机械通气导致小鼠认知障碍时对自噬和NLRP3炎症小体的影响  

Effects of mechanical ventilation on autophagy and NLRP3 inflammasome in mice with cognitive impairment

作　　者：张晗[1] 陈婷[1] 陈畅[1] 唐李娟 陈莹莹[1] 钟琦[1] 张宗泽[1] ZHANG Han;CHEN Ting;CHEN Chang;TANG Lijuan;CHEN Yingying;ZHONG Qi;ZHANG Zongze(Department of Anesthesiology,Zhongnan Hospital of Wuhan University,Wuhan 430071,Hubei,China)

机构地区：[1]武汉大学中南医院麻醉科,湖北武汉430071

出　　处：《贵州医科大学学报》2022年第2期141-148,共8页Journal of Guizhou Medical University

基　　金：国家自然科学基金(81901109,81771160)。

摘　　要：目的探讨机械通气导致小鼠认知障碍时对自噬和Nod样受体热蛋白结构域相关蛋白3(NLRP3)炎症小体的影响。方法将8~12周龄、体质量20~25 g的SPF级健康雄性C57BL/6小鼠110只分为对照组(C组,n=26)、机械通气组(MV组,n=28)、机械通气+雷帕霉素处理组(Rap组,n=28)及机械通气+6-氨基-3-甲基嘌呤处理组(3-MA组,n=28);C组自主呼吸,MV组机械通气6 h,其余2组在机械通气前1 h分别给予0.001 mg/g Rap或0.015 mg/g 3-MA,再机械通气6 h;机械通气结束各组小鼠行Morris水迷宫实验和旷场实验,采用Western blot法检测各组小鼠海马组织微管相关蛋白1轻链3Ⅰ(LC3-Ⅰ)、微管相关蛋白1轻链3Ⅱ(LC3-Ⅱ)、自噬相关蛋白p62、Nod样受体蛋白3(NLRP3)、效应蛋白半胱氨酸天冬氨酸酶-1(Caspase-1)、凋亡相关点状蛋白(ASC)的表达,免疫荧光法检测海马小胶质细胞离子钙接头蛋白-1(IBA-1)的阳性细胞数。结果与C组比较,MV组小鼠各时点中央区停留时间延长、跨格次数减少、逃避潜伏期延长、目标象限停留时间百分比降低,海马组织p62表达降低,LC3-Ⅱ/LC3-Ⅰ比值升高、NLRP3、ASC、Caspase-1表达增加,小胶质细胞IBA-1的阳性细胞数增加(P<0.05);与MV组比较,Rap组小鼠各时点中央区停留时间缩短、跨格次数增加、逃避潜伏期缩短、目标象限停留时间百分比升高,海马组织p62表达增加,LC3-Ⅱ/LC3-Ⅰ比值降低、NLRP3、ASC、Caspase-1表达减少,小胶质细胞IBA-1的阳性细胞数减少(P<0.05);3-MA组小鼠各时点中央区停留时间延长、跨格次数减少、逃避潜伏期延长、目标象限停留时间百分比降低,海马组织p62表达降低,LC3-Ⅱ/LC3-Ⅰ比值升高、NLRP3、ASC、Caspase-1表达增加,小胶质细胞IBA-1的阳性细胞数增加(P<0.05)。结论机械通气可导致小鼠神经认知功能障碍,其作用机制可能与自噬介导的NLRP3炎症小体激活有关;增强自噬可通过增加NLRP3炎症小体的降解来改善机械通气�Objective To evaluate the effect of mechanical ventilation on autophagy and NOD-like receptor protein 3(NLRP3)inflammasome in mice with cognitive impairment caused by mechanical ventilation.Methods One hundred and ten SPF healthy male C57BL/6 mice aged 8 to 12 weeks with weight of 20 to 25 g were divided into control group(group C,n=26),mechanical ventilation group(group MV,n=28),mechanical ventilation+rapamycin treatment group(group Rap,n=28),mechanical ventilation+3-Methyladenine treatment group(group 3-MA,n=28)by using a random number table.The mice breathed spontaneously for 6 h in group C,and the mice were mechanically ventilated for 6 h in group MV,the other two groups were given autophagy agonist rapamycin(Rap)0.001 mg/g ip and autophagy inhibitor 3-Methyladenine(3-MA)0.015 mg/g ip 1 h before mechanical ventilation and followed by mechanical ventilation for 6 h.Morris water maze test and open field test were conducted at 1 and 3 days after the end of mechanical ventilation.Hippocampal tissues were obtained for determination of the expression of autophagy-related proteins(LC3-Ⅱ,LC3-Ⅰ,p62)and NLRP3 inflammasomes(NLRP3,Caspase-1,ASC)by Western blot and the number of positive cells of hippocampal microglia IBA-1 was detected by immunofluorescence.Results Compared with group C,the residence time in the central area was prolonged at each time point,the number of crossing the grid was reduced,the escape latency was prolonged,the percentage of residence time in the target quadrant was decreased,the expression of p62 was decreased,the ratio of LC3-Ⅱ/LC3-Ⅰwas increased,the expression of NLRP3,ASC,Caspase-1 increased,and the number of IBA-1 positive cells in microglia increased in group MV(P<0.05).Compared with the MV group,the residence time in the central area at each time point was shortened,the number of crossing the grid increased,the escape latency was shortened,the percentage of residence time in the target quadrant increased,the expression of p62 increased,the ratio of LC3-Ⅱ/LC3-Ⅰdecreased,and the

关 键 词：自噬 NLRP3炎症小体 呼吸 人工 细胞凋亡 脑 认知障碍 

分 类 号：R743.3[医药卫生—神经病学与精神病学] R749.1[医药卫生—临床医学]