Estradiol improves pulmonary hemodynamics and vascular remodeling in perinatal pulmonary hypertension

Partial ligation of the ductus arteriosus (DA) in the fetal lamb causes sustained elevation of pulmonary vascular resistance (PVR) and hypertensive structural changes in small pulmonary arteries, providing an animal model for persistent pulmonary hypertension of the newborn. Based on its vasodilator and antimitogenic properties in other experimental studies, we hypothesized that estradiol (E(2)) would attenuate the pulmonary vascular structural and hemodynamic changes caused by pulmonary hypertension in utero. To test our hypothesis, we treated chronically instrumented fetal lambs (128 days, term = 147 days) with daily infusions of E(2) (10 microg; E(2) group, n = 6) or saline (control group, n = 5) after partial ligation of the DA. We measured intrauterine pulmonary and systemic artery pressures in both groups throughout the study period. After 8 days, we delivered the study animals by cesarean section to measure their hemodynamic responses to birth-related stimuli. Although pulmonary and systemic arterial pressures were not different in utero, fetal PVR immediately before ventilation was reduced in the E(2)-treated group (2.43 +/- 0.79 vs. 1.48 +/- 0.26 mmHg. ml(-1). min, control vs. E(2), P < 0.05). During the subsequent delivery study, PVR was lower in the E(2)-treated group in response to ventilation with hypoxic gas but was not different between groups with ventilation with 100% O(2). During mechanical ventilation after delivery, arterial partial O(2) pressure was higher in E(2) animals than controls (41 +/- 11 vs. 80 +/- 35 Torr, control vs. E(2), P < 0. 05). Morphometric studies of hypertensive vascular changes revealed that E(2) treatment decreased wall thickness of small pulmonary arteries (59 +/- 1 vs. 48 +/- 1%, control vs. E(2), P < 0.01). We conclude that chronic E(2) treatment in utero attenuates the pulmonary hemodynamic and histological changes caused by DA ligation in fetal lambs.     

Pulmonary hypertension impairs alveolarization and reduces lung growth in the ovine fetus

Persistent pulmonary hypertension of the newborn (PPHN) is a clinical disorder characterized by abnormal vascular structure, growth, and reactivity. Disruption of vascular growth during early postnatal lung development impairs alveolarization, and newborns with lung hypoplasia often have severe pulmonary hypertension. To determine whether pulmonary hypertension can directly impair vascular growth and alveolarization in the fetus, we studied the effects of chronic intrauterine pulmonary hypertension on lung growth in fetal lambs. We performed surgery, which included partial constriction of the ductus arteriosus (DA) to induce pulmonary hypertension (PH, n = 14) or sham surgery (controls, n = 13) in fetal lambs at 112-125 days (term = 147 days). Tissues were harvested near term for measurement of right ventricular hypertrophy (RVH), radial alveolar counts (RAC), mean linear intercepts (MLI), wall thickness, and vessel density of small pulmonary arteries. Chronic DA constriction caused RVH (P < 0.0001), increased wall thickness of small pulmonary arteries (P < 0.002), and reduced small pulmonary artery density (P < 0.005). PH also reduced alveolarization, causing a 27% reduction in RAC and 20% increase in MLI. Furthermore, prolonged DA constriction (21 days) not only decreased RAC and increased MLI by 30% but also caused a 25% reduction of lung-body weight ratio. We conclude that chronic PH reduces pulmonary arterial growth, decreases alveolar complexity, and impairs lung growth. We speculate that chronic hypertension impairs vascular growth, which disrupts critical signaling pathways regulating lung vascular and alveolar development, thereby interfering with alveolarization and ultimately resulting in lung hypoplasia.     

Intrauterine hypertension decreases lung VEGF expression and VEGF inhibition causes pulmonary hypertension in the ovine fetus

Although vascular endothelial growth factor (VEGF) plays a vital role in lung vascular growth in the embryo, its role in maintaining endothelial function and modulating vascular structure during late fetal life has not been studied. We hypothesized that impaired lung VEGF signaling causes pulmonary hypertension, endothelial dysfunction, and structural remodeling before birth. To determine whether lung VEGF expression is decreased in an experimental model of persistent pulmonary hypertension of the newborn (PPHN), we measured lung VEGF and VEGF receptor protein content from fetal lambs 7-10 days after ductus arteriosus ligation (132-140 days gestation; term = 147 days). In contrast with the surge in lung VEGF expression during late gestation in controls, chronic intrauterine pulmonary hypertension reduced lung VEGF expression by 78%. To determine whether VEGF inhibition during late gestation causes pulmonary hypertension, we treated fetal lambs with EYE001, an aptamer that specifically inhibits VEGF(165). Compared with vehicle controls, EYE001 treatment elevated pulmonary artery pressure and pulmonary vascular resistance by 22 and 50%, respectively, caused right ventricular hypertrophy, and increased wall thickness of small pulmonary arteries. EYE001 treatment reduced lung endothelial nitric oxide synthase protein content by 50% and preferentially impaired the pulmonary vasodilator response to ACh, an endothelium-dependent agent. We conclude that chronic intrauterine pulmonary hypertension markedly decreases lung VEGF expression and that selective inhibition of VEGF(165) mimics the structural and physiological changes of experimental PPHN. We speculate that hypertension downregulates VEGF expression in the developing lung and that impaired VEGF signaling may contribute to the pathogenesis of PPHN.     

Differential proinflammatory and prooxidant effects of bone morphogenetic protein-4 in coronary and pulmonary arterial endothelial cells

There is increasing evidence that TGF-beta family member cytokine bone morphogenetic protein (BMP)-4 plays different pathophysiological roles in the pulmonary and systemic circulation. Upregulation of BMP-4 has been linked to atherosclerosis and hypertension in the systemic circulation, whereas disruption of BMP-4 signaling is associated with the development of pulmonary hypertension. To test the hypothesis that BMP-4 elicits differential effects in the pulmonary and systemic circulation, we compared the prooxidant and proinflammatory effects of BMP-4 in cultured human coronary arterial endothelial cells (CAECs) and pulmonary arterial endothelial cells (PAECs). We found that BMP-4 (from 0.3 to 10 ng/ml) in CAECs increased O(2)(*-) and H(2)O(2) generation, induced NF-kappaB activation, upregulated ICAM-1, and induced monocyte adhesiveness to ECs. In contrast, BMP-4 failed to induce oxidative stress or endothelial activation in PAECs. Also, BMP-4 treatment impaired acetylcholine-induced relaxation and increased O(2)(*-) production in cultured rat carotid arteries, whereas cultured rat pulmonary arteries were protected from these adverse effects of BMP-4. Thus, we propose that BMP-4 exerts prooxidant, prohypertensive, and proinflammatory effects only in the systemic circulation, whereas pulmonary arteries are protected from these adverse effects of BMP-4. The vascular bed-specific endothelial effects of BMP-4 are likely to contribute to its differential pathophysiological role in the systemic and pulmonary circulation.     

Umbilical cord compression produces pulmonary hypertension in newborn lambs: a model to study the pathophysiology of persistent pulmonary hypertension in the newborn

We investigated the effects of chronic intrauterine hypoxaemia produced by prolonged partial umbilical cord compression on the circulation shortly after birth in lambs. Vascular catheters were inserted in 10 fetal sheep at 120 to 130 days gestation to measure descending aortic blood gases, arterial pH, and arterial O2 saturation. An inflatable silicone rubber balloon cuff was also placed around the umbilical cord. After recovery and the return of descending aortic blood gases to the normal range, the balloon was gradually inflated, decreasing the PaO2 from 21.2 +/- 3.6 to 17.5 +/- 1.3 mm Hg and the arterial O2 saturation from 57.1 +/- 9.2% to 37.2% +/- 5.2. After 14.3 +/- 3.7 days of partial umbilical cord compression, the lambs were delivered by Caesarean section, instrumented to measure systemic and pulmonary arterial, right atrial and pulmonary arterial wedge pressures, pulmonary and systemic blood flows, and mechanically ventilated. Five normal lambs were also studied. From 60 to 120 min after delivery, when compared to normal lambs, the umbilical compression lambs had an increased pulmonary arterial pressure (P less than 0.05) pulmonary vascular resistance (P less than 0.05), and right atrial pressure (P less than 0.05) with similar arterial blood gases. In both groups, hypoxic ventilation produced an increase in pulmonary arterial pressure (P less than 0.05) which on return to room air ventilation decreased to baseline in the normal lambs but not in the umbilical cord compression lambs (P less than 0.05). Prolonged partial umbilical cord compression produces chronic fetal hypoxaemia and pulmonary arterial hypertension after birth. This may represent a model to study the pathophysiology of persistent pulmonary hypertension syndrome.     

Pulmonary vascular effects of sildenafil on the development of chronic pulmonary hypertension in the ovine fetus

We investigated the pulmonary vascular effects of prophylactic use of sildenafil, a specific phosphodiesterase-5 inhibitor, in late-gestation fetal lambs with chronic pulmonary hypertension. Fetal lambs were operated on at 129 +/- 1 days gestation (term = 147 days). Ductus arteriosus (DA) was compressed for 8 days to cause chronic pulmonary hypertension. Fetuses were treated with sildenafil (24 mg/day) or saline. Pulmonary vascular responses to increase in shear stress and in fetal PaO2 were studied at, respectively, day 4 and 6. Percent wall thickness of small pulmonary arteries (%WT) and the right ventricle-to-left ventricle plus septum ratio (RVH) were measured after completion of the study. In the control group, DA compression increased PA pressure (48 +/- 5 to 72 +/- 8 mmHg, P < 0.01) and pulmonary vascular resistance (PVR) (0.62 +/- 0.08 to 1.15 +/- 0.11 mmHg x ml(-1) x min(-1), P < 0.05). Similar increase in PAP was observed in the sildenafil group, but PVR did not change significantly (0.54 +/- 0.06 to 0.64 +/- 0.09 mmHg x ml(-1) x min(-1)). Acute DA compression, after brief decompression, elevated PVR 25% in controls and decreased PVR 35% in the sildenafil group. Increased fetal PaO2 did not change PVR in controls but decreased PVR 60% in the sildenafil group. %WT and RVH were not different between groups. Prophylactic sildenafil treatment prevents the rise in pulmonary vascular tone and altered vasoreactivity caused by DA compression in fetal lambs. These results support the hypothesis that elevated PDE5 activity is involved in the consequences of chronic pulmonary hypertension in the perinatal lung.     

Postnatal changes in pulmonary vein responses to endothelin-1 in the normal and chronically hypoxic lung

The response of pulmonary arteries to endothelin-1 (ET-1) changes with age in normal pigs and is abnormal in pulmonary hypertension. The purpose of this study was to determine if the same is true of the pulmonary veins. We studied the wall structure and functional response to ET-1 in pulmonary veins from normal pigs from fetal life to adulthood and from pigs subjected to chronic hypobaric hypoxia either from birth for 3 days or from 3 to 6 days of age. In isolated normal veins, the contractile response decreased by 40% between late fetal life and 14 days of age with a concomitant twofold increase in endothelium-dependent relaxant response. The ET(A) antagonist BQ-123 reduced the contractile response significantly more in newborn than older animals, whereas the ET-B antagonist BQ-788 had no effect in fetal animals and maximally increased contraction at 14 days of age. Hypoxic exposure significantly increased pulmonary vein smooth muscle area and contractile response to ET-1. The relaxation response was impaired following hypoxic exposure from birth but not from 3 to 6 days of age. The ET(A) antagonist BQ-123 decreased contractile and increased dilator responses significantly more than in age-matched controls. Thus pulmonary veins show age-related changes similar to those seen in the pulmonary arteries with a decrease in ET(A)-mediated contractile and increase in ET-B-mediated relaxant response with age. Contractile response was also increased in hypoxia as in the arteries. This study suggests that pulmonary veins are involved in postnatal adaptation and the pathogenesis of pulmonary hypertension.     

Telokin expression and the effect of hypoxia on its phosphorylation status in smooth muscle cells from small and large pulmonary arteries

Small pulmonary arteries (SPA), <500 microm diameter of the cat, constrict when exposed to hypoxia, whereas larger arteries (large pulmonary arteries; LPA), >800 microm diameter, show little or no response. It is unknown why different contractile responses occur within the same vascular bed, but activator or repressor proteins within the smooth muscle cell (SMC) can modify myosin phosphatase and myosin light chain kinase (MLCK), thereby influencing the phosphorylation state of myosin light chain (MLC) and ultimately, contraction. Telokin, a protein with a sequence identical to the COOH-terminal domain of MLCK, is expressed in smooth muscle where in its phosphorylated state it inhibits myosin phosphatase, binds to unphosphorylated myosin, and helps maintain smooth muscle relaxation. We measured telokin mRNA and telokin protein in smooth muscle from different diameter feline pulmonary arteries and sought to determine whether changes in the phosphorylation status of telokin and MLC occurred during hypoxia. In pulmonary arteries, telokin expression varied inversely with artery diameter, but cerebral arteries showed neither telokin protein nor telokin mRNA. Although telokin and MLC were distributed uniformly throughout the SPA muscle cell cytoplasm, they were not colocalized. During hypoxia, telokin dephosphorylated, and MLC became increasingly phosphorylated in SPA SMC, whereas in LPA SMC there was no change in either telokin or MLC phosphorylation. When LPA SMC were exposed to phenylephrine, MLC phosphorylation increased with no change in telokin phosphorylation. These results suggest that in SPA, phosphorylated telokin may help maintain relaxation under unstimulated conditions, whereas in LPA, telokin's function remains undetermined.     

Role of platelet-activating factor in pulmonary vascular remodeling associated with chronic high altitude hypoxia in ovine fetal lambs

Platelet-activating factor (PAF) is implicated in pathogenesis of chronic hypoxia-induced pulmonary hypertension in some animal models and in neonates. Effects of chronic hypoxia on PAF receptor (PAF-R) system in fetal pulmonary vasculature are unknown. We investigated the effect of chronic high altitude hypoxia (HAH) in fetal lambs [pregnant ewes were kept at 3,801 m (12,470 ft) altitude from approximately 35 to 145 days gestation] on PAF-R-mediated effects in the pulmonary vasculature. Age-matched controls were kept at sea level. Intrapulmonary arteries were isolated, and smooth muscle cells (SMC-PA) were cultured from HAH and control fetuses. To determine presence of pulmonary vascular remodeling, lung tissue sections were subjected to morphometric analysis. Percentage medial wall thickness was significantly increased (P < 0.05) in arteries at all levels in the HAH lambs. PAF-R protein expression studied by immunocytochemistry and Western blot analysis on lung tissue SMC-PA demonstrated greater PAF-R expression in HAH lambs. PAF-R binding (femtomoles per 10(6) cells) in HAH SMC-PA was 90.3 +/- 4.08 and 66% greater than 54.3 +/- 4.9 in control SMC-PA. Pulmonary arteries from HAH fetuses synthesized >3-fold PAF than vessels from controls. Compared with controls SMC-PA of HAH lambs demonstrated 139% and 40% greater proliferation in 10% FBS alone and with 10 nM PAF, respectively. Our data demonstrate that exposure of ovine fetuses to HAH will result in significant upregulation of PAF synthesis, PAF-R expression, and PAF-R-mediated effects in pulmonary arteries. These findings suggest that increased PAF-R protein expression and increased PAF binding contribute to pulmonary vascular remodeling in these animals and may predispose them to persistent pulmonary hypertension after birth.     

ET(A)-receptor blockade and ET(B)-receptor stimulation in experimental congenital diaphragmatic hernia

The aim of this study was to assess the role of nitric oxide (NO) and endothelin (ET)-1 in the pathophysiology of persistent pulmonary hypertension of the newborn in fetal lambs with a surgically created congenital diaphragmatic hernia (CDH). The pulmonary vascular response to various agonists and antagonists was assessed in vivo between 128 and 132 days gestation. Age-matched fetal lambs served as control animals. Control and CDH lambs had similar pulmonary vasodilator responses to acetylcholine, sodium nitroprusside, zaprinast, and dipyridamole. The ET(A)-receptor antagonist BQ-123 caused a significantly greater pulmonary vasodilatation in CDH than in control animals. The ET(B)-receptor agonist sarafotoxin 6c induced a biphasic response, with a sustained pulmonary vasoconstriction after a transient pulmonary vasodilatation that was not seen in CDH animals. We conclude that the NO signaling pathway in vivo is intact in experimental CDH. In contrast, ET(A)-receptor blockade and ET(B)-receptor stimulation significantly differed in CDH animals compared with control animals. Imbalance of ET-1-receptor activation favoring pulmonary vasoconstriction rather than altered NO-mediated pulmonary vasodilatation is likely to account for persistent pulmonary hypertension of the newborn in fetal lambs with a surgically created CDH.     

Pulmonary vasodilation with structurally altered pulmonary vessels and pulmonary hypertension

To evaluate pulmonary vasodilation in a structurally altered pulmonary vascular bed, we gave endothelium-dependent (acetylcholine) and endothelium-independent [sodium nitroprusside, prostaglandin I2 (PGI2)] vasodilators in vivo and to isolated lobar pulmonary arteries from neonatal calves with severe pulmonary hypertension. Acetylcholine, administered by pulmonary artery infusion, decreased pulmonary arterial pressure from 120 +/- 7 to 71 +/- 6 mmHg and total pulmonary resistance from 29.4 +/- 2.6 to 10.4 +/- 0.9 mmHg.l-1.min without changing systemic arterial pressure (90 +/- 5 mmHg). Although both sodium nitroprusside and PGI2 lowered pulmonary arterial pressure to 86 +/- 4 and 96 +/- 4 mmHg, respectively, they also decreased systemic arterial pressure to 65 +/- 4 and 74 +/- 3 mmHg, respectively. Neither sodium nitroprusside nor PGI2 was as effective as acetylcholine at lowering total pulmonary resistance (18.0 +/- 3.6 and 19.1 +/- 2.2 mmHg.l-1.min, respectively). Right-to-left cardiac shunt through the foramen ovale was decreased by acetylcholine from 1.6 +/- 0.4 to 0.1 +/- 0.2 l/min but was not changed by sodium nitroprusside or PGI2. Isolated lobar pulmonary arteries from pulmonary hypertensive calves did not relax in response to acetylcholine, whereas isolated pulmonary arteries from age-matched control calves did relax in response to acetylcholine. Control and pulmonary hypertensive lobar pulmonary arteries relaxed equally well in response to sodium nitroprusside. We concluded that acetylcholine vasodilation was impaired in vitro in isolated lobar pulmonary arteries but was enhanced in vivo in resistance pulmonary arteries in neonatal calves with pulmonary hypertension.     

Quantitative characterization of postnatal growth trends in proximal pulmonary arteries in rats by phase-contrast magnetic resonance imaging

Malformations of the pulmonary arteries can increase right heart workload and result in morbidity, heart failure, and death. With the increased use of murine models to study these malformations, there is a pressing need for an accurate and noninvasive experimental technique that is capable of characterizing pulmonary arterial hemodynamics in these animals. We describe the growth trends of pulmonary arteries in 13 male Sprague-Dawley rats at 20, 36, 52, 100, and 160 days of age with the introduction of phase-contrast MRI as such a technique. PCMRI results correlated closely with cardiac output measurements by ultrasound echocardiography and with fluorescent microspheres in right-left lung flow split (flow partition). Mean flow, average cross-sectional area, distensibility, and shear rates for the right and left pulmonary arteries (RPA and LPA) were calculated. The RPA was larger and received more flow at all times than the LPA (P < 0.0001). Right-left flow split did not change significantly with age, and arterial distensibility was not significantly different between RPA and LPA, except at 160 days (P < 0.01). Shear rates were much higher for the LPA than the RPA (P < 0.0001) throughout development. The RPA and LPA showed different structure-function relationships but obeyed similar allometric scaling laws, with scaling exponents comparable to those of the main pulmonary artery. This study is the first to quantitatively describe changes in RPA and LPA flows and sizes with development and to apply phase-contrast MRI techniques to pulmonary arteries in rats.     

Chronic O2 exposure in the newborn rat results in decreased pulmonary arterial nitric oxide release and altered smooth muscle response to isoprostane.

Chronic oxygen exposure in the newborn rat results in lung isoprostane formation, which may contribute to the pulmonary hypertension evident in this animal model. The purpose of this study was to investigate the pulmonary arterial smooth muscle responses to 8-iso-prostaglandin F(2alpha) (8-iso-PGF(2a)) in newborn rats exposed to 60% O2 for 14 days. Because, in the adult rat, 8-iso-PGF(2alpha) may have a relaxant effect, mediated by nitric oxide (NO), we also sought to evaluate the pulmonary arterial NO synthase (NOS) protein content and NO release in the newborn exposed to chronic hyperoxia. Compared with air-exposed control animals, 8-iso-PGF(2a) induced a significantly greater force (P < 0.01) and reduced (P < 0.01) relaxation of precontracted pulmonary arteries in the 60% O2-treated animals. These changes were reproduced in control pulmonary arteries by NOS blockade by using NG-nitro-L-arginine methyl ester. Pulmonary arterial endothelial NOS was unaltered, but the inducible NOS protein content was significantly decreased (P < 0.01) in the experimental group. Pulmonary (P < 0.05) and aortic (P < 0.01) tissue ex vivo NO accumulation was significantly reduced in the 60% O2-treated animals. We speculate that impaired pulmonary vascular tissue NO metabolism after chronic O2 exposure potentiates 8-iso-PGF(2alpha)-induced vasoconstriction in the newborn rat, thus contributing to pulmonary hypertension.     

Hemodynamic effects of periodic G(z) acceleration in meconium aspiration in pigs.

The hemodynamic effects of periodic acceleration (pG(z)), induced in the spinal axis with noninvasive motion ventilation (NIMV), were studied in a piglet model of pulmonary hypertension associated with meconium aspiration. Animals (n = 12) were anesthetized, paralyzed, intubated, and supported by conventional mechanical ventilation (CMV). Thirty minutes after tracheal instillation of meconium solution (6 ml/kg), either CMV (n = 6) was continued or NIMV (n = 6) was initiated. Changes in systemic and pulmonary hemodynamics and arterial blood gases were tracked for 2 h after aspiration. Thermodilution, cardiac output, and heart rate were not significantly different after meconium aspiration in the pG(z) group relative to the CMV controls. Aortic pressure and systemic vascular resistance were significantly lower (approximately 30%) after meconium aspiration in NIMV animals relative to CMV animals. Pulmonary arterial pressure and pulmonary vascular resistance were also significantly lower, by 100%, after aspiration of meconium in the NIMV animals compared with the CMV controls. Meconium aspiration significantly decreased total respiratory compliance by approximately 50% and increased total respiratory resistance by approximately 100% in both CMV and NIMV animals, but such alterations did not differ between the two groups. Both CMV and NIMV satisfactorily supported ventilation in these paralyzed animals. In conclusion, NIMV through pG(z) in the spinal axis decreased systemic and pulmonary vascular resistance in piglets after meconium aspiration.     

Effects of BAY 41-2272, a soluble guanylate cyclase activator, on pulmonary vascular reactivity in the ovine fetus

Nitric oxide (NO)-cGMP signaling plays a critical role during the transition of the pulmonary circulation at birth. BAY 41-2272 is a novel NO-independent direct stimulator of soluble guanylate cyclase that causes vasodilation in systemic and local circulations. However, the hemodynamic effects of BAY 41-2272 have not been studied in the perinatal pulmonary circulation. We hypothesized that BAY 41-2272 causes potent and sustained fetal pulmonary vasodilation. We performed surgery on 14 fetal lambs (125-130 days gestation; term = 147 days) and placed catheters in the main pulmonary artery, aorta, and left atrium to measure pressures. An ultrasonic flow transducer was placed on the left pulmonary artery (LPA) to measure blood flow, and a catheter was placed in the LPA for drug infusion. Pulmonary vascular resistance (PVR) was calculated as pulmonary artery pressure minus left atrial pressure divided by LPA blood flow. BAY 41-2272 caused dose-related increases in pulmonary blood flow up to threefold above baseline and reduced PVR by 75% (P < 0.01). Prolonged infusion of BAY 41-2272 caused sustained pulmonary vasodilation throughout the 120-min infusion period. The pulmonary vasodilator effect of BAY 41-2272 was not attenuated by N(omega)-nitro-l-arginine, a NO synthase inhibitor. In addition, compared with sildenafil, a phosphodiesterase 5 inhibitor, the pulmonary vasodilator response to BAY 41-2272 was more prolonged. We conclude that BAY 41-2272 causes potent and sustained fetal pulmonary vasodilation independent of NO release. We speculate that BAY 41-2272 may have therapeutic potential for pulmonary hypertension associated with failure to circulatory adaptation at birth, especially in the setting of impaired NO production.     

Chronic intrauterine pulmonary hypertension decreases calcium-sensitive potassium channel mRNA expression

Calcium-sensitive potassium (K(Ca)) channels play a critical role in mediating perinatal pulmonary vasodilation. Because infants with persistent pulmonary hypertension of the newborn (PPHN) have blunted vasodilator responses to birth-related stimuli, we hypothesized that lung K(Ca) channel gene expression is decreased in PPHN. To test this hypothesis, we measured K(Ca) channel gene expression in distal lung homogenates from both fetal lambs with severe pulmonary hypertension caused by prolonged compression of the ductus arteriosus and age-matched, sham-operated animals (controls). After at least 9 days of compression of the ductus arteriosus, fetal lambs were killed. To determine lung K(Ca) channel mRNA levels, primers were designed against the known sequence of the K(Ca) channel and used in semiquantitative RT-PCR, with lung 18S rRNA content as an internal control. Compared to that in control lambs, lung K(Ca) channel mRNA content in the PPHN group was reduced by 26 +/- 6% (P < 0.02), whereas lung voltage-gated K(+) 2.1 mRNA content was unchanged. We conclude that lung K(Ca) channel mRNA expression is decreased in an ovine model of PPHN. Decreased K(Ca) channel gene expression may contribute to the abnormal pulmonary vascular reactivity associated with PPHN.     

Hypoxia-induced inhibition of converting enzyme activity: role in vascular regulation

Systemic and pulmonary vascular reactivity to graded doses of angiotensin I (ANG I), angiotensin II (ANG II), and, as a control, phenylephrine were examined in 14- or 28-day hypoxia-exposed and air control rats. Hypoxic rats exhibited pulmonary hypertension that was reversible on return to room air, but systemic arterial pressure was not altered by hypoxia. Systemic pressor responses to ANG I and ANG II were significantly less in the hypoxic rats than in the control rats at 14 and 28 days but returned to control levels in hypoxic animals that were then returned to room air, demonstrating reversibility of the hypoxia-induced changes in vascular reactivity. Pulmonary pressor responses to ANG I were significantly less at 14 days, whereas responses to ANG II were significantly greater at 28 days, in hypoxic rats than in controls. There were no significant differences in systemic and pulmonary pressor responses to phenylephrine between the hypoxic and air control animals. The altered systemic and pulmonary pressor responsiveness to ANG I and ANG II in hypoxic rats is probably related to mechanisms specific to the renin-angiotensin system, such as inhibition of intrapulmonary angiotensin-converting enzyme activity and down regulation of ANG II receptors in the systemic circulation. Further study is needed to elucidate these mechanisms.     

Cyclooxygenase-2 and an early stage of chronic hypoxia-induced pulmonary hypertension in newborn pigs.

Our objective was to determine whether cyclooxygenase (COX)-2-dependent metabolites contribute to the altered pulmonary vascular responses that manifest in piglets with chronic hypoxia-induced pulmonary hypertension. Piglets were raised in either room air (control) or hypoxia for 3 days. The effect of the COX-2 selective inhibitor NS-398 on responses to arachidonic acid or acetylcholine (ACh) was measured in endothelium-intact and denuded pulmonary arteries (100- to 400-microm diameter). Pulmonary arterial production of the stable metabolites of thromboxane and prostacyclin was assessed in the presence and absence of NS-398. Dilation to arachidonic acid was greater for intact control than for intact hypoxic arteries, was unchanged by NS-398 in intact arteries of either group, and was augmented by NS-398 in denuded hypoxic arteries. ACh responses, which were dilation in intact control arteries but constriction in intact and denuded hypoxic arteries, were diminished by NS-398 treatment of all arteries. NS-398 reduced prostacyclin production by control pulmonary arteries and reduced thromboxane production by hypoxic pulmonary arteries. COX-2-dependent contracting factors, such as thromboxane, contribute to aberrant pulmonary arterial responses in piglets exposed to 3 days of hypoxia.     

Chronic hypoxia attenuates nitric oxide-dependent hemodynamic responses to acute hypoxia

Alterations in the nitric oxide (NO) pathway have been implicated in the pathogenesis of chronic hypoxia-induced pulmonary hypertension. Chronic hypoxia can either suppress the NO pathway, causing pulmonary hypertension, or increase NO release in order to counteract elevated pulmonary arterial pressure. We determined the effect of NO synthase inhibitor on hemodynamic responses to acute hypoxia (10% O(2)) in anesthetized rats following chronic exposure to hypobaric hypoxia (0.5 atm, air). In rats raised under normoxic conditions, acute hypoxia caused profound systemic hypotension and slight pulmonary hypertension without altering cardiac output. The total systemic vascular resistance (SVR) decreased by 41 +/- 5%, whereas the pulmonary vascular resistance (PVR) increased by 25 +/- 6% during acute hypoxia. Pretreatment with N(omega)-nitro-L-arginine methyl ester (L-NAME; 25 mg/kg) attenuated systemic vasodilatation and enhanced pulmonary vasoconstriction. In rats with prior exposure to chronic hypobaric hypoxia, the baseline values of mean pulmonary and systemic arterial pressure were significantly higher than those in the normoxic group. Chronic hypoxia caused right ventricular hypertrophy, as evidenced by a greater weight ratio of the right ventricle to the left ventricle and the interventricular septum compared to the normoxic group (46 +/- 4 vs. 28 +/- 3%). In rats which were previously exposed to chronic hypoxia (half room air for 15 days), acute hypoxia reduced SVR by 14 +/- 6% and increased PVR by 17 +/- 4%. Pretreatment with L-NAME further inhibited the systemic vasodilatation effect of acute hypoxia, but did not enhance pulmonary vasoconstriction. Our results suggest that the release of NO counteracts pulmonary vasoconstriction but lowers systemic vasodilatation on exposure to acute hypoxia, and these responses are attenuated following adaptation to chronic hypoxia.