小鼠内毒素性肺损伤时SIRT3与线粒体功能的关系  

Relationship between SIRT3 and mitochondrial function in mice with endotoxin-induced lung injury

作　　者：谢子雷 杜越 史佳[1] 李娜[1] 李玉婷 刘沙沙[1] 张艳芳[1] 余剑波[1] Xie Zilei;Du Yue;Shi Jia;Li Na;Li Yuting;Liu Shasha;Zhang Yanfang;Yu Jianbo(Tianjin Medical University Nankai Hospital Department of Anesthesiology and Critical Care Medicine,TianjinNankai Hospital,Tianjin 300100,China)

机构地区：[1]天津医科大学南开临床学院天津市南开医院麻醉科和重症医学科,天津300100

出　　处：《中华麻醉学杂志》2022年第2期226-230,共5页Chinese Journal of Anesthesiology

基　　金：国家自然科学基金面上项目(81772106);天津市卫生健康委员会科技项目青年项目(QN20027);天津市自然科学基金面上项目(20JCYBJC00540)。

摘　　要：目的:评价小鼠内毒素性肺损伤时沉默信息调节因子2相关酶3(SIRT3)与线粒体功能的关系。方法:清洁级健康成年雄性野生C57BL/6(SIRT3^(+/+))小鼠20只,SIRT3基因敲除(SIRT3^(-/-))小鼠20只,体重20~25 g,6~8周龄,采用随机数字表法,将SIRT3^(+/+)小鼠和SIRT3^(-/-)小鼠分别分为4组(n=5):空白对照组(C组、SIRT3^(-/-)C组)、内毒素性肺损伤组(L组、SIRT3^(-/-)L组)、内毒素性肺损伤+白藜芦醇组(L+R组、SIRT3^(-/-)L+R组)、白藜芦醇组(R组、SIRT3^(-/-)R组)。L+R组、R组、SIRT3^(-/-)L+R组和SIRT3^(-/-)R组腹腔注射白藜芦醇15 mg/kg,1次/d,连续7 d,其余组给予等容量生理盐水。第7天注射白藜芦醇后30 min时L+R组与SIRT3^(-/-)L+R组、L组与SIRT3^(-/-)L组于相应时点尾静脉注射LPS 15 mg/kg制备内毒素性肺损伤模型,其余组注射等容量生理盐水。注射生理盐水或LPS后12 h时,于眼眶静脉丛采血,分别采用二甲酚橙法和ABTS比色法测定血清总氧化状态(TOS)和总抗氧化状态(TAS)水平,计算氧化应激指数(OSI);小鼠安乐死后取肺组织,行肺损伤评分,采用JC-1法测定线粒体膜电位(MMP),采用特异性荧光探针法测定细胞氧消耗率(OCR),采用Western blot法测定SIRT3表达水平。结果:与C组或SIRT3^(-/-)C组比较,L组与L+R组、SIRT3^(-/-)L组与SIRT3^(-/-)L+R组肺损伤评分、血清TOS浓度和OSI升高,TAS浓度、MMP和OCR降低,SIRT3表达下调(P<0.05)。与L组比较,L+R组肺损伤评分、血清TOS浓度和OSI降低,TAS浓度、MMP及OCR升高,SIRT3表达上调,而SIRT3^(-/-)L组肺损伤评分、血清TOS浓度和OSI升高,TAS浓度、MMP及OCR降低,SIRT3表达下调(P<0.05)。与L+R组比较,SIRT3^(-/-)L+R组肺损伤评分、血清TOS浓度和OSI升高,TAS浓度、MMP及OCR降低,SIRT3表达下调(P<0.05)。SIRT3^(-/-)L+R组与SIRT3^(-/-)L组上述指标比较差异无统计学意义(P>0.05)。结论:SIRT3表达下调可导致线粒体功能受损,参与小鼠内毒素性肺损伤的病理生理机制。Objective To evaluate the relationship between silent information regulator 2 homologue 3(SIRT3)and mitochondrial function in mice with endotoxin-induced lung injury.Methods Twenty clean-grade healthy adult male wild C57BL/6(SIRT3^(+/+))mice,20 SIRT3 knockout(SIRT3^(-/-))mice,weighing 20-25 g,aged 6-8 weeks,were studied.SIRT3^(+/+)mice and SIRT3^(-/-)mice were divided into 4 groups(n=5 each)according to the random number table method:blank control group(group C,group SIRT3^(-/-)C),endotoxin-induced lung injury group(group L,group SIRT3^(-/-)L),endotoxin-induced lung injury plus resveratrol group(group L+R,group SIRT3^(-/-)L+R),and resveratrol group(group R,group SIRT3^(-/-)R).Resveratrol 15 mg/kg was intraperitoneally injected once a day for 7 consecutive days in L+R,R,SIRT3^(-/-)L+R and SIRT3^(-/-)R groups,while the equal volume of normal saline was injected in the rest groups.Lipopolysaccharid 15 mg/kg was injected via the tail vein to develop a mouse model of endotoxin-induced lung injury at 30 min after resveratrol injection on 7th day,in L+R and SIRT3^(-/-)L+R groups and at the corresponding time points in L and SIRT3^(-/-)L groups,while the equal volume of normal saline was injected in the other groups.Blood samples were collected from the orbital venous plexus at 12 h after injection of normal saline or lipopolysaccharid for determination of serum total oxidation state(TOS)and total antioxidant state(TAS)levels by the xylenol orange method and ABTS colorimetric method,and the oxidative stress index(OSI)was calculated.After the mice were sacrificed,the lung tissues were taken for microscopic examination of the pathological changes which were scored and for determination of the mitochondrial membrane potential(MMP)(by JC-1 method),cellular oxygen consumption rate(OCR)(by the specific fluorescent probe method),and expression of SIRT3(by Western blot).Results Compared with group C or group SIRT3^(-/-)C,the lung injury score,serum TOS concentration and OSI were significantly increased,TAS concentration,MMP and

关 键 词：内毒素血症 急性肺损伤 线粒体 抗衰老酶3 

分 类 号：R563[医药卫生—呼吸系统]