抑制核因子-kB活性对肝肺综合征大鼠肺血管内巨噬细胞iNOS表达的影响  

Effect of inhibiting nuclear factor-kB activity on expression of inducible nitric oxide synthase in pulmonary intravascular macrophages of rats with hepatopulmonary syndrome

作　　者：高君[1] 王宇[2] 张忠涛[2] 李建设[2] 马雪梅[2] 

机构地区：[1]首都医科大学附属北京朝阳医院京西院区肝胆胰脾外科,100043 [2]首都医科大学附属北京友谊医院普外科,100059

出　　处：《中华肝胆外科杂志》2009年第8期616-620,共5页Chinese Journal of Hepatobiliary Surgery

摘　　要：目的研究抑制核因子-kB（NFKB）活性对肝肺综合征（HPS）大鼠肺血管内巨噬细胞（PIM）内诱导型一氧化氮合酶（iNOS）表达的影响。方法Sprague—Dawley（SD）大鼠随机分为4组：对照组，吡咯醛二硫氨基甲酸（PDTC）对照组，CCl4组，CCl4＋PDTC组。对照组不做任何处理。CCl4组，肌注CCl4（3ml／kg·4d），共16次。CCl4＋PDTC组，注射CCl4第4次后，腹腔注射PDTC（20mg／kg·48h）。PDTC对照组，仅按CCl4＋PDTC组方法注射PDTC。取动脉血作血气分析，静脉血测内毒素和肝功能。取肠系膜淋巴结作细菌培养。应用免疫组化方法观察PIM的黏附、聚集情况，以及iNOS和NF—kB的蛋白表达和定位。以非放射性凝胶电泳阻滞实验（EMSA）检测NF—kB活性，sYBRGreen Ⅰ实时定量PCR方法检测肺组织中iNOS的mRNA表达。结果CCl4组发展成HPS模型，表现为PaO2、PaCO2降低和肺泡～动脉血氧分压差（A—aDO2）升高，肝功能异常，内毒素血症，所有数值都与对照组和PDTC对照组有显著性差异（P〈0．05）。CCl4组肠系膜淋巴结培养阳性率为62．5％（5／8），与CCl4＋PDTC组的66．7％（6／9）无统计学差异（P=1．000）。CCl4组中超过10个巨噬细胞的血管为60．8％（292／480），CCl4＋PDTC组中超过10个巨噬细胞的血管仅为19．6％（106／540），两组间存在非常显著性差异（P〈10．01）。对照组和PDTC对照组中肺血管内无巨噬细胞聚集。肺组织切片免疫组化染色检测NF-kB和iNOS蛋白均在CCI。组的PIM中表达。CCl4组的NF—kB活性明显高于对照组和PDTC对照组（P〈0．05）。CCl4＋PDTC组的NFKB活性明显低于CCl4组（P〈0.05），而与对照组和PDTC对照组之间无显著性差异（P〉0．05）。CCl4组的iNOS的mRNA表达明显高于对照组和PDTC对照组（P〈0．05）。CCl4＋PDTC组的iNOS的mRNA表达明显低于CCl4组（P〈0．05），而与对照组和PDTC对照组之间无显著性差�Objective To investigate the effect of inhibiting nuclear factor kB（NF-kB） activity on expression of inducible nitric oxide synthase（iNOS） in pulmonary intravascular maerophages （PIM） of rats with hepatopulmonary syndrome （HPS）. Methods The SD rats were randomized into four groups： control, control＋ PDTC, CCl4, CCl4＋PDTC groups. The rats in the control group received no pretreatment. In CCl4 group, CCl4 （3 ml/kg ·4day） was given by intramuscular injection and repeated for l6 times. In CCI4 ＋PDTC group, PDTC was given by intraperitoneal injection （20mg/kg ·48 h） after given CCl4 for 4 times. In control＋PDTC group, only PDTC was given according to that of CCl4 ＋PDTC group. Arterial blood was collected for measurement of blood gas. Venous blood was sampled for determination of hepatic function and endotoxin level. The mesenteric lymph nodes were dissected for bacteriological analysis. Immunolocalization of macrophages was performed using monoclonal macrophages antibody ED1 on paraffin sections. Proteins of NF-kB and iNOS of lung tissue were examined with immunohistochemistry. The activity of NF-kB in lung tissues was determined u sing electrophoretic mobility shift assay （EMSA）. By real-time polymerase chain reaction （PCR） u-sing SYBR Green I, the mRNA expression of iNOS in lung tissues was detected. Results CCl4 group developed HPS with decreased PaO2 and PaCO2 , increased alveolar arterial oxygen difference （AaDO2）, abnormal hepatic function and increased endotoxin level, which were significantly different from those in the control and control ＋PDTC groups （P〈0.05）. Culture positive mesenteric lymph nodes were found in 62.5% （5/8） of CCl4 group and 66.7% （6/9） of CCl4 ＋ PDTC group （P = l. 000）. There were no culture-positive mesenteric lymph nodes in control and control ＋PDTC groups. All lungs from CCl4 and CCl4 ＋PDTC group showed accumulation of large mononuelear macrophagelike cells within the lumen of numerous small muscular and nonmusc

关 键 词：肝肺综合征 肺血管内巨噬细胞 核因子-KB 诱导型氧化氮合酶 

分 类 号：R686[医药卫生—骨科学]