatohepatitis

目的:研究NLRP3炎症小体小分子抑制剂MCC950在蛋氨酸胆碱缺乏(MCD)饲料诱导小鼠非酒精性脂肪性肝类(NASH)形成中的作用及分子机制。方法:C57BL/6小鼠用MCD饲料喂养4周的方式构建小鼠NASH模型。将实验分为Control组、单纯MCD饲养组(MCD组)以及MCD饲养且每隔2周经腹腔注射50 mg/kg的MCC950(MCC950组),4周后,测量各组小鼠及小鼠肝脏的质量,HE染色检测肝组织的病理改变,ELISA法检测血清中炎症因子IL-1β和TNF-α水平。提取小鼠肝脏枯否细胞(KCs),检测NLRP3、NLRP3上游因子TLR4以及NLRP3下游因子IL-1β和IL18的蛋白表达。结果:C57BL/6小鼠通过MCD饲料喂养4周的方式成功构建小鼠NASH模型,体现在MCD组小鼠肝细胞气球样改变、大量炎症细胞浸润以及大量纤维结缔组织增生,血清炎症因子显著升高。MCD喂养也诱导KCs NLRP3活化以及下游细胞因子IL-1β和IL18的表达。MCC950组小鼠质量和肝脏质量显著高于MCD组,肝脏病理改变也显著减轻,炎症细胞浸润减少,纤维结缔组织增生减少。腹腔注射MCC950可以显著抑制MCD喂养介导的小鼠KCs NLRP3的活化,抑制下游细胞因子IL-1β和IL18的表达,但对NLRP3上游因子TLR4的表达无显著影响。结论:MCC950可以显著抑制MCD诱导的小鼠NASH形成,其分子机制可能与MCC950靶向抑制KCs NLRP3活化相关。

Objective:To investigate the role and molecular mechanism of MCC950,a small molecule inhibitor of NLRP3,in the formation of NASH induced by MCD feeding in mice.Methods:The NASH model of mice was established by feeding of MCD for 4 weeks.The experiment was divided into control group,MCD feeding group and MCD feeding group with intraperitoneal injection of 50 mg/kg MCC950 every 2 weeks.4 weeks later,we measured the mass of mice and liver of mice.HE staining was used to detected the pathological changes of liver tissues,and ELISA was used to detected the inflammatory factors IL-1β and TNF-α in serum.The mouse liver KCs cells were extracted,and the protein expressions of NLRP3,NLRP3 upstream factor TLR4 and NLRP3downstream factors IL-1β and IL18 were detected.Results:The NASH model of mice was established successfully,which was reflected in balloon-like changes of liver cells,infiltration of a large number of inflammatory cells,proliferation of fibrous connective tissue and significant increase of serum inflammatory factors in MCD group.MCD feeding also induced KCs NLRP3 activation and the expression of downstream cytokines IL-1β and IL18.The weight of mice and liver in MCC950group were significantly higher than those in MCD group,and the pathological changes of liver were also significantly reduced,the infiltration of inflammatory cells and the proliferation of fibrous connective tissue were reduced.Conclusion:MCC950 can significantly inhibit the formation of NASH induced by MCD in mice,and its molecular mechanism may be related to the targeted inhibition of KCs NLRP3 activation by MCC950.