酸化脂肪酸酯通过抑制NLRP3炎症小体活化减少脓毒症致死  

Ozonated triglyceride protects against septic lethality via preventing the activation of NLRP3 inflammasome

作　　者：王丹[1,2] 刘袁洪[1,2] 宗秀乐 闫思聿 鲁建云 WANG Dan;LIU Yuanhong;ZONG Xiule;YAN Siyu;LU Jianyun(Department of Dermatology,Third Xiangya Hospital,Central South University,Changsha 410013;Medical Ozone Research Center,Central South University,Changsha 410013,China)

机构地区：[1]中南大学湘雅三医院皮肤科,长沙410013 [2]中南大学医学臭氧研究中心,长沙410013

出　　处：《中南大学学报（医学版）》2023年第6期809-820,共12页Journal of Central South University ：Medical Science

基　　金：supported by the National Natural Science Foundation(82202391),Outstanding Youth Project of Natural Science Foundation of Hunan Province(2022JJ20093);the Wisdom Accumulation and Talent Cultivation Project of the Third Xiangya Hospital of Central South University(YX202201),China.

摘　　要：目的:脓毒症是一种宿主对感染的反应失调,具有病死率高的特点,目前的治疗方法难以达到满意的疗效。研究表明臭氧治疗因其具有加强抗感染和免疫调节的作用,在多种感染及炎症相关疾病中起治疗作用。臭氧油是重要的臭氧治疗方式,其中关键成分为酸化脂肪酸酯。而酸化脂肪酸酯在脓毒症中的作用并不明确。本研究旨在探索酸化脂肪酸酯在脓毒症小鼠模型中的作用及其分子机制。方法:利用脂多糖(lipopolysaccharide,LPS)腹腔注射及盲肠结扎穿孔术(cecal ligation and puncture,CLP)构建小鼠脓毒症模型,取小鼠血清检测炎症因子,采集肺组织进行苏木精和伊红(hematoxylin and eosin,HE)染色,评估不同时间点及不同剂量酸化脂肪酸酯对脓毒症炎症因子释放及相关肺损伤的影响。在构建脓毒症模型后观察96 h,分析小鼠存活率。此外,用酸化脂肪酸酯处理原代腹腔巨噬细胞、人急性单核细胞白血病细胞系THP-1,采用酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)测定炎症因子水平,蛋白质印迹法检测半胱氨酸蛋白酶-1(caspase-1)和下游消皮素D(gasdermin-D,GSDMD)蛋白的裂解。结果:不同时间和剂量的酸化脂肪酸酯处理均可减少脓毒症小鼠炎症小体相关细胞因子白细胞介素(interleukin,IL)-1β和IL-18的释放(均P<0.05),但对细胞因子IL-6和肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)的释放无显著影响(均P>0.05)。酸化脂肪酸酯可显著提高脓毒症小鼠存活率,减轻脓毒症小鼠肺损伤(均P<0.05)。酸化脂肪酸酯可抑制经典及非经典途径诱导的NOD样受体热蛋白结构域相关蛋白3(NOD-like receptor family pyrin domain containing 3,NLRP3)炎症小体活化(均P<0.05),但不影响黑色素瘤缺乏因子2(absent in melanoma 2,AIM2)和含NLR家族CARD结构域4(NLR family CARD domain-containing protein 4,NLRC4)炎症小体的活化(均P>0.05)。酸化脂肪酸酯可抑制caspase-1Objective:Sepsis is a critical dysregulated host response with high mortality and current treatment is difficult to achieve optimal efficacy.Ozone therapy has been revealed to protect infection and inflammation-related diseases due to its role in antibiotic and immunoregulatory effect.Ozonated triglyceride is a key component of ozonated oil that is one of ozone therapy dosage form.However,the potential role of ozonated triglyceride in sepsis remains unclear.This study aims to explore the effect of ozonated triglyceride on septic mouse model and the molecular mechanism.Methods:Intraperitoneal injection of lipopolysaccharide(LPS),cecal ligation and puncture(CLP)were applied to construct septic mouse model.The mouse serum was obtained for detection of cytokines,and lung tissues were collected for hematoxylin and eosin(HE)staining to evaluate the extent of lung injury in septic mouse with ozonated triglyceride treatment at different time and doses.The survival of septic mice was observed for 96 h and Kaplan-Meier analysis was used to analyze the survival rates.In addition,primary peritoneal macrophages and human acute monocytic-leukemia cell line(THP-1)were treated with inflammasome activators with or without ozonated triglyceride.The level of cytokines was detected by enzyme-linked immunosorbent assay(ELISA).The cleavage of caspase-1 and gasdermin-D(GSDMD)was detected by Western blotting.Results:Ozonated triglyceride at different time and doses reduced the release of inflammasome-related cytokines[interleukin(IL)-1βand IL-18](all P<0.05)but not proinflammatory cytokines such as IL-6 and tumor necrosis factor-α(TNF-α)in septic mice(all P>0.05).Ozonated triglyceride significantly improved the survival rate of septic mice and reduced sepsis-induced lung injury(all P<0.05).Ozonated triglyceride significantly domain containing 3(NLRP3)inflammasome(all P<0.05)but not affected absent in melanoma 2(AIM2)and NLR family CARD domain-containing protein 4(NLRC4)inflammasomes in vitro(all P>0.05).Ozonated triglyceride reduced

关 键 词：脓毒症 酸化脂肪酸酯 NLRP3炎症小体 臭氧治疗 臭氧油 

分 类 号：R459.7[医药卫生—急诊医学]