外源性硫化氢对内毒素血症大鼠肺组织线粒体途径细胞凋亡的影响  

Effect of exogenous hydrogen sulfide on mitochondrion-dependent apoptosis in lung tissues of rats with endotoxemia

作　　者：杜全胜[1] 任珊[1] 于娣[2] 张萌[3] 张楠[4] 李国风[5] 张建新[5] 

机构地区：[1]河北省人民医院重症医学科,石家庄市050051 [2]河北省胸科医院重症医学科,石家庄市050041 [3]河北医科大学第三医院胸外科,石家庄市050051 [4]河北省儿童医院药剂科,石家庄市050031 [5]河北省疾病预防控制中心药物研究所,石家庄市050021

出　　处：《中华麻醉学杂志》2017年第7期874-878,共5页Chinese Journal of Anesthesiology

基　　金：河北省自然科学基金（H2014307002）;河北省医学科学研究重点课题计划（ZD20140001）;河北省食品药品监督管理局2015年度食品药品安全科技项目计划（ZD2015009）;河北省中医药管理局科研计划项目（2015143）

摘　　要：目的 评价外源性硫化氢对内毒素血症大鼠肺组织线粒体途径细胞凋亡的影响.方法 健康雄性SD大鼠40只,2-3月龄,体重250-310 g,采用随机数字表法分为5组（n=8）：对照组（C组）、LPS组、低剂量硫氢化钠（NaHS）组（L-NaHS组）、中剂量NaHS组（M-NaHS组）和高剂量NaHS组（H-Na-HS组）.采用静脉注射LPS 5 mg/kg的方法制备内毒素血症模型.C组静脉注射等容量0.9％氯化钠溶液.L-NaHS组、M-NaHS组和H-NaHS组分别于静脉注射LPS后3h时腹腔注射硫化氢供体NaHS 0.78、1.56和3.12 mg/kg.于静脉注射LPS或0.9％氯化钠溶液后6h时处死大鼠,取肺组织,观察肺组织线粒体超微结构,采用流式细胞术检测肺细胞凋亡率,分别采用Western blot法和RT-PCR法检测肺组织caspase-3、caspase-9、Bcl-2和Bax及其mRNA的表达水平,计算Bcl-2和Bax表达水平的比值（Bcl-2/Bax比值）,采用Western blot法检测肺组织胞浆及线粒体细胞色素c（Cyt c）的表达水平.结果 与C组比较,LPS组细胞凋亡率升高,肺组织caspase-3、caspase-9、Bax和Bcl-2及其mRNA表达上调,Bcl-2/Bax比值降低,胞浆Cyt c表达上调,线粒体Cyt c表达下调（P〈0.05或0.01）;与LPS组比较,L-NaHS组、M-NaHS组和H-NaHS组细胞凋亡率降低,肺组织caspase-3、Bax及其mRNA表达下调,Bcl-2及其mRNA表达上调,Bcl-2/Bax比值升高,胞浆Cyt c表达下调,线粒体Cytc表达上调,M-NaHS组和H-NaHS组caspase-9及其mRNA表达下调,L-NaHS组caspase-9表达下调（P〈0.05或0.01）,caspase-9 mRNA表达差异无统计学意义（P〉0.05）,M-NaHS组和H-NaHS组线粒体超微结构损伤程度减轻.结论 外源性硫化氢抑制内毒素血症大鼠肺组织细胞凋亡的机制可能与抑制线粒体凋亡途径有关.Objective To evaluate the effect of exogenous hydrogen sulfide （H2S） on mitochondrion-dependent apoptosis in lung tissues of rats with endotoxemia.Methods Forty healthy male SpragueDawley rats,aged 2-3 months,weighing 250-310 g,were divided into 5 groups （n=8 each） using a random number table：control group （group C）,lipopolysaccharide （LPS） group,low dose sodium hydrosulphide （NaHS） group （group L-NaHS）,moderate dose NaHS group （group M-NaHS） and high dose NaHS group （group H-NaHS）.Endotoxemia was induced by Ⅳ LPS 5 mg/kg in chloral hydrate-anesthetized rats.The equal volume of 0.9% sodium chloride solution was intravenously injected in group C.NaHS （an exogenous donor of H2S） 0.78,1.56 and 3.12 mg/kg were intraperitoneally injected at 3 h after LPS injection in L-NaHS,M-NaHS and H-NaHS groups,respectively.The rats were sacrificed at 6 h after injection of LPS or 0.9% sodium chloride solution,and lungs were removed for examination of the mitochondrial ultrastructure of lung tissues and for determination of apoptosis in lung cells （by flow cytometry） and expression of caspase-3,caspase-9,Bcl-2 and Bax protein and mRNA in lung tissues （by Western blot or real-timne polymerase chain reaction）.The apoptosis rate and ratio of Bcl-2 expression to Bax expression （Bcl-2/Bax ratio） were calculated.The expression of cytochrome c （Cyt c） in cytoplasm and mitochondria of lung tissues was detected by Western blot.Results The apoptosis rate was significantly increased,the expression of caspase-3,caspase-9,Bcl-2 and Bax protein and mRNA was up-regulated,Bcl-2/Bax ratio was decreased,the expression of Cyt c in cytoplasm was up-regulated,and the expression of Cyt c in mitochondria was down-regulated in group LPS （P 〈0.05 or 0.01）.Compared with group LPS,the apoptosis rate was significantly decreased,the expression of caspase-3 and Bax protein and mRNA was down-regulated,the expression of Bcl-2 protein and mRNA was up-regulated,Bcl-2/Bax ratio was increased,the expressi

关 键 词：硫化氢 内毒素血症 肺 细胞凋亡 线粒体 

分 类 号：R563.8[医药卫生—呼吸系统]