scholarly journals Tetrahydrobiopterin oral therapy recouples eNOS and ameliorates chronic hypoxia-induced pulmonary hypertension in newborn pigs

2016 ◽  
Vol 311 (4) ◽  
pp. L743-L753 ◽  
Author(s):  
Anna Dikalova ◽  
Judy L. Aschner ◽  
Mark R. Kaplowitz ◽  
Marshall Summar ◽  
Candice D. Fike
Keyword(s):  
Pulmonary Hypertension ◽  
Chronic Hypoxia ◽  
Disease Onset ◽  
Pulmonary Arteries ◽  
Pulmonary Vasculature ◽  
Hypoxic Exposure ◽  
No Production ◽  
Major Purpose ◽  
Newborn Piglets ◽  
Enos Uncoupling

We previously showed that newborn piglets who develop pulmonary hypertension during exposure to chronic hypoxia have diminished pulmonary vascular nitric oxide (NO) production and evidence of endothelial NO synthase (eNOS) uncoupling (Fike CD, Dikalova A, Kaplowitz MR, Cunningham G, Summar M, Aschner JL. Am J Respir Cell Mol Biol 53: 255–264, 2015). Tetrahydrobiopterin (BH4) is a cofactor that promotes eNOS coupling. Current clinical strategies typically invoke initiating treatment after the diagnosis of pulmonary hypertension, rather than prophylactically. The major purpose of this study was to determine whether starting treatment with an oral BH4 compound, sapropterin dihydrochloride (sapropterin), after the onset of pulmonary hypertension would recouple eNOS in the pulmonary vasculature and ameliorate disease progression in chronically hypoxic piglets. Normoxic (control) and hypoxic piglets were studied. Some hypoxic piglets received oral sapropterin starting on day 3 of hypoxia and continued throughout an additional 7 days of hypoxic exposure. Catheters were placed for hemodynamic measurements, and pulmonary arteries were dissected to assess eNOS dimer-to-monomer ratios (a measure of eNOS coupling), NO production, and superoxide (O2·−) generation. Although higher than in normoxic controls, pulmonary vascular resistance was lower in sapropterin-treated hypoxic piglets than in untreated hypoxic piglets. Consistent with eNOS recoupling, eNOS dimer-to-monomer ratios and NO production were greater and O2·− generation was less in pulmonary arteries from sapropterin-treated than untreated hypoxic animals. When started after disease onset, oral sapropterin treatment inhibits chronic hypoxia-induced pulmonary hypertension at least in part by recoupling eNOS in the pulmonary vasculature of newborn piglets. Rescue treatment with sapropterin may be an effective strategy to inhibit further development of pulmonary hypertension in newborn infants suffering from chronic cardiopulmonary conditions associated with episodes of prolonged hypoxia.

scholarly journals Combined l-citrulline and tetrahydrobiopterin therapy improves NO signaling and ameliorates chronic hypoxia-induced pulmonary hypertension in newborn pigs

2020 ◽  
Vol 318 (4) ◽  
pp. L762-L772 ◽  
Author(s):  
Anna Dikalova ◽  
Judy L. Aschner ◽  
Mark R. Kaplowitz ◽  
Gary Cunningham ◽  
Marshall Summar ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Chronic Hypoxia ◽  
Pulmonary Arteries ◽  
No Production ◽  
Enos Uncoupling ◽  
No Signaling ◽  
Newborn Pigs ◽  
Clinical Conditions ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide

Newborn pigs with chronic hypoxia-induced pulmonary hypertension (PH) have evidence of endothelial nitric oxide synthase (eNOS) uncoupling. In this model, we showed that therapies that promote eNOS coupling, either tetrahydrobiopterin (BH4), a NOS cofactor, or l-citrulline, a NO-l-arginine precursor, inhibit PH. We wanted to determine whether cotreatment with l-citrulline and a BH4 compound, sapropterin dihydrochloride, improves NO signaling and chronic hypoxia-induced PH more markedly than either alone. Normoxic (control) and hypoxic piglets were studied. Some hypoxic piglets received sole treatment with l-citrulline or BH4, or were cotreated with l-citrulline and BH4, from day 3 through day 10 of hypoxia. Catheters were placed for hemodynamic measurements, and pulmonary arteries were dissected to assess eNOS dimer-to-monomer ratios and NO production. In untreated hypoxic piglets, pulmonary vascular resistance (PVR) was higher and NO production and eNOS dimer-to-monomer ratios were lower than in normoxic piglets. Compared with the untreated hypoxic group, PVR was lower in hypoxic piglets cotreated with l-citrulline and BH4 and in those treated with l-citrulline alone but not for those treated solely with BH4. NO production and eNOS dimer-to-monomer ratios were greater for all three treated hypoxic groups compared with the untreated group. Notably, greater improvements in PVR, eNOS dimer-to-monomer ratios, and NO production were found in hypoxic piglets cotreated with l-citrulline and BH4 than in piglets treated with either alone. Cotreatment with l-citrulline and BH4 more effectively improves NO signaling and inhibits chronic hypoxia-induced PH than either treatment alone. Combination therapies may offer enhanced therapeutic capacity for challenging clinical conditions, such as chronic neonatal PH.

Thapsigargin stimulates increased NO activity in hypoxic hypertensive rat lungs and pulmonary arteries

1996 ◽  
Vol 80 (4) ◽  
pp. 1336-1344 ◽  
Author(s):  
M. Muramatsu ◽  
R. C. Tyler ◽  
D. M. Rodman ◽  
I. F. McMurtry
Keyword(s):  
Pulmonary Artery ◽  
Chronic Hypoxia ◽  
Pulmonary Arteries ◽  
Male Rats ◽  
Pulmonary Vasculature ◽  
No Production ◽  
Low Concentrations ◽  
Pulmonary Vascular Endothelium ◽  
Increased Sensitivity ◽  
Stimulation Of

This study addressed the controversy of whether endothelium-derived nitric oxide (NO) activity is increased or decreased in the hypertensive pulmonary vasculature of chronically hypoxic rats. Thapsigargin, a receptor-independent Ca2+ agonist and stimulator of endothelial NO production, was used to compare NO-mediated vasodilation in perfused lungs and conduit pulmonary artery rings isolated from adult male rats either kept at Denver's altitude of 5,280 ft (control pulmonary normotensive rats) or exposed for 4-5 wk to the simulated altitude of 17,000 ft (chronically hypoxic pulmonary hypertensive rats). Under baseline conditions, thapsigargin (10(-9)-10(-7) M) caused vasodilation in hypertensive lungs and vasoconstriction in normotensive lungs. Whereas the sustained vasodilation in hypertensive lungs was reversed to vasoconstriction by the inhibitor of NO synthase N(omega)-nitro-L-arginine (L-NNA; 10(-4) M), a transient vasodilation to thapsigargin in acutely vasoconstricted normotensive lungs was potentiated. As measured by a chemiluminescence assay, the recirculated perfusate of hypertensive lungs accumulated considerably higher levels of NO-containing compounds that did normotensive lungs, and thapsigargin-induced stimulation of NO-containing compounds accumulation was greater in hypertensive than in normotensive lungs. Similarly, low concentrations of thapsigargin (10(-10)-10(-9) M) caused greater endothelium-dependent L-NNA-reversible relaxation of hypertensive than of normotensive pulmonary artery rings. The increased sensitivity of hypertensive arteries to thapsigargin-induced relaxation was eliminated in nominally Ca(2+)-free medium and was not mimicked by ryanodine, a releaser of intracellular Ca2+. These results with thapsigargin, which acts on endothelial cells to stimulate Ca2+ influx and a sustained rise in intracellular Ca2+ concentration, support the idea that pulmonary vascular endothelium-derived NO activity is increased rather than decreased in chronic hypoxia-induced pulmonary hypertension in rats.

Impaired NO signaling in small pulmonary arteries of chronically hypoxic newborn piglets

2004 ◽  
Vol 286 (6) ◽  
pp. L1244-L1254 ◽  
Author(s):  
Candice D. Fike ◽  
Judy L. Aschner ◽  
Yongmei Zhang ◽  
Mark R. Kaplowitz
Keyword(s):  
Chronic Hypoxia ◽  
Pulmonary Arteries ◽  
Exposure Period ◽  
No Production ◽  
Resistance Level ◽  
Caveolin 1 ◽  
Selective Antagonist ◽  
Newborn Piglets ◽  
No Signaling ◽  
Nos Isoforms

We performed studies to determine whether chronic hypoxia impairs nitric oxide (NO) signaling in resistance level pulmonary arteries (PAs) of newborn piglets. Piglets were maintained in room air (control) or hypoxia (11% O2) for either 3 (shorter exposure) or 10 (longer exposure) days. Responses of PAs to a nonselective NO synthase (NOS) antagonist, Nω-nitro-l-arginine methylester (l-NAME), a NOS-2-selective antagonist, aminoguanidine, and 7-nitroindazole, a NOS-1-selective antagonist, were measured. Levels of NOS isoforms and of two proteins involved in NOS signaling, heat shock protein (HSP) 90 and caveolin-1, were assessed in PA homogenates. PAs from all groups constricted to l-NAME but not to aminoguanidine or 7-nitroindazole. The magnitude of constriction to l-NAME was similar for PAs from control and hypoxic piglets of the shorter exposure period but was diminished for PAs from hypoxic compared with control piglets of the longer exposure period. NOS-3, HSP90, and caveolin-1 levels were similar in hypoxic and control PAs. These findings indicate that NOS-3, but not-NOS 2 or NOS-1, is involved with basal NO production in PAs from both control and hypoxic piglets. After 10 days of hypoxia, NO function is impaired in PAs despite preserved levels of NOS-3, HSP90, and caveolin-1. The development of NOS-3 dysfunction in resistance level PAs may contribute to the progression of chronic hypoxia-induced pulmonary hypertension in newborn piglets.

scholarly journals l-Citrulline ameliorates chronic hypoxia-induced pulmonary hypertension in newborn piglets

2009 ◽  
Vol 297 (3) ◽  
pp. L506-L511 ◽  
Author(s):  
Madhumita Ananthakrishnan ◽  
Frederick E. Barr ◽  
Marshall L. Summar ◽  
Heidi A. Smith ◽  
Mark Kaplowitz ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Chronic Hypoxia ◽  
Vascular Endothelial Cells ◽  
No Production ◽  
Nitrate Accumulation ◽  
Newborn Piglets ◽  
Pulmonary Vascular Endothelium ◽  
Isolated Lungs ◽  
Nitrite Nitrate

Newborn piglets develop pulmonary hypertension and have diminished pulmonary vascular nitric oxide (NO) production when exposed to chronic hypoxia. NO is produced by endothelial NO synthase (eNOS) in the pulmonary vascular endothelium using l-arginine as a substrate and producing l-citrulline as a byproduct. l-Citrulline is metabolized to l-arginine by two enzymes that are colocated with eNOS in pulmonary vascular endothelial cells. The purpose of this study was to determine whether oral supplementation with l-citrulline during exposure of newborn piglets to 10 days of chronic hypoxia would prevent the development of pulmonary hypertension and increase pulmonary NO production. A total of 17 hypoxic and 17 normoxic control piglets were studied. Six of the 17 hypoxic piglets were supplemented with oral l-citrulline starting on the first day of hypoxia. l-Citrulline supplementation was provided orally twice a day. After 10 days of hypoxia or normoxia, the animals were anesthetized, hemodynamic measurements were performed, and the lungs were perfused in situ. Pulmonary arterial pressure and pulmonary vascular resistance were significantly lower in hypoxic animals treated with l-citrulline compared with untreated hypoxic animals ( P < 0.001). In vivo exhaled NO production ( P = 0.03) and nitrite/nitrate accumulation in the perfusate of isolated lungs ( P = 0.04) were significantly higher in l-citrulline-treated hypoxic animals compared with untreated hypoxic animals. l-Citrulline supplementation ameliorated the development of pulmonary hypertension and increased NO production in piglets exposed to chronic hypoxia. We speculate that l-citrulline may benefit neonates exposed to prolonged periods of hypoxia from cardiac or pulmonary causes.

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

2007 ◽  
Vol 293 (6) ◽  
pp. L1475-L1482 ◽  
Author(s):  
Christine E. Bixby ◽  
Basil O. Ibe ◽  
May F. Abdallah ◽  
Weilin Zhou ◽  
Alison A. Hislop ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Protein Expression ◽  
High Altitude ◽  
Vascular Remodeling ◽  
Chronic Hypoxia ◽  
Pulmonary Arteries ◽  
Platelet Activating Factor ◽  
Pulmonary Vasculature ◽  
Pulmonary Vascular Remodeling ◽  
Mediated Effects

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 ∼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 106 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.

Exhaled NO is reduced at an early stage of hypoxia-induced pulmonary hypertension in newborn piglets

2003 ◽  
Vol 284 (3) ◽  
pp. L489-L500 ◽  
Author(s):  
Joyce E. Turley ◽  
Leif D. Nelin ◽  
Mark R. Kaplowitz ◽  
Yongmei Zhang ◽  
Candice D. Fike
Keyword(s):  
Nitric Oxide ◽  
Pulmonary Hypertension ◽  
Early Stage ◽  
Pulmonary Arteries ◽  
No Production ◽  
Newborn Piglets ◽  
Exhaled No ◽  
Isolated Lungs ◽  
Air Control ◽  
Nos Isoforms

Altered nitric oxide (NO) production could contribute to the pathogenesis of hypoxia-induced pulmonary hypertension. To determine whether parameters of lung NO are altered at an early stage of hypoxia-induced pulmonary hypertension, newborn piglets were exposed to room air (control, n = 21) or 10% O2 (hypoxia, n = 19) for 3–4 days. Some lungs were isolated and perfused for measurement of exhaled NO output and the perfusate accumulation of nitrite and nitrate (NOx−), the stable metabolites of NO. Pulmonary arteries (20–600-μm diameter) and their accompanying airways were dissected from other lungs and incubated for NOx− determination. Abundances of the nitric oxide synthase (NOS) isoforms endothelial NOS and neural NOS were assessed in homogenates of PAs and airways. The perfusate NOx− accumulation was similar, whereas exhaled NO output was lower for isolated lungs of hypoxic, compared with control, piglets. The incubation solution NOx− did not differ between pulmonary arteries (PAs) of the two groups but was lower for airways of hypoxic, compared with control, piglets. Abundances of both eNOS and nNOS proteins were similar for PA homogenates from the two groups of piglets but were increased in airway homogenates of hypoxic compared with controls. The NO pathway is altered in airways, but not in PAs, at an early stage of hypoxia-induced pulmonary hypertension in newborn piglets.

scholarly journals L‐Citrulline Inhibits Chronic Hypoxia‐Induced Pulmonary Hypertension in Newborn Piglets

2008 ◽  
Vol 22 (S1) ◽  
Author(s):  
Candice Fike ◽  
Madhumita Ananthakrishnan ◽  
Mark Kaplowitz ◽  
Frederick Barr ◽  
Marshall Summar
Keyword(s):  
Pulmonary Hypertension ◽  
Chronic Hypoxia ◽  
Newborn Piglets

Abstract 15303: Dimethylarginine Dimethylaminohydrolase-1 is Not Involved in Chronic Hypoxic Pulmonary Hypertension and Right Ventricular Hypertrophy in Mice

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Juliane Hannemann ◽  
Antonia Glatzel ◽  
Jonas Hillig ◽  
Julia Zummack ◽  
Rainer H Boeger
Keyword(s):  
Pulmonary Hypertension ◽  
Right Ventricular ◽  
Ventricular Hypertrophy ◽  
Chronic Hypoxia ◽  
Right Ventricular Hypertrophy ◽  
Cardiomyocyte Hypertrophy ◽  
No Production ◽  
Knock Out ◽  
Knock Out Mice ◽  
Adma Concentration

Introduction: Chronic hypoxia causes persistent pulmonary vasoconstriction and leads to pulmonary hypertension and right ventricular hypertrophy. Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of NO synthesis; its level increases in hypoxia concomitantly with reduced activity of dimethylarginine dimethylaminohydrolases (DDAH-1 and DDAH-2), the enzymes metabolizing ADMA. DDAH knockout models may therefore help to understand the pathophysiological roles of this enzyme and its substrate, ADMA, in the development of hypoxia-associated pulmonary hypertension. Hypothesis: We hypothesized that DDAH1 knock-out mice have an attenuated hypoxia-induced elevation of ADMA and reduced right ventricular hypertrophy. Methods: DDAH1 knock-out mice (KO) and their wild-type littermates (WT) were subjected to normoxia (NX) or hypoxia (HX) during 21 days. We measured ADMA concentration in plasma, DDAH1 and DDAH2 expression in the lung, right ventricular hypertrophy by the Fulton index, cardiomyocyte hypertrophy by dystrophin staining of heart tissues, and muscularization of pulmonary arterioles by CD31 and α-actin staining of lung sections. Results: DDAH1 KO mice had higher ADMA concentration than WT under NX (2.31±0.33 μmol/l vs. 1.20±0.17 μmol/l; p < 0.05). ADMA significantly increased in WT-HX (to 1.74±0.86 μmol/l; p < 0.05 vs. normoxia), whilst it did not further increase in KO-HX (2.58±0.58 μmol/l; p = n.s.). This was paralleled by a 38±13% reduction in DDAH1 mRNA but not DDAH2 mRNA expression, and reduced DDAH protein expression. We observed right ventricular hypertrophy under hypoxia in both, WT and KO mice, with no significant differences between both genotypes. Further, cardiomyocyte hypertrophy and pulmonary arteriolar muscularization were significantly increased by hypoxia, but not significantly different between WT and KO mice. Conclusions: We conclude that chronic hypoxia causes an elevation of ADMA, which impairs NO production and leads to endothelial dysfunction and vasoconstriction. Downregulation of DDAH expression and activity may be involved in this; however, knockout of DDAH1 does not modify the pathophysiological changes in remodeling of the pulmonary vasculature and the right ventricle.

Abstract 14372: Identification of the Genes That Are Involved in Severe Pulmonary Hypertension in Type 2 Diabetic Mice

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Qiuyu Zheng ◽  
Atsumi Tsuji-Hosokawa ◽  
Jody T Cabrera ◽  
Jian Wang ◽  
Ayako Makino
Keyword(s):  
Pulmonary Hypertension ◽  
Endothelial Function ◽  
Protein Level ◽  
Chronic Hypoxia ◽  
Systolic Pressure ◽  
Vascular Complications ◽  
Hypoxic Exposure ◽  
Diabetic Mice ◽  
Type 2 Diabetic

Pulmonary hypertension (PH) is a progressive disease characterized by increased pulmonary vascular resistance. Increasing evidence shows that diabetes increases the risks of PH, and diabetic priming leads to severe PH. However, the molecular mechanism by which preconditioning of diabetes results in severe PH is still unknown. It has been shown that endothelium serves as a key regulator of vascular tone, and endothelial cell dysfunction is implicated in the development of PH and diabetes-related vascular complications. Therefore, we identified the genes that contribute to the development of severe PH in diabetic mice with a focus on endothelial function. We first determined the effect of chronic hypoxia (10% O 2 , 4 weeks) on hemodynamics in type 2 diabetic (T2D) mice. We used inducible T2D mice (generated by high-fat diet and a low-dose streptozotocin injection) and spontaneous T2D mice (TALLYHO/Jng). Diabetic mice exhibited a slight increase in right ventricular systolic pressure (RVSP), and chronic hypoxia led to a further rise in RVSP in both inducible and spontaneous T2D mice. We isolated pulmonary endothelial cells (MPEC) from normoxia-exposed control mice (CN), hypoxia-exposed control mice (CH), normoxia-exposed diabetic mice (DN), and hypoxia-exposed diabetic mice (DH) to examine the levels of 92 genes using real-time PCR. Nighty two genes were selected based on their functions, which are significantly related to endothelial function. We found that 27 genes were significantly changed among 4 groups. We then examined the protein levels of genes that were related to apoptosis and glycolysis. Western Blot data indicated that the protein level of GAPDH was significantly increased in CH and DH compared to CN and DN. In addition, hypoxic exposure in diabetic mice (DH) significantly increased HK2 protein level compare to hypoxia-exposed control mice (CH). These data suggest that precondition of diabetes increases susceptibility to developing PH due partly to altering gene expression of HK2 and Gapdh in MPECs. Since HK2 and GAPDH are a crucial regulator of glycolysis, alteration of glycolysis is expected in hypoxia-exposed diabetic mice. Our study revealed the key molecules which could be used for treating severe PH in diabetes.

Sign in / Sign up

Export Citation Format

Share Document