sildenafil-citrate and Hyperoxia

Bronchopulmonary dysplasia (BPD), a common complication of preterm birth, is associated with pulmonary hypertension (PH) in 25% of infants with moderate to severe BPD. Neonatal mice exposed to hyperoxia for 14d develop lung disease similar to BPD, with evidence of associated PH. The cyclic guanosine monophosphate (cGMP) signaling pathway has not been well studied in BPD-associated PH. In addition, there is little data about the natural history of hyperoxia-induced PH in mice or the utility of phosphodiesterase-5 (PDE5) inhibition in established disease. C57BL/6 mice were placed in room air or 75% O2 within 24h of birth for 14d, followed by recovery in room air for an additional 7 days (21d). Additional pups were treated with either vehicle or sildenafil for 7d during room air recovery. Mean alveolar area, pulmonary artery (PA) medial wall thickness (MWT), RVH, and vessel density were evaluated at 21d. PA protein from 21d animals was analyzed for soluble guanylate cyclase (sGC) activity, PDE5 activity, and cGMP levels. Neonatal hyperoxia exposure results in persistent alveolar simplification, RVH, decreased vessel density, increased MWT, and disrupted cGMP signaling despite a period of room air recovery. Delayed treatment with sildenafil during room air recovery is associated with improved RVH and decreased PA PDE5 activity, but does not have significant effects on alveolar simplification, PA remodeling, or vessel density. These data are consistent with clinical studies suggesting inconsistent effects of sildenafil treatment in infants with BPD-associated PH.

Topics: Animals; Animals, Newborn; Bronchopulmonary Dysplasia; Cyclic GMP; Cyclic Nucleotide Phosphodiesterases, Type 5; Disease Models, Animal; Guanylate Cyclase; Hyperoxia; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Lung; Mice; Mice, Inbred C57BL; Oxygen; Phosphodiesterase 5 Inhibitors; Pulmonary Alveoli; Pulmonary Artery; Signal Transduction; Sildenafil Citrate; Vascular Remodeling

Pulmonary hypertension (PH) and right ventricular hypertrophy (RVH) affect 25-35% of premature infants with significant bronchopulmonary dysplasia (BPD), increasing morbidity and mortality. We sought to determine the role of phosphodiesterase 5 (PDE5) in the right ventricle (RV) and left ventricle (LV) in a hyperoxia-induced neonatal mouse model of PH and RVH. After birth, C57BL/6 mice were placed in room air (RA) or 75% O2 (CH) for 14 days to induce PH and RVH. Mice were euthanized at 14 days or recovered in RA for 14 days or 42 days prior to euthanasia at 28 or 56 days of age. Some pups received sildenafil or vehicle (3 mg·kg(-1)·dose(-1) sc) every other day from P0. RVH was assessed by Fulton's index [RV wt/(LV + septum) wt]. PDE5 protein expression was analyzed via Western blot, PDE5 activity was measured by commercially available assay, and cGMP was measured by enzyme-linked immunoassay. Hyperoxia induced RVH in mice after 14 days, and RVH did not resolve until 56 days of age. Hyperoxia increased PDE5 expression and activity in RV, but not LV + S, after 14 days. PDE5 expression normalized by 28 days of age, but PDE5 activity did not normalize until 56 days of age. Sildenafil given during hyperoxia prevented RVH, decreased RV PDE5 activity, and increased RV cGMP levels. Mice with cardiac-specific overexpression of PDE5 had increased RVH in RA. These findings suggest normal RV PDE5 function is disrupted by hyperoxia, and elevated PDE5 contributes to RVH and remodeling. Therefore, in addition to impacting the pulmonary vasculature, sildenafil also targets PDE5 in the neonatal mouse RV and decreases RVH.

Topics: Animals; Animals, Newborn; Antihypertensive Agents; Cyclic AMP; Cyclic Nucleotide Phosphodiesterases, Type 5; Disease Models, Animal; Down-Regulation; Heart Ventricles; Hyperoxia; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Mice, Inbred C57BL; Mice, Transgenic; Phosphodiesterase 5 Inhibitors; Piperazines; Purines; Second Messenger Systems; Sildenafil Citrate; Sulfonamides; Time Factors; Ventricular Function, Right; Ventricular Remodeling

Pulmonary hypertension (PH) occurs in 25 to 35% of premature infants with significant bronchopulmonary dysplasia (BPD). Neonatal mice exposed to 14 days of hyperoxia develop BPD-like lung injury and PH. To determinne the impact of hyperoxia on pulmonary artery (PA) cyclic guanosine monophosphate (cGMP) signaling in a murine model of lung injury and PH, neonatal C57BL/6 mice were placed in room air, 75% O2 for 14 days (chronic hyperoxia [CH]) or 75% O2 for 24 hours, followed by 13 days of room air (acute hyperoxia with recovery [AHR]) with or without sildenafil. At 14 days, mean alveolar area, PA medial wall thickness (MWT), right ventricular hypertrophy (RVH), and vessel density were assessed. PA protein was analyzed for cGMP, soluble guanylate cyclase, and PDE5 activity. CH and AHR mice had RVH, but only CH mice had increased alveolar area and MWT and decreased vessel density. In CH and AHR PAs, soluble guanylate cyclase activity was decreased, and PDE5 activity was increased. In CH mice, sildenafil attenuated MWT and RVH but did not improve mean alveolar area or vessel density. In CH and AHR PAs, sildenafil decreased PDE5 activity and increased cGMP. Our results indicate that prolonged hyperoxia leads to lung injury, PH, RVH, and disrupted PA cGMP signaling. Furthermore, 24 hours of hyperoxia causes RVH and disrupted PA cGMP signaling that persists for 13 days. Sildenafil reduced RVH and restored vascular cGMP signaling but did not attenuate lung injury. Thus, hyperoxia can rapidly disrupt PA cGMP signaling in vivo with sustained effects, and concurrent sildenafil therapy can be protective.

Topics: Animals; Cyclic GMP; Guanosine Monophosphate; Hyperoxia; Hypertension, Pulmonary; Lung; Lung Injury; Mice; Mice, Inbred C57BL; Piperazines; Pulmonary Artery; Purines; Signal Transduction; Sildenafil Citrate; Sulfones

Congenital diaphragmatic hernia (CDH) is clinically challenging because of associated lung hypoplasia (LH). There have been no validated parameters to evaluate fetal LH severity. Sildenafil has been shown to improve LH mass in nitrofen-induced pulmonary artery (PA) models, but the pulmonary vascular tone has not been evaluated in vivo. The aim of this study was to identify the PA Doppler parameter that best predicts LH severity and to investigate the efficacy of antenatal sildenafil treatment in experimental CDH.. Nitrofen (50-60% CDH in offspring) or vehicle on E9.5 and sildenafil or vehicle on E11.5-20.5 were administrated to pregnant rats. On E20.5, PA Doppler indices were investigated with and without maternal hyperoxia. The presence/absence of CDH, lung/body weight ratio and radial saccular count were assessed at E20.5.. At baseline, CDH rats had lower PA Doppler acceleration/ejection time ratios and pulsatility index (PI). Maternal hyperoxia resulted in a significant decrease in the PA/PI suggesting pulmonary vasodilation. In contrast, in CDH fetuses, the ipsilateral PA/PI showed little or no response to hyperoxia (P > .05), and in those with LH, PI response to maternal hyperoxia correlated positively with hernia, lung/body weight ratio (r = 0.70, P = .01). Maternal sildenafil therapy significantly improved PA response to hyperoxia and lung growth in CDH fetuses (P < .01).. Pulmonary vasodilation that occurs in E20.5 fetal rats in response to maternal hyperoxia is blunted in CDH. Change in PA/PI with hyperoxia is a useful predictor of LH severity. Sildenafil improves pulmonary vascular response and lung growth in fetal CDH.

Topics: Animals; Female; Hernia, Diaphragmatic; Hernias, Diaphragmatic, Congenital; Hyperoxia; Lung; Piperazines; Pregnancy; Pulmonary Artery; Purines; Rats; Rats, Sprague-Dawley; Regional Blood Flow; Sildenafil Citrate; Sulfones; Vasodilation

Hyperoxia exposure leads to the development of lung injury and bronchial hyperreactivity (BHR) via involvement of nitric oxide (NO) pathway. We aimed at characterizing whether the stimulation of the NO pathway by sildenafil or vasoactive intestinal peptide (VIP) is able to prevent the hyperoxia-induced development of BHR. The respective roles of the preserved lung volume and alveolar architecture, the anti-inflammatory and anti-apoptotic potentials of these treatments in the diminished lung responsiveness were also characterized.. Immature (28-day-old) rats were exposed for 72 hours to room air (Group C), hyperoxia (>95%, Group HC), or hyperoxia with the concomitant administration of vasoactive intestinal peptide (VIP, Group HV) or sildenafil (Group HS). Following exposure, the end-expiratory lung volume (EELV) was assessed plethysmographically. Airway and respiratory tissue mechanics were measured under baseline conditions and following incremental doses of methacholine to assess BHR. Inflammation was assessed by analyzing the bronchoalveolar lavage fluid (BALF), while biochemical and histological analyses were used to characterize the apoptotic and structural changes in the lungs.. The BHR, the increased EELV, the aberrant alveolarization, and the infiltration of inflammatory cells into the BALF that developed in Group HC were all suppressed significantly by VIP or sildenafil treatment. The number of apoptotic cells increased significantly in Group HC, with no evidence of statistically significant effects on this adverse change in Groups HS and HV.. These findings suggest that stimulating the NO pathway by sildenafil and VIP exert their beneficial effect against hyperoxia-induced BHR via preserving normal EELV, inhibiting airway inflammation and preserving the physiological lung structure, whereas the antiapoptotic potential of these treatments were not apparent in this process.

Topics: Animals; Bronchial Hyperreactivity; Hyperoxia; Lung; Male; Piperazines; Purines; Rats; Rats, Sprague-Dawley; Sildenafil Citrate; Sulfonamides; Vasoactive Intestinal Peptide

Blood flow through intrapulmonary arteriovenous anastomoses (IPAVAs) is known to increase in healthy humans during exercise while breathing room air, but is prevented or significantly reduced during exercise while breathing 100% O2, potentially due to vasoconstriction of IPAVAs. Thus, pharmacological interventions that target known pathways regulating the cardiopulmonary circulation may be able to prevent the hyperoxia-induced reduction in IPAVA blood flow (Q̇ IPAVA ) during exercise. In nine healthy human subjects, we investigated the effects of sildenafil (100 mg p.o.), nifedipine (20 mg p.o.) and acetazolamide (250 mg p.o. three times a day for 3 days) on Q̇ IPAVA at rest and during cycle ergometer exercise at 50, 100, 150, 200 and 250 W, while breathing room air (normoxia) and 100% O2 (hyperoxia). Transthoracic saline contrast echocardiography and a 0-5 bubble scoring system were used to detect and assess Q̇ IPAVA qualitatively; ultrasound was used to assess the blood flow velocity oftricuspid regurgitation and the left ventricular outflow tract blood flow to calculate pulmonary artery systolic pressure (PASP) and cardiac output, respectively. Without drugs, bubble scores increased significantly to ≥2 at 150 W in normoxia and to ≤2 at 200 W in hyperoxia. Only nifedipine consistently increased cardiac output at rest and during low-intensity exercise in normoxia and hyperoxia. However, there was no detectable effect of any drug on Q̇ IPAVA ; specifically, bubble scores were the same during exercise in either normoxia or hyperoxia. Accordingly, the reduction in Q̇ IPAVA during exercise while breathing 100% O2 is likely not to be due to the independent pharmacological mechanisms of action associated with sildenafil, nifedipine or acetazolamide.

Topics: Acetazolamide; Adult; Arteriovenous Anastomosis; Blood Flow Velocity; Exercise; Exercise Test; Female; Hemodynamics; Humans; Hyperoxia; Lung; Male; Nifedipine; Oxygen; Piperazines; Pulmonary Circulation; Purines; Regional Blood Flow; Respiration; Sildenafil Citrate; Sulfonamides; Young Adult

Oxygen is a pulmonary vasodilator, but data suggest high O(2) concentrations impede that response. We previously reported 24 h of 100% O(2) increased phosphodiesterase 5 (PDE5) activity in fetal pulmonary artery smooth muscle cells (FPASMC) and in ventilated neonatal lambs. PDE5 degrades cyclic GMP (cGMP) and inhibits nitric oxide (NO)-mediated cGMP-dependent vasorelaxation. We sought to determine the mechanism by which hyperoxia initiates reactive oxygen species (ROS) production and regulates PDE5.. Thirty minutes of hyperoxia increased mitochondrial ROS versus normoxia (30.3±1.7% vs. 21.1±2.8%), but had no effect on cytosolic ROS, measured by roGFP, a ratiometric protein thiol redox sensor. Hyperoxia increased PDE5 activity (220±39%) and decreased cGMP responsiveness to NO (37±17%). Mitochondrial catalase overexpression attenuated hyperoxia-induced mitochondrial roGFP oxidation, compared to FPASMC infected with empty adenoviral vector (50±3% of control) or mitochondrial superoxide dismutase. MitoTEMPO, mitochondrial catalase, and DT-3, a cGMP-dependent protein kinase I alpha inhibitor, decreased PDE5 activity (32±13%, 26±21%, and 63±10% of control, respectively), and restored cGMP responsiveness to NO (147±16%,172±29%, and 189±43% of control, respectively). C57Bl6 mice exposed to 90%-100% O(2) for 45 min±mechanical ventilation had increased PA PDE5 activity (206±39% and 235±75%, respectively).. This is the first description that hyperoxia induces ROS in the mitochondrial matrix prior to the cytosol. Our results indicate that short hyperoxia exposures can produce significant changes in critical cellular signaling pathways.. These results indicate that mitochondrial matrix oxidant signals generated during hyperoxia, specifically H(2)O(2), activate PDE5 in a cGMP-dependent protein kinase-dependent manner in pulmonary vascular smooth muscle cells.

Topics: Animals; Antioxidants; Catalase; Cell Hypoxia; Cells, Cultured; Cyclic GMP; Cyclic GMP-Dependent Protein Kinase Type I; Cyclic GMP-Dependent Protein Kinases; Cyclic Nucleotide Phosphodiesterases, Type 5; Enzyme Activation; Fetus; Hydrogen Peroxide; Hyperoxia; Male; Mice; Mice, Inbred C57BL; Mitochondria, Muscle; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Nitric Oxide; Organophosphorus Compounds; Oxidation-Reduction; Phosphodiesterase 5 Inhibitors; Piperazines; Piperidines; Pulmonary Artery; Purines; Sheep, Domestic; Sildenafil Citrate; Sulfones; Superoxide Dismutase

Phosphodiesterase-5 inhibition with sildenafil has been used to treat severe pulmonary hypertension and bronchopulmonary dysplasia (BPD), a chronic lung disease in very preterm infants who were mechanically ventilated for respiratory distress syndrome.. Sildenafil treatment was investigated in 2 models of experimental BPD: a lethal neonatal model, in which rat pups were continuously exposed to hyperoxia and treated daily with sildenafil (50-150 mg/kg body weight/day; injected subcutaneously) and a neonatal lung injury-recovery model in which rat pups were exposed to hyperoxia for 9 days, followed by 9 days of recovery in room air and started sildenafil treatment on day 6 of hyperoxia exposure. Parameters investigated include survival, histopathology, fibrin deposition, alveolar vascular leakage, right ventricular hypertrophy, and differential mRNA expression in lung and heart tissue.. Prophylactic treatment with an optimal dose of sildenafil (2 x 50 mg/kg/day) significantly increased lung cGMP levels, prolonged median survival, reduced fibrin deposition, total protein content in bronchoalveolar lavage fluid, inflammation and septum thickness. Treatment with sildenafil partially corrected the differential mRNA expression of amphiregulin, plasminogen activator inhibitor-1, fibroblast growth factor receptor-4 and vascular endothelial growth factor receptor-2 in the lung and of brain and c-type natriuretic peptides and the natriuretic peptide receptors NPR-A, -B, and -C in the right ventricle. In the lethal and injury-recovery model we demonstrated improved alveolarization and angiogenesis by attenuating mean linear intercept and arteriolar wall thickness and increasing pulmonary blood vessel density, and right ventricular hypertrophy (RVH).. Sildenafil treatment, started simultaneously with exposure to hyperoxia after birth, prolongs survival, increases pulmonary cGMP levels, reduces the pulmonary inflammatory response, fibrin deposition and RVH, and stimulates alveolarization. Initiation of sildenafil treatment after hyperoxic lung injury and continued during room air recovery improves alveolarization and restores pulmonary angiogenesis and RVH in experimental BPD.

Topics: Animals; Animals, Newborn; Disease Models, Animal; Fibrin; Humans; Hyperoxia; Hypertrophy, Right Ventricular; Lung Injury; Phosphodiesterase Inhibitors; Piperazines; Pneumonia; Purines; Rats; Rats, Wistar; Sildenafil Citrate; Sulfones; Survival Analysis; Survival Rate; Treatment Outcome