内源性一氧化氮在内毒素引起的肺动脉高压和肺损伤中的作用

本实验观察了家兔静脉内注入内毒素的主要成分脂多糖(LPS)后平均动脉血压(MAP)、肺动脉压(PAP)及入、出肺血NO含量的变化,并观察了静脉内预注入NO生成抑制剂Nω-硝基-L-精氨酸(L-NNA)及诱生型NO生成抑制剂氨基胍(AG)后PAP和肺损伤的变化.结果观察到:家兔LPS注入后,MAP均明显下降,LPS注入后0.5、1、1.5、2h PAP明显增高(P<0.05).LPS注入后PAP的高峰期(1h)入肺血NO含量明显降低,出肺血NO无明显变化.与对照组相比,LPS注入后3h出肺血NO含量和5h入、出肺血NO含量均明显增多.相关分析表明,兔LPS注入前和LPS注入后1h PAP与入肺血NO含量呈明显的负相关,而LPS注入后 3h和5h两者相关不明显.静脉预注入L-NNA后,LPS处理组的动物PAP明显增高,入、出肺血丙二醛(MDA)含量也明显增高,动物生存率明显降低.肺组织光镜下可见肺萎陷和小血管淤血加重,白细胞明显增加.静脉预注入AG后,LPS处理组的动物MAP在3～5h明显增高,此时PAP无明显改变,但5h时血中MDA含量明显减低,5h时与LPS组相比肺萎陷和小血管淤血减轻,白细胞也明显减少.以上结果提示,内毒素入血后较早期阶段可出现PAP的升高,此时入肺血NO的减少是参与肺动脉压增高(PAH)的机制之一.家兔内毒素进入血后较早期阶段NO对减轻内毒素引起的PAH和肺损伤起重要作用,而较晚的时期当诱生型NO合酶(iNOS)诱生后释放的NO则参与内毒素引起的肺组织炎症反应和肺损伤.     

Effect of position, nitric oxide, and almitrine on lung perfusion in a porcine model of acute lung injury.

In a porcine model of oleic acid-induced lung injury, the effects of inhaled nitric oxide (iNO) and intravenous almitrine bismesylate (ivALM), which enhances the hypoxic pulmonary vasoconstriction on the distribution of regional pulmonary blood flow (PBF), were assessed. After injection of 0.12 ml/kg oleic acid, 20 anesthetized and mechanically ventilated piglets [weight of 25 +/- 2.6 (SD) kg] were randomly divided into four groups: supine position, prone position, and 10 ppm iNO for 40 min followed by 4 microg x kg(-1) x min(-1) ivALM for 40 min in supine position and in prone position. PBF was measured with positron emission tomography and H(2)15O. The redistribution of PBF was studied on a pixel-by-pixel basis. Positron emission tomography scans were performed before and then 120, 160, and 200 min after injury. With prone position alone, although PBF remained prevalent in the dorsal regions it was significantly redistributed toward the ventral regions (P < 0.001). A ventral redistribution of PBF was also obtained with iNO regardless of the position (P = 0.043). Adjunction of ivALM had no further effect on PBF redistribution. PP and iNO have an additive effect on ventral redistribution of PBF.     

Birth upregulates nitric oxide synthase activity in the porcine lung

Developmental changes in the lung occur at birth, allowing for the transition from placental to air breathing. Here we have measured nitric oxide synthase (NOS) activity in the porcine lung pre and post partum. NOS activity, which was predominantly calcium dependent, was low in full term fetal tissue compared to that present in lungs from the newborn (5 minutes post partum), 1, 3, 6 and 14 day old animals. No increase in activity was seen when fetal pigs were allowed to breathe for 5 minutes. Specific activity remained low in fetal tissue following partial purification. By contrast, levels of NOS III protein in tissue extracts and in pulmonary arterial endothelial cells, demonstrated by immunohistochemistry, were similar in tissue from the fetal and newborn animals. Thus NOS activity is significantly lower in fetal when compared to postnatal lung tissue despite comparable amounts of NOS III protein being expressed, and birth is followed by an abrupt increase in enzyme activity in animals born at term which correlates with an increase in protein expression.     

PAF antagonists inhibit monocrotaline-induced lung injury and pulmonary hypertension.

Lung platelet-activating factor (PAF) levels increased in some rats at 1-3 wk after subcutaneous injection of monocrotaline (MCT). We tested the effect of specific PAF antagonists, WEB 2086 and WEB 2170, on MCT-induced lung injury and subsequent pulmonary hypertension and right ventricular hypertrophy. Treatment with either agent decreased MCT-induced pulmonary hypertension and right ventricular hypertrophy at 3 wk after injection. Treatment with WEB 2170 reduced MCT-induced pulmonary vascular leak at 1 wk after injection, and WEB 2086-treatment exclusively during the early leak phase also decreased MCT-induced right ventricular hypertrophy at 3 wk. Treatment with WEB 2170 between the 3rd and 4th wk after MCT injection inhibited the progression of right ventricular hypertrophy at 4 wk. These results suggest that PAF contributes to the early pulmonary vascular leak, and this leak phase is important for the development of pulmonary hypertension and right ventricular hypertrophy in MCT-treated rats. Furthermore, it appears that PAF action contributes to the maintenance of a chronic inflammatory process that involves the synthesis of other lipid mediators (prostaglandins and leukotrienes) and leads to pulmonary hypertension. We conclude that PAF has a role in the MCT-induced inflammatory lung injury and pulmonary hypertension.     

Altered function of pulmonary surfactant in fatty acid lung injury

To determine whether acute fatty acid lung injury impairs pulmonary surfactant function, we studied anesthetized ventilated rabbits given oleic acid (55 mg/kg iv, n = 11) or an equivalent volume of saline (n = 8). Measurements of pulmonary mechanics indicated a decrease in dynamic compliance within 5 min of injury and a decrease in lung volume that was disproportionately large at low pressures, consistent with diminished surfactant activity in vivo. Bronchoalveolar lavage fluid obtained 1 h after injury had significantly increased erythrocytes and total leukocytes, largely polymorphonuclear cells. The phospholipid content and composition of the cell-free fraction had only minor changes from those of controls, but the protein content was increased 35-fold. Measurements of lavage surface activity in vitro showed an increase in average minimum surface tension from 1.3 +/- 0.4 (SE) dyn/cm in controls to 20.2 +/- 3.9 dyn/cm in injured animals. The alterations in static pressure-volume curves and decrease in lavage surface activity suggest a severe alteration of surfactant function in this form of lung injury that occurs despite the presence of normal amounts of surfactant phospholipids.     

Pathophysiology of pulmonary hypertension in acute lung injury

Acute lung injury (ALI) and acute respiratory distress syndrome are characterized by protein rich alveolar edema, reduced lung compliance, and acute severe hypoxemia. A degree of pulmonary hypertension (PH) is also characteristic, higher levels of which are associated with increased morbidity and mortality. The increase in right ventricular (RV) afterload causes RV dysfunction and failure in some patients, with associated adverse effects on oxygen delivery. Although the introduction of lung protective ventilation strategies has probably reduced the severity of PH in ALI, a recent invasive hemodynamic analysis suggests that even in the modern era, its presence remains clinically important. We therefore sought to summarize current knowledge of the pathophysiology of PH in ALI.     

Inhaled nitric oxide attenuates pulmonary inflammation and fibrin deposition and prolongs survival in neonatal hyperoxic lung injury

Administration of inhaled nitric oxide (iNO) is a potential therapeutic strategy to prevent bronchopulmonary dysplasia (BPD) in premature newborns with respiratory distress syndrome. We evaluated this approach in a rat model, in which premature pups were exposed to room air, hyperoxia, or a combination of hyperoxia and NO (8.5 and 17 ppm). We investigated the anti-inflammatory effects of prolonged iNO therapy by studying survival, histopathology, fibrin deposition, and differential mRNA expression (real-time RT-PCR) of key genes involved in the development of BPD. iNO therapy prolonged median survival 1.5 days (P = 0.0003), reduced fibrin deposition in a dosage-dependent way up to 4.3-fold (P < 0.001), improved alveolar development by reducing septal thickness, and reduced the influx of leukocytes. Analysis of mRNA expression revealed an iNO-induced downregulation of genes involved in inflammation (IL-6, cytokine-induced neutrophilic chemoattractant-1, and amphiregulin), coagulation, fibrinolysis (plasminogen activator inhibitor 1 and urokinase-type plasminogen activator receptor), cell cycle regulation (p21), and an upregulation of fibroblast growth factor receptor-4 (alveolar formation). We conclude that iNO therapy improves lung pathology and prolongs survival by reducing septum thickness, inhibiting inflammation, and reducing alveolar fibrin deposition in premature rat pups with neonatal hyperoxic lung injury.     

A perinatal nitric oxide donor increases renal vascular resistance and ameliorates hypertension and glomerular injury in adult fawn-hooded hypertensive rats

Enhancing perinatal nitric oxide (NO) availability persistently reduces blood pressure in spontaneously hypertensive rats. We hypothesize that this approach can be generalized to other models of genetic hypertension, for instance those associated with renal injury. Perinatal exposure to the NO donor molsidomine was studied in fawn-hooded hypertensive (FHH) rats, a model of mild hypertension, impaired preglomerular resistance, and progressive renal injury. Perinatal molsidomine increased urinary NO metabolite excretion at 8 wk of age, i.e., 4 wk after treatment was stopped (P < 0.05). Systolic blood pressure was persistently reduced after molsidomine (42-wk females: 118 +/- 3 vs. 141 +/- 5 and 36-wk males: 139 +/- 4 vs. 158 +/- 4 mmHg; both P < 0.001). Perinatal treatment decreased glomerular filtration rate (P < 0.05) and renal blood flow (P < 0.01) and increased renal vascular resistance (P < 0.05), without affecting filtration fraction, suggesting persistently increased preglomerular resistance. At 4 wk of age natriuresis was transiently increased by molsidomine (P < 0.05). Molsidomine decreased glomerulosclerosis (P < 0.05). Renal blood flow correlated positively with glomerulosclerosis in control (P < 0.001) but not in perinatally treated FHH rats. NO dependency of renal vascular resistance was increased by perinatal molsidomine. Perinatal enhancement of NO availability can ameliorate development of hypertension and renal injury in FHH rats. Paradoxically, glomerular protection by perinatal exposure to the NO donor molsidomine may be due to persistently increased preglomerular resistance. The mechanisms by which increased perinatal NO availability can persistently reprogram kidney function and ameliorate hypertension deserve further study.     

Combined inhaled nitric oxide and inhaled prostacyclin during experimental chronic pulmonary hypertension.

Inhaled nitric oxide (NO) and inhaled prostacyclin (PGI2) produce selective reductions in pulmonary vascular resistance (PVR) through differing mechanisms. NO decreases PVR via cGMP, and PGI2 produces pulmonary vasodilation via cAMP. As a general pharmacological principle, two drugs that produce similar effects via different mechanisms should have additive or synergistic effects when combined. We designed this study to investigate whether combined inhaled NO and PGI2 therapy results in additive effects during chronic pulmonary hypertension in the rat. Monocrotaline injected 4 wk before study produced pulmonary hypertension in all animals. Inhaled NO (20 parts/million) reversibly and selectively decreased pulmonary artery pressure (Ppa) with a mean reduction of 18%. Four concentrations of PGI2 were administered via inhalation (5, 10, 20, and 80 microg/ml), both alone and combined with inhaled NO. Inhaled PGI2 alone decreased Ppa in a dose-dependent manner with no change in mean systemic arterial pressure. Combined inhaled NO and PGI2 selectively and significantly decreased Ppa more did than either drug alone. The effects were additive at the lower concentrations of PGI2 (5, 10, and 20 microg/ml). The combination of inhaled NO and inhaled PGI2 may be useful in the management of pulmonary hypertension.     

Effect of recombinant SP-C surfactant in a porcine lavage model of acute lung injury.

Synthetic surfactants allow examination of the effects of specific components of natural surfactant. To determine whether surfactant containing apoprotein C, dipalmitoyl-phosphatidylcholine, phosphatidylglycerol, and palmitic acid restores gas-exchanging function in acute lung injury (ALI), we administered such surfactant (in doses of 50 or 100 mg/kg and in volumes from 1 to 6 ml/kg) or phospholipid (PL) alone, by intratracheal instillation, to pigs with ALI induced by massive saline lavage. Animals ventilated with 100% O(2) and receiving 1, 2, 4, or 6 ml/kg of 50 mg/kg recombinant surfactant apoprotein C (rSP-C) surfactant or 2 ml/kg of 50 mg/kg PL (control) had mean arterial PO(2) values, 4 h after treatment, of 230, 332, 130, 142, or 86 Torr, respectively. Animals receiving 1, 2, or 4 ml/kg of 100 mg/kg rSP-C surfactant or 2 ml/kg of 100 mg/kg PL (control) had mean arterial PO(2) values of 197, 214, 148, or 88 Torr, respectively. Surfactant PL distribution was homogeneous. Hyaline membrane formation was reduced in treated animals. Thus, in this model of ALI, rSP-C with PL has the capacity to improve gas exchange and possibly modify lung injury.     

Role of inhibition of nitric oxide production in monocrotaline-induced pulmonary hypertension

Mathew, Rajamma, Elizabeth S. Gloster, T. Sundararajan, Carl I. Thompson, Guillermo A. Zeballos, andMichael H. Gewitz. Role of inhibition of nitric oxide productionin monocrotaline-induced pulmonary hypertension. J. Appl. Physiol. 82(5): 1493-1498, 1997.Monocrotaline (MCT)-induced pulmonary hypertension (PH) isassociated with impaired endothelium-dependent nitric oxide(NO)-mediated relaxation. To examine the role of NO in PH,Sprague-Dawley rats were given a single subcutaneous injection ofnormal saline [control (C)], 80 mg/kg MCT, or the same doseof MCT and a continuous subcutaneous infusion of 2 mg · kg¹ · day¹of molsidomine, a NO prodrug (MCT+MD). Two weeks later, plasma NO₃ levels, pulmonary arterialpressure (Ppa), ratio of right-to-left ventricular weights (RV/LV) toassess right ventricular hypertrophy, and pulmonary histology wereevaluated. The plasma NO₃ level inthe MCT group was reduced to 9.2 ± 1.5 µM(n = 12) vs. C level of 17.7 ± 1.8 µM (n = 8; P < 0.02). In the MCT+MD group,plasma NO₃ level was 12.3 ± 2.0 µM (n = 8). Ppa and RV/LV in theMCT group were increased compared with C [Ppa, 34 ± 3.4 mmHg(n = 6) vs. 19 ± 0.8 mmHg(n = 8) and 0.41 ± 0.01 (n = 9) vs. 0.25 ± 0.008 (n = 8), respectively;P < 0.001]. In the MCT+MDgroup, Ppa and RV/LV were not different when compared with C [19 ± 0.5 mmHg (n = 5) and 0.27 ± 0.01 (n = 9), respectively;P < 0.001 vs. MCT]. Medial wall thickness of lung vessels in the MCT group was increased comparedwith C [31 ± 1.5% (n = 9)vs. 13 ± 0.66% (n = 9);P < 0.001], and MDpartially prevented MCT-induced pulmonary vascular remodeling [22 ± 1.2% (n = 11);P < 0.001 vs. MCT and C].These results indicate that a defect in the availability of bioactive NO may play an important role in the pathogenesis of MCT-induced PH.

     

Modeling pulmonary nitric oxide exchange.

Nitric oxide (NO) was first detected in the exhaled breath more than a decade ago and has since been investigated as a noninvasive means of assessing lung inflammation. Exhaled NO arises from the airway and alveolar compartments, and new analytical methods have been developed to characterize these sources. A simple two-compartment model can adequately represent many of the observed experimental observations of exhaled concentration, including the marked dependence on exhalation flow rate. The model characterizes NO exchange by using three flow-independent exchange parameters. Two of the parameters describe the airway compartment (airway NO diffusing capacity and either the maximum airway wall NO flux or the airway wall NO concentration), and the third parameter describes the alveolar region (steady-state alveolar NO concentration). A potential advantage of the two-compartment model is the ability to partition exhaled NO into an airway and alveolar source and thus improve the specificity of detecting altered NO exchange dynamics that differentially impact these regions of the lungs. Several analytical techniques have been developed to estimate the flow-independent parameters in both health and disease. Future studies will focus on improving our fundamental understanding of NO exchange dynamics, the analytical techniques used to characterize NO exchange dynamics, as well as the physiological interpretation and the clinical relevance of the flow-independent parameters.     

Detection of porcine oleic acid-induced acute lung injury using pulmonary acoustics.

To evaluate the utility of monitoring the sound-filtering characteristics of the respiratory system in the assessment of acute lung injury (ALI), we injected a multifrequency broadband sound signal into the airway of five anesthetized, intubated pigs, while recording transmitted sound over the trachea and on the chest wall. Oleic acid injections effected a severe lung injury predominantly in the dependent lung regions, increasing venous admixture from 6 +/- 1 to 54 +/- 8% (P < 0.05) and reducing dynamic respiratory system compliance from 19 +/- 0 to 12 +/- 2 ml/cmH(2)O (P < 0.05). A two- to fivefold increase in sound transfer function amplitude was seen in the dependent (P < 0.05) and lateral (P < 0.05) lung regions; no change occurred in the nondependent areas. High within-subject correlations were found between the changes in dependent lung sound transmission and venous admixture (r = 0.82 +/- 0.07; range 0.74-0.90) and dynamic compliance (r = -0.87 +/- 0.05; -0.80 to -0.93). Our results indicate that the acoustic changes associated with oleic acid-induced lung injury allow monitoring of its severity and distribution.     

一氧化氮供体对过氧化氢引起的心肌细胞损伤的保护作用

关于一氧化氮(NO)对心肌细胞是否具有保护作用目前尚存在争议,为探讨NO对过氧化氢(H2O2)引起的心肌细胞损伤是否具有保护作用及其可能的机制,实验将体外培养的新生大鼠心肌细胞分为3组(1)阴性对照组(Normal组);(2)H2O2组H2O2(0.1mmol/L)与心肌细胞共育4h;(3)S-亚硝基-N-乙酰青霉胺(SNAP)+H2O2组NO供体SNAP(0.5mmol/L)处理心肌细胞10min后,加入H2O2与心肌细胞共育4 h.用流式细胞术检测心肌细胞凋亡率,心肌细胞损伤程度以心肌细胞存活率和乳酸脱氢酶(lactate dehydrogenase,LDH)活性来表示,同时检测心肌细胞超氧化物歧化酶(superoxide dismutase,SOD)活性和丙二醛(MDA)含量.通过激光共聚焦显微术检测在不同处理条件下心肌细胞胞内钙的变化.结果表明,正常心肌细胞LDH活性和细胞存活率分别为631.4±75.6 U/L和93.1±6.2%,细胞凋亡率为0;H2O2处理细胞后可使细胞LDH活性显著增高(1580.5±186.7 U/L,P＜0.01),细胞存活率明显下降(58.3±7.6%,P＜0.01),流式细胞仪检测到大量心肌细胞凋亡,凋亡率为26.4±5.7%;SOD活性较正常细胞19.67±0.85 NU/ml显著下降,为14.73±1.68 NU/m(P＜0.01),MDA含量较正常细胞6.95±0.83μmol/L显著增高,为15.35±3.49μmol/L(P＜0.01).SNAP预处理细胞可显著提高心肌细胞存活率(79.7±9.3%,P＜0.01),降低LDH活性和细胞凋亡率(分别为957.8±110.9 U/L和9.1±3.3%,P＜0.01);并提高细胞抗氧化能力,表现为较H2O2处理组的SOD活性增高(21.36±3.11 NU/ml,P＜0.01),MDA含量下降(9.12±1.47 μmol/L,P＜0.01).激光共聚焦显微镜检测结果表明,H2O2可升高细胞内钙,而SNAP则可降低细胞内钙,SNAP预处理细胞后可取消H2O2升高细胞内钙的作用.上述结果提示,NO供体SNAP可对抗H2O2对心肌细胞的损伤,其机制与提高心肌细胞抗氧化损伤能力和对抗H2O2引起的细胞内钙超载有关.     

Sex differences in renal injury and nitric oxide production in renal wrap hypertension

To investigate the faster rate of renal disease progression in men compared with women, we addressed the following questions in the renal wrap (RW) model of hypertension: 1) Do sex differences exist in RW-induced renal injury, which are independent of sex differences in blood pressure? 2) Do sex differences in nitric oxide (NO) production exist in RW hypertension? Male (M) and female (F) rats underwent sham-operated (M-Sham, n = 7; F-Sham, n = 10) or RW (M-RW, n = 13; F-RW, n = 14) surgery for 9 wk. Markers of renal injury, including the glomerulosclerosis index (F-RW, 0.70 +/- 0.1 vs. M-RW, 2.2 +/- 0.6; P < 0.05), mean glomerular volume (F-RW, 1.05 +/- 0.050 x 10(6) vs. M-RW, 1.78 +/- 0.15 x 10(6) microm(3); P < 0.001), and proteinuria (F-RW, 68.7 +/- 15 vs. M-RW, 124 +/- 7.7 mg/day; P < 0.001) were greater in RW males compared with RW females. Endothelial NO synthase protein expression was elevated in the renal cortex (3.2-fold) and medulla (2.2-fold) 9 wk after RW in males, whereas no differences were observed in females. Neuronal NO synthase protein expression was unchanged in the renal cortex in males and in both the renal cortex and medulla in females, whereas in the male medulla, neuronal NOS was decreased by 57%. These data suggest the degree of renal injury is greater in male compared with female rats in RW hypertension despite similar degrees of hypertension and renal function and may involve sex differences in renal NO metabolism.