Balance of Superoxide and Nitric Oxide in Hyperoxic Lung Injury

高氧性肺损伤中超氧化物和一氧化氮的平衡

• 批准号: 10670560
• 负责人: TANIGAKI Toshimori
• 金额: $ 0.77万
• 依托单位: Tokai University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10670560/
• 关键词: hyperoxia; oxygen toxicity; acute lung injury; superoxide; nitric oxide synthase; selective inhibitor; L-NMMA; L-NIL; _L-NIL; _L-NMMA; 酸化窒素

EXPERIMENT 1: To study the interaction of superoxide (OィイD22ィエD2-) and nitric oxide (NO) production in the pathogenesis of acute lung injury, we controlled NO production in the isolated lung exposed to OィイD22ィエD2- by alienation of the L-arginine concentration in the perfusate following in vivo exposure to lipopolysaccharide (LPS). Rats were divided lo three groups (n=6 in each): (G1) saline+ィイD2LィエD2-Arginine (3mM), (G2) LPS+ィイD2LィエD2-Arginine (3mM), (G3) LPS+ィイD2LィエD2-Arginine (0.1mM). The animals in G2 and G3 received LPS 6 hr before isolation of the lungs. Xathine oxidase (0.1 U/ml) was infused to generate OィイD22ィエD2- and 2-methyl-6-(p-methoxyphenyl)-3,7-dihydro-imiduo (1,2-a) pyrazin-3-one (MCLA; 6mM), OィイD22ィエD2-specific chemiluminescence probe was added to the perfusate. We estimated lung injury by measuring the concentration of lactate dehydrogenase (LDH) in the perfusate. Nitrite and nitrate in the perfusate were assayed using the Griess reaction. LPS administration increased O … More ィイD22ィエD2- production only slightly. Both NO production and LDH concentration increased significantly in G2 but not in G3 compared with G1. In conclusion, excess production of NO may exacerbate ROS mediated acute lung injury.EXPERIMENT 2: The role of nitric oxide (NO) in pathogenesis of oxidant lung injury remains uncertain. We investigated the effect of two NO synthase (NOS) inhibitors, ィイD2LィエD2-NィイD16ィエD1-(1-iminoethyl)lysine (ィイD2LィエD2-NIL), highly selective inhibitor of iNOS, and ィイD2LィエD2-NG-monomethyl arginine (ィイD2LィエD2-NMMA), non-selective NOS inhibitor on hyperoxic lung injury in rats. Animals received ィイD2LィエD2-NIL, ィイD2LィエD2-NMMA or diluent intraperitoneally every 12 hr beginning 12 hr before exposure to 100% oxygen (OィイD22ィエD2) or air for a period of 50 hr. Absorption of the NOS inhibitors was confirmed by measurement of changes in the nitrite/nitrate concentration in serum. We measured wet-to-dry weight ratio (W/D) of lung tissue as an index of pulmonary edema and total protein concentration in bronchoalveolar lavage fluid (BALP) as an index of lung protein flux. We also evaluated hyperoxia-induced alternations in inducible NOS (iNOS) and endothelial NOS (eNOS) mRNA expression in lung tissue. OィイD22ィエD2 exposure increased W/D, BALP and serum nitrite/nitrate concentration. ィイD2LィエD2-NIL and ィイD2LィエD2-NMMA attenuated the OィイD22ィエD2 induced increases in nitrite/nitrate concentration but only ィイD2LィエD2-NMMA produced further significant increases in W/D and BALP above those with OィイD22ィエD2 exposure. Exposure to 100% OィイD22ィエD2 increased the expression of both iNOS and eNOS mRNA and this effect was not altered by ィイD2LィエD2-NIL or ィイD2LィエD2-NMMA, which acts as competitive NOS inhibitors. In conclusion, ィイD2LィエD2-NMMA but not ィイD2LィエD2-NIL accentuates hyperoxic lung injury suggesting that endogenous endothelial NO play a protective role in pulmonary oxygen toxicity. Less

实验1：为了研究超氧阴离子（O_（22）与一氧化氮（NO）的相互作用在急性肺损伤发病机制中的作用，我们用O_（22）D_（2-）处理离体肺，通过在体暴露于脂多糖（LPS）的灌流液中L-精氨酸浓度的异化来控制NO的产生。将大鼠分为三组（每组n=6）：（G1）生理盐水+LPS D2 L β-D2-精氨酸（3 mM），（G2）LPS+ LPS D2 L β-D2-精氨酸（3 mM），（G3）LPS+ LPS D2 L β-D2-精氨酸（0.1mM）。G2和G3中的动物在肺分离前6小时接受LPS。输注黄嘌呤氧化酶（0.1U/ml）以产生O-甲基-D22-亚氨基-D2-和2-甲基-6-（对甲氧基苯基）-3，7-二氢-亚氨基（1，2-a）吡嗪-3-酮（MCLA; 6 mM），将O-甲基-D22-亚氨基-D2-特异性化学发光探针加入灌注液中。我们通过测量灌流液中乳酸脱氢酶（LDH）的浓度来估计肺损伤。使用Griess反应测定灌流液中的亚硝酸盐和硝酸盐。LPS给药增加了O ...更多信息 D22-D22-生产只有轻微。与G1相比，G2的NO产生和LDH浓度均显著增加，但G3的NO产生和LDH浓度均不显著增加。实验二：一氧化氮（NO）在氧化性肺损伤发病机制中的作用尚不明确。本实验观察了两种一氧化氮合酶（NOS）抑制剂：高选择性诱导型一氧化氮合酶（iNOS）抑制剂NIL D2 L-1-（1-亚氨乙基）赖氨酸（NIL）和非选择性NOS抑制剂NMMA D2 L-1-NG-单甲基精氨酸（NMMA）对大鼠高氧肺损伤的影响。动物在暴露于100%氧气（O NOD 22 NOD 2）或空气之前12小时开始，每12小时腹膜内接受NOS D2 L NOS D2-NIL、NOS D2 L NOS D2-NMMA或稀释剂，持续50小时。以肺组织湿干重比（W/D）作为肺水肿的指标，以支气管肺泡灌洗液（BALP）中总蛋白浓度作为肺蛋白通量的指标。我们还评估了高氧诱导的肺组织诱导型一氧化氮合酶（iNOS）和内皮型一氧化氮合酶（eNOS）mRNA表达的变化。O_（22）O_（2 NIL和NMMA减弱了O-22-D2诱导的亚硝酸盐/硝酸盐浓度的增加，但只有NMMA使W/D和BALP进一步显著增加，高于O-22-D2暴露。暴露于100%O-22-D2可增加iNOS和eNOS mRNA的表达，并且这种作用不被作为竞争性NOS抑制剂的D2-D2-NIL或D2-D2-NMMA改变。总之，内皮细胞NO在高氧肺损伤中起保护作用，而内皮细胞D2-NMMA则无此作用。少