脂多糖诱导脓毒症小鼠心肌细胞及线粒体自噬

发布时间：2018-02-11 06:49

  本文关键词： 脓毒症 心肌损伤 线粒体自噬 微管相关蛋白轻链(LC) pink parkin　出处：《细胞与分子免疫学杂志》2016年02期 　论文类型：期刊论文

【摘要】：目的探讨脓毒症心肌损伤时心肌细胞及线粒体自噬水平的变化。方法雄性C57BL/J小鼠随机分为空白对照组(NC)、LPS处理6、12、24、36 h组。LPS处理组小鼠腹腔注射10 mg/kg LPS建立脓毒症模型,空白对照组注射等量生理盐水。分别于以上时间点处死小鼠,收集血液及心脏组织,提取心肌组织胞质蛋白、线粒体及线粒体蛋白,另取心肌组织进行冰冻切片;应用ELISA试剂盒测定血清心肌肌钙蛋白I(c Tn I)含量变化;JC-1染色结合荧光酶标仪检测LPS刺激后不同时间点线粒体膜电位变化;Western blot法检测自噬相关蛋白微管相关蛋白1轻链3(LC3)、PTEN诱导激酶1(pink1)、E3泛素连接酶帕金森病蛋白(parkin)水平;免疫荧光组织化学染色检测心肌组织LC3、pink1/parkin蛋白的定位及其在不同处理组的表达差异。结果与对照组相比,LPS诱导的脓毒症小鼠血清c Tn I在6 h即开始明显升高;LPS处理组线粒体膜电位下降,在LPS 12 h组最低;细胞自噬相关蛋白LC3-Ⅱ/LC3-Ⅰ在腹腔注射LPS 12 h明显升高,然后逐渐下降;pink1/parkin在腹腔注射LPS 6 h明显升高,然后逐渐下降。结论脓毒症心肌损伤时心肌细胞及线粒体自噬水平升高,自噬应激可能更早发于心肌线粒体。
[Abstract]:Objective to investigate the changes of cardiac myocyte and mitochondrial autophagy during myocardial sepsis. Methods male C57BL / J mice were randomly divided into two groups: control group (n = 36) treated with LPS-treated with LPS-61212O / 24 h. LPS-treated mice were injected intraperitoneally with 10 mg/kg LPS to establish septic model. Mice were killed at the above time points, blood and heart tissues were collected, myocardial cytoplasmic protein, mitochondria and mitochondrial protein were extracted, and myocardial tissue was taken for frozen section. Detection of changes of serum cardiac troponin I c Tn I by ELISA kit and detection of mitochondrial membrane potential at different time points after LPS stimulation by using JC-1 staining and fluorescence enzyme labeling. Western blot method for the detection of autophagy associated protein microtubule-associated protein 1. The level of PTEN-induced kinase 1 Pink1PK E3 ubiquitin ligase Parkinsonase (Parkinin) was detected in light chain L3 LC3 and PTEN. Immunofluorescence histochemical staining was used to detect the localization of LC3pink1 / parkin protein and its expression in different treatment groups. Results compared with the control group, the serum c TN I of LPS induced sepsis mice began to increase significantly at 6 h after lipopolysaccharide (LPS) treatment. The mitochondrial membrane potential decreased in the group, In LPS 12h group, LC3- 鈪，

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