线粒体靶向抗氧化剂Mito-Tempo对脓毒症小鼠急性肝损伤的保护作用

Protective effect of the mitochondria-targeted antioxidant Mito-Tempoagainst acute liver injury in mice with sepsis

目的:探讨线粒体靶向抗氧化剂Mito-Tempo对脓毒症小鼠急性肝损伤的影响及可能机制。方法:将C57BL/6小鼠随机分为对照组(Control组)、盲肠结扎穿刺组(CLP组)、Mito-Tempo治疗组(CLP+Mito-Tempo组)。采用盲肠结扎穿刺术(cecal ligation and puncture,CLP)构建小鼠脓毒症急性肝损伤模型,CLP+Mito-Tempo组在CLP处理前1 h使用Mito-Tempo腹腔注射进行预处理。24 h后麻醉并处死小鼠,取小鼠肝脏组织进行HE染色观察肝组织病理损伤;ELISA法监测血清中炎症因子,免疫荧光染色检测肝组织中活性氧(reactive oxygen species,ROS)的水平;电镜观察线粒体形态,Western blot法检测细胞焦亡相关蛋白剪切型含半胱氨酸的天冬氨酸蛋白水解酶-1(cleaved cysteinyl aspartate specific proteinase 1,cleaved-Caspases-1)、剪切型焦孔素D(cleaved-gasdermin D,GSDMD)、白细胞介素-1α(interleukin 1α,IL-1α)、白细胞介素-1β(interleukin 1β,IL-1β)的表达水平。结果:与Control组相比,CLP组肝损伤加重,ROS水平增加,线粒体形态出现破坏,细胞焦亡相关蛋白cleavedCaspase-1、cleaved-GSDMD、IL-1α、IL-1β的表达水平增加;与CLP组相比,CLP+Mito-Tempo组肝组织损伤程度减轻,ROS水平降低,线粒体形态部分恢复,细胞焦亡相关蛋白cleaved-Caspase-1、cleaved-GSDMD、IL-1α、IL-1β的表达水平降低。结论:Mito-Tempo可以减轻脓毒症小鼠急性肝损伤,其机制可能与降低氧化应激水平抑制细胞焦亡的发生有关。

Objective:To investigate the effect of the mitochondria-targeted antioxidant Mito-Tempo on acute liver injury in mice with sepsis and its possible mechanisms.Methods:C57BL/6 mice were randomly divided into control group(Control group),cecal ligation puncture group(CLP group),and Mito-Tempo treatment group(CLP+Mito-Tempo group).Cecal ligation and puncture(CLP)was used to establish a mouse model of sepsis and acute liver injury,and the mice in the CLP+Mito-Tempo group were pretreated with in-traperitoneal injection of Mito-Tempo at 1 hour before CLP treatment.After 24 hours,the mice were anesthetized and sacrificed,and HE staining was used to observe liver histopathological injury;ELISA was used to measure the serum levels of inflammatory factors,and immunofluorescent staining was used to measure the level of reactive oxygen species(ROS)in liver tissue;an electron microscope was used to observe the morphology of mitochondria;Western blot was used to measure the expression levels of cleaved cysteinyl aspar-tate specific proteinase 1(cleaved caspase-1),cleaved gasdermin D(GSDMD),interleukin-1α(IL-1α),and interleukin-1β(IL-1β).Results:Compared with the Control group,the CLP group had aggravation of liver injury,an increase in ROS level,destroyed mitochon-drial morphology,and increases in the expression levels of the pyroptosis-related proteins cleaved caspase-1,cleaved GSDMD,IL-1α,and IL-1β.Compared with the CLP group,the CLP+Mito-Tempo group had alleviation of liver injury,a reduction in ROS level,partial restoration of mitochondrial morphology,and significant reductions in the expression levels of the pyroptosis-related proteins cleaved caspase-1,cleaved GSDMD,IL-1α,and IL-1β.Conclusion:Mito-Tempo can alleviate acute liver injury in mice with sepsis,possibly by reducing the level of oxidative stress and inhibiting pyroptosis.