eNOS基因转染对兔肺动脉高压及肺动脉高压危象的实验研究  

Effects of adenovius-mediated eNOS gene transfection on pulmonary artery hypertension and hypertension crisis in rabbits

作　　者：张凤伟[1] 吴树明[2] 曹广庆[2] 陆现硕[2] 

机构地区：[1]临沂市人民医院心外科,山东省276000 [2]山东大学齐鲁医院心外科

出　　处：《中国心血管病研究》2010年第7期528-533,共6页Chinese Journal of Cardiovascular Research

基　　金：2005年山东省自然科学基金资助项目（Y2005C30）

摘　　要：目的 探讨携带内皮型一氧化氮合酶（eNOS）基因的复制缺陷型重组腺病毒载体（AdCMVeNOS）转染对左向右分流所致的肺动脉高压及肺动脉高压危象兔模型的影响.方法 18只肺动脉高压模型兔随机分成实验组（9只）和对照组（9只）.实验组气管内滴入AdCMVeNOS病毒转染液（5×109 PFU/ml）2 ml,对照组气管内滴入生理盐水2 ml.4 d后手术结扎左向右分流,气管插管吸入10%氧气,检测两组模型缺氧前、后肺动脉收缩压（SPAP）、肺动脉平均压（MPAP）和平均动脉压（MAP）的变化.处死模型后,硝酸还原酶法测定肺组织的NO浓度,检测外源性eNOS基因mRNA及蛋白表达.结果 AdCMVeNOS转染后4 d和缺氧前,实验组肺动脉压较转染前明显下降（P〈0.01）,而对照组肺动脉压无明显改变,与实验组比较,差异具有统计学意义（P〈0.05）.实验组在缺氧1 h后,有4只兔出现肺动脉高压危象,对照组在缺氧30 min时,3只兔出现肺动脉高压危象,缺氧1 h后,全部出现肺动脉高压危象.两组在低氧通气后均出现氧分压（PaO2）降低,SPAP和MPAP升高,二氧化碳分压（PaCO2）升高,MAP降低.恢复供氧后,实验组各测得值恢复至正常水平,对照组SPAP和MPAP高于基础值,MAP低于基础值（P〈0.01）,实验组SPAP、MPAP和PaCO2的升高幅度及MAP和PaO2的降低幅度均低于对照组（P〈0.01）.免疫组化染色示,实验组肺泡内皮细胞、肺中小血管的平滑肌细胞和内皮细胞eNOS表达较对照组明显增加.实验组肺组织NO浓度明显高于对照组（P〈0.01）.RT-PCR产物经琼脂糖凝胶电泳后,实验组在3.77 kb处扩增出特异条带,而对照组在3.77 kb处未扩增出特异条带.Western blot分析显示,实验组兔肺组织eNOS（135 kDa）蛋白表达与对照组相比明显增加.结论 AdCMVeNOS转染左向右分流所致的兔肺动脉高压动物模型后,可有效降低肺动脉高压兔的肺动脉压和由缺氧诱发肺动脉高压危象的发�Objective The effects of transfer of the endothelial nitric oxide synthase （eNOS） gene to the lung were studied in both pulmonary hypertension rabbits induced by left to right shunt and pulmonary hypertension crisis rabbits induced by hypoxia. Methods Adenoviral vector encoding eNOS was intratraeheally transfeeted into the rabbit＇s lung with flow-induced pulmonary hypertension. Rabbits instilled intratraeheally with control vector containing no gene in the expression cassette served as control. Hemodynamic data were recorded before transfection,after transfeetion and before hypoxia, 10 rain, 30 rain, 1 h after hypoxia, and after inspired oxygen. The concentration of NO and transgene expression was investigated. Results There were no significant differences in SPAP and MPAP between two groups before transfection. Four days after intratraeheal administration of AdCMVeNOS, there was a significant reduction in SPAP and MPAP when compared with values obtained in control animals treated with saline （P〈0.01）. Pulmonary hypertension crisis occurred in 3 rabbits in control group 30 min after hypoxia, but no one was observed in AdCMVeNOS group. One hour after hypoxia, all rabbits presented pulmonary hypertension crisis in control group, and only 4 rabbits in AdCMVeNOS group developed pulmonary hypertension crisis. Comparing the variables after hypoxia versus before hypoxia in control group with AdCMVceNOS group, the increasing extent of SPAP, MPAP and PaC02 and decreasing extent of MAP, CO and Pa02 in AdCMVeNOS group were lower than that in control group. Immunohistochemical staining showed expression of the eNOS gene was detected mainly in endothelial cells of small pulmonary vessels. Transgene expression was confirmed using Western blot analysis. Conclusion Intratracheal adenoviral-mediated eNOS gene transfer to rabbit lungs reduces pulmonary hypertension induced by left-to-right shunt and decrease occurrence rate of pulmonary hypertension crisis.

关 键 词：一氧化氮合酶基因 腺病毒载体 肺动脉高压 基因治疗 兔 

分 类 号：R654.2[医药卫生—外科学] Q95-33[医药卫生—临床医学]