Ratio Of Right Ventricular Weight Research Articles

Increased Hemoglobin Affinity for Oxygen with GBT021601 Reduces Pulmonary Hypertension and Lung Vascular Inflammation in a Murine Model of Sickle Cell Disease

Background: Sickle cell disease (SCD) is caused by a single nucleotide change in the β-globin gene coding sequence that results in the production of hemoglobin S (HbS). Under conditions of red blood cell (RBC) deoxygenation, HbS polymerizes and leads to RBC sickling, resulting in hemolysis, anemia, diminished microvascular blood flow, vascular endothelial cell activation, vaso-occlusion, and ischemia. Despite improvements in treatment options, SCD remains a debilitating condition, and most patients experience progressive end-organ damage with associated complications and repeated hospitalizations. Pulmonary hypertension (PH), which is associated with increased mortality, is a common complication in patients with SCD, with nearly 10% of the SCD population reporting PH. PH in patients with SCD is multifactorial and may be caused by hemolysis, nitric oxide scavenging, and/or tissue hypoxia. Pharmacological increase of the concentration of oxygenated HbS (oxyHbS) in RBCs by voxelotor, a first-generation HbS polymerization inhibitor, reduces RBC sickling and preserves oxygen (O2) delivery by reducing anemia and improving blood rheology. Limited data are currently available on whether inhibiting HbS polymerization by increasing oxyHbS concentration may improve end-organ damage and associated complications, including PH. Objective: To examine the effects of GBT021601, a small-molecule, investigational, second-generation HbS polymerization inhibitor, on hemoglobin (Hb)-O2 affinity, PH, right ventricular dysfunction, and lung inflammation in a murine model of SCD. Methods: Townes HbSS mice treated with GBT021601 (75 mg/kg orally once daily for 14 days) were exposed to acute hypoxia (8% O2) for 1 hour and reoxygenated for 72 hours. Untreated HbAA Townes mice were used as controls. Lung inflammation was assessed in terms of accumulation of inflammatory cells, activation of nuclear factor kappa B subunit 1, and expression of intercellular adhesion molecule 1 and vascular cell adhesion molecule 1. To evaluate the impact of GBT021601 treatment on Hb-O2 affinity and anemia, p50 (partial O2 pressure at which Hb is 50% saturated with O2) and hematocrit levels were measured in SCD mice. In vivo pulmonary hemodynamic measurements included pulmonary artery pressure and right atrial pressure, whereas right ventricular hypertrophy was assessed using the Fulton index, which is the ratio of right ventricular weight to the weight of the left ventricle plus septum. Results: The total number of inflammatory cells and the specific numbers of lymphocytes, macrophages, and neutrophils were lower in untreated HbAA Townes mice and in HbSS Townes mice treated with GBT021601 compared with vehicle-dosed HbSS mice (Figure, panel A). Chronic treatment with GBT021601 increased Hb-O2 affinity, reducing the p50 in SCD mice from 32 mm Hg to 14 mm Hg. The treatment significantly reduced anemia, increasing hematocrit from 27.2% in the vehicle-dosed HbSS mice to 38.5% in the mice treated with GBT021601. Chronic GBT021601 treatment in HbSS mice also reduced pulmonary artery and right atrial pressure compared with those in untreated HbSS mice. Additionally, GBT021601 reduced right ventricular hypertrophy in HbSS mice, as evidenced by a reduction in the Fulton ratio (Figure, panel B). Conclusions: GBT021601 reduced PH, right ventricular dysfunction, and lung inflammation in a murine model of SCD. Our findings suggest that reducing RBC sickling, anemia, and hemolysis by increasing HbS affinity for O2 plays a critical role in modulating the immune system in PH and lung vasculature. These findings highlight the potential therapeutic efficacy of GBT021601 based on its ability to increase HbS affinity for O2 and prevent HbS polymerization and RBC sickling to reduce PH in SCD. Funding: Global Blood Therapeutics. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

The unexpected role of PI3-kinase gamma in pulmonary hypertension

Abstract Introduction Pulmonary hypertension (PH) is a pulmonary vascular disease that is associated with unacceptably high morbidity and mortality. PH is characterized by chronically increased pulmonary arterial pressure, increased pulmonary vascular resistance and right ventricular (RV) dysfunction and hypertrophy. Underlying mechanisms include increased proliferation and reduced apoptosis of both vascular smooth muscle cells (SMC) and endothelial cells (EC), as well as dysregulated immune responses. We have previously shown that class IA phosphatidylinositol-3-kinase (PI3K) isoforms, activated via receptor tyrosine kinases, are critically involved in the pathogenesis of PH. However, recent findings suggest that the class IB isoform PI3Kγ, which is activated downstream of G protein coupled receptors, is also important. It has been shown that PI3Kγ is involved in numerous processes that promote both vascular remodelling and maladaptive cardiac hypertrophy, including leukocyte recruitment, expression of proinflammatory chemokines and cytokines, as well as SMC and EC proliferation and survival. Therefore, the aim of our study was to investigate the role of PI3Kγ in the pathogenesis of PH. Methods The impact of PI3Kγ on the pathogenesis of PH was analysed in vivo using mice expressing a catalytically inactive form of PI3Kγ (PI3KγKD/KD) in the hypoxia-induced mouse model of PH. Mice were kept at 10%O2 (HOX) for 21 days or left under normoxic conditions (NOX). Subsequently, systolic right ventricular pressure (RVSP) was measured with a pressure catheter. RV hypertrophy was expressed as the ratio of RV weight to left ventricular + septum weight. Migration and proliferation of human pulmonary arterial SMC (hPASMC) as well as EC (hMVEC) were analysed using a PI3Kγ isoform-specific inhibitor (AS605240 [0.1; 0.3; 1μM]). Chemotaxis was determined by means of a modified Boyden chamber, and proliferation was quantified by a Bromodeoxyuridine (BrdU) incorporation assay. Results Whereas PI3Kγ inactivation had no effect on NOX animals, hypoxia led to increased RVSP and RV hypertrophy in WT animals (34.67±2.02 mmHg; 0.38±0.087) which were unexpectedly further increased in PI3KγKD/KD mice (37.67±1.3 mmHg, p=0.0104 vs. HOX WT; 0.47±0.06, p=0.0155 vs. HOX WT). Heart rate and systemic blood pressure remained unchanged. Inhibition of PI3Kγ by means of AS605240 did not affect proliferation of hPASMC and hMVEC, induced by multiple stimuli (FCS [10%], PDGF-BB [30ng/ml], or CXCL12 [100ng/ml], VEGF [50ng/ml]), respectively. However, FCS-induced migration of these cells was significantly reduced by AS605240 [0.3μM] (p<0.05). Conclusion Contrary to our expectations, the results show that kinase inactivation of PI3Kγ was not able to attenuate the pathogenesis of PH, but surprisingly led to a significant increase without critically changing cellular responses of SMC and EC. Therefore, our results indicate an unexpected protective effect of PI3Kγ on PH. Funding Acknowledgement Type of funding source: None

Triple-tyrosine kinase inhibition attenuates pulmonary arterial hypertension and neointimal formation

The present study examined the effects of simultaneous inhibition of vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF) and fibroblast growth factor (FGF) receptor signaling with BIBF1000, a novel triple tyrosine kinase inhibitor on preventing and reversing the progression of severe pulmonary arterial hypertension (PAH) in an experimental model in rats. Left pneumonectomized male Wistar rats were injected with monocrotaline to induce PAH. Treatment with BIBF1000 from day 1 to day 21 after monocrotaline injection attenuated PAH development, as evidenced by lower values for pulmonary artery pressure (mPAP), right ventricular pressure (RVSP), pulmonary arterial neointimal formation, and the ratio of right ventricular weight to left ventricular and septum weight [RV/(LV+S)] on day 21 compared to control rats. Treatment with BIBF1000 from day 21 to day 42 after monocrotaline injection reversed established PAH as shown by normalized values for mPAP and RVSP, RV/(LV+S) ratio, pulmonary arterial occlusion scores, levels of heart and lung fibrosis, as well as improved survival. Treatment with BIBF1000 reduced inflammatory cell recruitment in bronchoalveolar lavage and lung tissues, reduced CD-68 positive macrophages and expression of proliferating cell nuclear antigen in the perivascular areas, and reduced TNF-α and growth factor productions, and inhibited the phosphorylation of AKT and GSK3β in lungs. In addition, BIBF1000 inhibited pulmonary artery smooth muscle cells migration and proliferation from rat pulmonary artery explant cultures. Simultaneous inhibition of VEGF, PDGF, and FGF receptor signaling by BIBF1000 prevents and reverses the progression of severe pulmonary arterial hypertension and vascular remodeling in this experimental model.

Autoimmune disease mouse model exhibits pulmonary arterial hypertension.

BackgroundPulmonary arterial hypertension is often associated with connective tissue disease. Although there are some animal models of pulmonary hypertension, an autoimmune disease-based model has not yet been reported. MRL/lpr mice, which have hypergammaglobulinemia, produce various autoimmune antibodies, and develop vasculitis and nephritis spontaneously. However, little is known about pulmonary circulation in these mice. In the present study, we examined the pulmonary arterial pressure in MRL/lpr mice.Methods and resultsWe used female MRL/lpr mice aged between 12 and 14 weeks. Fluorescent immunostaining showed that there was no deposition of immunoglobulin or C3 in the lung tissue of the MRL/lpr mice. Elevation of interferon-γ and interleukin-6 was recognized in the lung tissue of the MRL/lpr mice. Right ventricular systolic pressure, Fulton index and the ratio of right ventricular weight to body weight in the MRL/lpr mice were significantly higher than those in wild type mice with same background (C57BL/6). The medial smooth muscle area and the proportion of muscularized vessels in the lung tissue of the MRL/lpr mice were larger than those of the C57BL/6 mice. Western blot analysis demonstrated markedly elevated levels of prepro-endothelin-1 and survivin as well as decreased endothelial nitric oxide synthase phosphorylation in the lung tissue of the MRL/lpr mice. Terminal deoxynucleotidyl-transferase-mediated dUTP nick end-labeling assay showed the resistance against apoptosis of pulmonary arterial smooth muscle cells in the MRL/lpr mice.ConclusionWe showed that MRL/lpr mice were complicated with pulmonary hypertension. MRL/lpr mice appeared to be a useful model for studying the mechanism of pulmonary hypertension associated with connective tissue diseases.

Abstract 11814: Pulmonary Hypertension With Altered eNOS and ALK-1 Signaling in the Autoimmuno-disease Mice

Background: Pulmonary artery hypertension is often associated with connective tissue disease and major determinant of prognosis. MRL/lpr mouse, which has hypergammaglobulinemia and products various autoimmuno antibody, develops vasculitis and nephritis spontaneously and is widely used as the model of lupus nephritis or Sjogren syndrome. In this study, we examined the right ventricular pressure overload, pathological features and the signaling molecules in the lung tissues of MRL/lpr mice. Methods: We used the female MRL/lpr mice at the age of 12 to 18 weeks. We measured right ventricular systolic pressure (RVSP) by a micro-manometer catheter and the ratio of right ventricular weight to left ventricular weight including ventricular septum (RV/LV). Lung sections were stained with α-smooth muscle actin, and the medial smooth muscle area was expressed as a percentage of the external area of the vessel. Levels of endothelial nitric oxide synthase (eNOS), survivin, endothelin-1 (ET-1), and activin receptor-like kinase-1 (ALK-1) were analyzed by Western blotting. Results: RVSP in the MRL/lpr mice was significantly higher (29.8 ± 7.2 vs. 19.2 ± 1.9 mmHg, P < 0.05) than that in WT C57/Bl6 mice. RV/LV was increased in the MRL/lpr mice compared to the WT C57/Bl6 mice (0.33 ± 0.58 vs. 0.20 ± 0.67, P < 0.05). The medial smooth muscle area of MRL/lpr mice were larger than that of WT C57/bl6 (56.3 ± 4.0 vs. 39.5 ± 6.1%, P < 0.05). Western blot analysis revealed the markedly elevated levels of ET-1 and survivin as well as decreased ALK-1 expression and eNOS phosphorylation in the lung tissue of MRL/lpr mice. Conclusion: We demonstrated that MRL/lpr mice were complicated with pulmonary hypertension. The possible mechanisms include the vasoconstriction enhanced by increased ET-1 level and impaired vasodilatation caused by decreased eNOS phosphorylation and ALK-1 expression. In addition, upregulation of survivin promoted the medial wall thickening of pulmonary vessels. MRL/lpr mice appeared to be a useful model in considering the mechanism of pulmonary hypertension associated with connective tissue diseases.

Calpain mediated pulmonary vascular remodeling in hypoxia induced pulmonary hypertension

To explore the role of calpain in pulmonary vascular remodeling in hypoxia-induced pulmonary hypertension and the underlying mechanisms. Sprague-Dawley rats were randomly divided into the hypoxia group and the normoxia control group. Right ventricular systolic pressure (RVSP) and mean pulmonary artery pressure (mPAP) were monitored by a method with right external jugular vein cannula. Right ventricular hypertrophy index was presented as the ratio of right ventricular weight to left ventricular weight (left ventricle plus septum weight). Levels of calpain-1, -2 and -4 mRNA in pulmonary artery were determined by real-time PCR. Levels of calpain-1, -2 and -4 protein were determined by Western blot. Primary rat pulmonary arterial smooth muscle cells (PASMCs) were divided into 4 groups: a normoxia control group, a normoxia+MDL28170 group, a hypoxia group and a hypoxia+MDL28170 group. Cell proliferation was detected by MTS and flow cytometry. Levels of Ki-67 and proliferating cell nuclear antigen (PCNA) mRNA were determined by real-time PCR. RVSP, mPAP and right ventricular remodeling index were significantly elevated in the hypoxia group compared to those in the normoxia group. In the hypoxia group, pulmonary vascular remodeling was significantly developed, accompanied by up-regulation of calpain-1, -2 and -4. MDL28170 significantly inhibited hypoxia-induced proliferation of PASMCs concomitant with the suppression of Ki-67 and PCNA mRNA expression. Calpain mediates vascular remodeling via promoting proliferation of PASMCs in hypoxia-induced pulmonary hypertension.

Protective effects of drag-reducing polymers in a rat model of monocrotaline-induced pulmonary hypertension.

Drag-reducing polymers (DRPs) are blood-soluble macromolecules which may increase blood flow and reduce vascular resistance. The purpose of the present study was to observe the effect of DRPs on monocrotaline-induced pulmonary hypertension (PH) in the rat model. A total of 64 male Wistar rats were randomly divided into four groups: GroupI (pulmonary hypertension model+ DRP treatment); GroupII (pulmonary hypertension model + saline treatment); GroupIII (control + DRP treatment); GroupIV (control + saline treatment). After five weeks, comparisons were made of the following indices: survival rate, body weight, blood pressure, right ventricular systolic pressure, right ventricular hypertrophy, wall thickness of pulmonary arteries, the internal diameter of small pulmonary arteries, plasma IL-1β and IL-6. The survival rate after 5 weeks varied significantly across all groups (P=0.013), but the survival rates of Groups I and II were not statistically significantly different. Administration of DRP (intravenous injection twice weekly) attenuated the PH-induced increase in right ventricular systolic pressure and suppressed the increases in right ventricular (RV) weight and the ratio of right ventricular weight to left ventricle plus septum weight (RV/LV + S). DRP treatment also significantly decreased the wall thickness of pulmonary arteries, augmented the internal diameter of small pulmonary arteries, and suppressed increases in the plasma levels of IL-1β and IL-6. DRP treatment with intravenous injection effectively inhibited the development of monocrotaline-induced pulmonary hypertension in the rat model. DRPs may have potential application for the treatment of pulmonary hypertension.

Effect of thoracic epidural blockade on hypoxia-induced pulmonary arterial hypertension in rats.

The present study was aimed to investigate the influence of thoracic epidural blockade on hypoxia-induced pulmonary hypertension in rats. Forty eight Wistar rats were randomly divided into 4 equal groups, named normoxia hypoxia hypoxia/ ropivacaine and hypoxia/saline. Animals were placed in a hypoxia chamber and instrumented with epidural catheters at the thoracic level. Rats were injected with saline or ropivacaine. Haemodynamic measurements included pulmonary artery pressure and right ventricular hypertrophy. Degree of pulmonary vascular remodeling was determined by Hematoxylin and Eosin (HE) staining. Serum cyclic GMP (cGMP) and TNF-α were measured using radioimmuno assay. Real-time PCR and western boltting were employed to examine the expression of cAMP responding-element binding protein (CREB). We found that the thoracic epidural blockade significantly decreased chronic hypoxia-induced pulmonary hypertension and vascular remodeling in rats. Ropivacaine-treated rats exhibited significantly lower mean pulmonary artery pressure (mPAP), ratio of right ventricular weight to left ventricular plus septal weight (RV/(LV+S)) and wall thickness of pulmonary artery compared with those of control rats. Hypoxia-induced increase in levels of serum cGMP and TNF-α was reversed by thoracic epidural blockade. Moreover, hypoxia increased expression of CREB at mRNA and protein levels which could be suppressed by thoracic epidural blockade. Thoracic epidural blockade reduced mPAP and serum level of TNF-α and increased cGMP. The treatment reversed upregulated expression of CREB at mRNA and protein production.

Gene Transfer Therapy by Either Type 1 or Type 2 Adeno-Associated Virus Expressing Human Prostaglandin I2 Synthase Gene is Effective for Treatment of Pulmonary Arterial Hypertension

Prostaglandin I(2) (PGI(2)) plays an important role in the clinical treatment of pulmonary arterial hypertension (PAH). However, the administration of PGI(2) involves continuous intravenous infusion using an indwelling catheter, which limits the patient's quality of life and increases the risk of infection. We therefore investigated whether human PGI(2) synthase (hPGIS) gene transfer using an adeno-associated virus (AAV) vector is still effective in a mouse model of PAH and tested for differences in the therapeutic efficacy of PAH among AAV serotypes. The PAH was induced by subjecting mice to hypoxia (10% O(2)). Type 1 AAV expressing hPGIS (AAV1-hPGIS) or type 2 AAV expressing hPGIS (AAV2-hPGIS) was injected into the thigh muscle of mice. Both vectors expressing hPGIS produced strong hPGIS protein expression in the mouse thigh skeletal muscles after 8 weeks of hypoxia. The administration of AAV1-hPGIS or AAV2-hPGIS also significantly inhibited the hypoxia-induced increase in right ventricular systolic pressure, the ratio of right ventricular weight to body weight (RV/BW), and the ratio of RV weight to left ventricular plus septal weight (RV/LV + S), and significantly attenuated the hypoxia-induced increase in medial wall thickness of peripheral pulmonary arteries. Furthermore, there were no significant differences in the degree of amelioration in RV systolic pressure, RV/BW, RV/LV + S, and percentage of wall thickness of peripheral pulmonary arteries between AAV1-hPGIS and AAV2-hPGIS administrations. In conclusion, we revealed that type 1 and type 2 AAV are equally effective for the treatment of PAH in a hypoxia-induced mouse model. Gene-transfer therapy using AAV expressing hPGIS is, therefore, a potential therapeutic breakthrough for PAH.

Benzenesulfonamide attenuates monocrotaline-induced pulmonary arterial hypertension in a rat model

In this study, we examined the effects of LASSBio-965 (N-[2-(3,4-dimethoxyphenyl) ethyl]-benzenesulfonamide), a compound designed as a simplified structure of a non-selective phosphodiesterase 4 inhibitor, on vascular smooth muscle in vitro as well as in a rat model of monocrotaline (MCT)-induced pulmonary arterial hypertension. LASSBio-965 (50mg/kg) treatment caused a significant decrease in right systolic ventricular pressure (32.47±3.09mmHg) compared to the MCT-vehicle group (51.88±3.23mmHg; P<0.05) and in the ratio of right ventricular weight to left ventricular weight plus septum (0.42±0.03g compared to 0.59±0.06g, respectively; MCT-vehicle group; P<0.05). LASSBio-965 induced a concentration-dependent relaxation of rat aortic rings, which was decreased by mechanical removal of the endothelium. Milrinone, rolipram, and sildenafil reduced the maximum relaxation (100%) to 22.4±5.8, 69.5±5.6 and 80.1±10.7%, respectively (P<0.05). Maximum relaxation responses of aortic and pulmonary artery rings were decreased in the MCT-vehicle group (54.80±5.69 and 35.87±4.78, respectively) compared to the control (91.51±4.79 and 54.32±2.39, respectively) but improved with LASSBio-965 treatment (50mg/kg; 88.43±4.54 and 59.36±4.83, respectively). These results indicate that LASSBio-965 can attenuate the pulmonary arterial hypertension in an animal model most likely through the nonselective inhibition of phosphodiesterases 3, 4, and 5.

Genetic correlation between heart ratio and body weight as a function of ascites frequency in broilers split up into sex and health status

Ascites or pulmonary hypertension syndrome is a metabolic disorder in broilers. Male broilers have a higher BW and are therefore expected to be more prone to developing ascites than females. As genetic parameters might be affected by the ascites incidence, genetic parameters might differ between male and female broilers. The aims of this study were to estimate the heritability for the ratio of right ventricular weight to total ventricular weight (RATIO) and BW in male and female broilers, the genetic correlation between RATIO and BW separately for male and female broilers, and the genetic correlations between BW for ascitic and nonascitic broilers. Data were available from 7,856 broilers (3,819 males and 4,037 females). The broilers in the experiment were kept under a cold temperature regimen and increased CO2 levels. In this study, we showed that the incidence of ascites is higher in male than in female broilers. Heritability estimates for BW at 7 wk of age were higher for male (0.22) than for female (0.17) broilers, and for RATIO heritability, estimates were higher for female (0.44) than for male (0.32) broilers. The genetic correlations between RATIO and BW measured at different ages changed from slightly positive at 2 wk of age to moderately negative at 7 wk. The change in genetic correlation was more extreme for male (from 0.01 to −0.62) than for female (from 0.13 to −0.24) broilers. The difference in ascites incidence between male and female broilers is the most likely reason for the difference in genetic correlations. The genetic correlation between BW traits measured in broilers with fluid in the abdomen and without fluid in the abdomen decreased from 0.91 at 2 wk to 0.69 at 7 wk. We conclude that under circumstances with ascites, data from male and female broilers should be analyzed separately.

Effects of Autologous Bone Marrow Mononuclear Cells Implantation in Canine Model of Pulmonary Hypertension

We investigated the safety and feasibility of intratracheal administration of autologous bone marrow-derived mononuclear cells (ABM-MNCs) and observed the effects in a canine model of pulmonary hypertension (PH). The PH model was induced by intravenous injection of 3mg/kg dehydromonocrotaline (DMCT) via the right atrium. Two weeks after DMCT administration, the animals received 4 different treatments (n=10 in each group): (I) negative control group; (II): ABM-MNCs group; (III) PH group; (IV) PH+ABM-MNCs group. Six weeks after injection of cells (10⁷), the hemodynamic data were significantly improved in group IV compared with group III (P<0.05). The ratio of right ventricular weight to left ventricular plus septal weight was significantly decreased in group IV compared with group III (P<0.05). The mRNA levels of vascular endothelial growth factor, preproendothelin-1, interleukin-6 and tumor necrosis factor-α were significantly improved in group IV compared with group III (P<0.05). The immunofluorescence result showed that 6 weeks after administration ABM-MNCs could differentiate into pulmonary vascular endothelial cells. Six weeks after intratracheal administration, ABM-MNCs significantly improved the impairment caused by DMCT in a canine model of PH (ie, decreased pulmonary arteriolar narrowing, alveolar septum thickening and right ventricular hypertrophy, enhanced angiogenesis) and this provides a firm foundation for a clinical trial.

Effects of prostaglandin E1 (PGE1) on pulmonary hypertension and lung vascular remodeling in a rat monocrotaline model of human pulmonary hypertension

Prostaglandin E1 (PGE1) has been shown to be a potent pulmonary vasodilator in humans and in many animals. The effects of PGE1 on the development of pulmonary hypertension and on pulmonary vascular remodeling were studied in a rat monocrotaline (MCT) model of human pulmonary hypertension. By 3 weeks after injection, MCT (80 mg/kg S.C.) had resulted in high values of mean pulmonary arterial pressure and of the ratio of right ventricular weight to left ventricle+septum weight (RV/LV+S). PGE1 inhibited the development of pulmonary hypertension (300 micrograms/kg) and right ventricular hypertrophy (300 and 100 micrograms/kg) induced by MCT. Three weeks after the injection, the media walls of pulmonary arteries in lungs from rats given MCT were significantly thicker than those from lungs of control rats. PGE1 (300, 100, and 30 micrograms/kg) resulted in significantly less of this morphologic change, in a dose-dependent manner. These results indicate that PGE1 inhibits the development of pulmonary hypertension associated with lung vascular thickening induced by MCT. PGE1 may be useful for the treatment of pulmonary hypertension in humans.

Role of leukotriene B4 in monocrotaline-induced pulmonary hypertension

Monocrotaline (MCT) causes lung inflammation and chronic pulmonary hypertension associated with lung vascular thickening in rats. We hypothesized that leukotriene B4 (LTB4) and LTB4-induced accumulation of leukocytes in the lung play a role in MCT-induced lung disease, and therefore measured LTB4 and myeloperoxidase (MPO) levels in lung tissue of MCT-treated rats. Next, we examined the effect of an orally active LTB4 receptor antagonist (ONO4057) on MPO levels in lung tissue, on pulmonary hypertension, and on pulmonary vascular remodeling induced by MCT. Lung LTB4 and MPO levels had increased by 3 days after MCT injection. In the ONO4057-treated MCT rats, lung MPO levels were significantly lower than in the rats given MCT but not ONO4057. By the third week after injection. MCT had caused increases in mean pulmonary arterial pressure, in the ratio of right ventricular weight to left ventricle+septum weight (RV/[VS + S]), and in media wall thickness of the muscular arteries of the lung. Treatment with ONO4057, either for 3 weeks or during the first week after MCT injection, significantly reduced pulmonary hypertension, right ventricular hypertrophy, and lung vascular thickening induced by MCT. These results indicate that ONO4057 reduces both the accumulation of leukocytes in lung tissue and the chronic pulmonary hypertension induced by MCT, and they suggest a role for LTB4 in the inflammatory process that contributes to pulmonary hypertension and lung vascular remodeling induced by MCT in rats.

Generation of hydroxyl radicals during ascites experimentally induced in broilers

1. Increased metabolic rates, pulmonary hypertension and cardiac dysfunction are the most important features of the ascites syndrome in broiler chickens. However, the mechanism of cell injury causing the pathogenesis of the syndrome is not clearly understood. Our study aimed to examine the generation of hydroxyl radicals (OH•) in broiler chickens experiencing ascites. 2. Three hundred and fifty 1-d-old chickens were purchased from a local hatchery and reared in an open poultry house for 46 d. They were divided at random into three groups and ascites was induced in two groups by exposing them to low temperature or administration of triiodothyronine (T3). The third group served as control and was reared normally. 3. Haematological, biochemical and pathological tests were used to determine the incidence of ascites; including total red blood cell (RBC), packed cell volume (PCV), release of alanine transaminase (ALT) and aspartate transaminase (AST) and ratio of right ventricular weight to total ventricular weight (RV/TV). 4. A salicylate hydroxylation method was used to examine the generation of hydroxyl radicals (OH•) in treated groups. Two hydroxylated salicylic acid metabolites, 2,3- and 2,5-dihydroxy benzoic acid (2,3- and 2,5-DHBA), were measured by HPLC to detect the generation of OH•. 5. An ascites syndrome was observed in T3 and low-temperature treated groups, as shown by necropsy changes and increases in f RBC, PCV, ALT, AST and the ratio of RV/TV. Concentrations of 2,3- and 2,5-DHBA were increased in groups experiencing ascites compared to control group. 6. It is suggested that reactive oxygen species, that is OH• ions, may be involved in the pathogenesis of the ascites syndrome in broiler chickens.

A Bivariate Mixture Model Analysis of Body Weight and Ascites Traits in Broilers

The objective of the present study was to use bivariate mixture models to study the relationships between body weight (BW) and ascites indicator traits. Existing data were used from an experiment in which birds were housed in 2 groups under different climate conditions. In the first group, BW, the ratio of right ventricular weight to total ventricular weight (RV:TV), and hematocrit value (HCT) were measured in 4,202 broilers under cold conditions; in the second group, the same traits were measured in 795 birds under normal temperature conditions. Cold-stress conditions were applied to identify individuals that were susceptible to ascites. The RV:TV and HCT were approximately normally distributed under normal temperature conditions, whereas the distributions of these traits were skewed under cold temperature conditions, suggesting different underlying distributions. Fitting a bivariate mixture model to the observations showed that there was only one homogeneous population for ascites traits under normal temperature conditions, whereas there was a mixture of (2) distributions under cold conditions. One distribution contained nonascitic birds and the other distribution contained ascitic birds. In the distribution of nonascitic birds, the inferred phenotypic correlations (phenotypic correlations with 2 distinguishing underlying distributions) of BW with RV:TV and HCT were close to zero (0.10 and -0.07, respectively), whereas in the distribution of ascitic birds, the inferred phenotypic correlations of BW with RV:TV and HCT were negative (-0.39 and -0.4, respectively). The negative inferred correlations of BW with RV:TV and HCT in the distribution of ascitic birds resulted in negative overall correlations (correlations without 2 distinguishing distributions) of BW with RV:TV (-0.30) and HCT (-0.37) under cold conditions. The present results indicate that the overall correlations between BW and ascites traits are dependent on the relative frequency of ascitic and nonascitic birds in the population.

A Long-Acting Prostacyclin Agonist with Thromboxane Inhibitory Activity for Pulmonary Hypertension

The balance between prostacyclin and thromboxane plays an important role in the regulation of pulmonary vascular tone. Recently, we developed ONO-1301, a novel, long-acting prostacyclin agonist with thromboxane synthase inhibitory activity. We investigated whether modulation of prostacyclin/thromboxane balance by ONO-1301 ameliorates monocrotaline-induced pulmonary hypertension in rats. After subcutaneous injection of monocrotaline or vehicle, rats were randomized to receive repeated subcutaneous administration of ONO-1301 or vehicle twice per day for 3 wk. There was significant development of pulmonary hypertension 3 wk after monocrotaline injection. Treatment with ONO-1301 significantly attenuated the increases in right ventricular systolic pressure and ratio of right ventricular weight to body weight in monocrotaline rats. Furthermore, ONO-1301 significantly attenuated the increase in medial wall thickness of peripheral pulmonary arteries in monocrotaline rats. The half-life of plasma ONO-1301 concentration after a single subcutaneous administration was approximately 5.6 h. A single administration of ONO-1301 increased plasma cyclic adenosine 3', 5'-monophosphate level, which lasted at least up to 8 h. Treatment with ONO-1301 significantly decreased plasma 11-dehydro-thromboxane B2, a metabolite of thromboxane, in monocrotaline rats. Finally, Kaplan-Meier survival curves demonstrated that repeated administration of ONO-1301 improved survival rate in monocrotaline rats compared with vehicle administration (80 vs. 30% in 6-wk survival). Subcutaneous administration of a novel prostacyclin agonist (ONO-1301) markedly attenuated monocrotaline-induced pulmonary hypertension and improved survival in rats. The beneficial effects of ONO-1301 may occur through its long-lasting stimulation of cyclic adenosine 3', 5'-monophosphate and inhibition of thromboxane synthase.

Cyclin-Dependent Kinase Inhibitor p27 Kip1 , But Not p21 WAF1/Cip1 , Is Required for Inhibition of Hypoxia-Induced Pulmonary Hypertension and Remodeling by Heparin in Mice

Heparin has growth inhibitory effects on pulmonary artery smooth muscle cell (PASMC) in vitro and in vivo. However, the mechanism has not been fully defined. In this study, we investigated the role of cyclin-dependent kinase inhibitors, p21(WAF1/cip1) (p21) and p27Kip1 (p27), in the inhibitory effect of heparin on PASMC proliferation in vitro and on hypoxia-induced pulmonary hypertension in vivo using p21 and p27-null mice. In vitro, loss of the p27 gene negated the inhibitory effect of heparin on PASMC proliferation, but p21 was not critical for this inhibition. In vivo, heparin significantly inhibited the development of hypoxia-induced pulmonary hypertension and remodeling, as evidenced by decreased right ventricular systolic pressure, ratio of right ventricular weight to left ventricle plus septum weight, and percent wall thickness of pulmonary artery, in p21(+/+), p21(-/-), p27(+/+), and p27(+/-), but not in p27(-/-) mice. We also observed that hypoxia decreased p27 expression significantly in mouse lung, which was restored by heparin. Heparin inhibited Ki67 proliferative index in terminal bronchial vessel walls in p27(+/+) and p27(+/-), but not in p27(-/-) mice exposed to hypoxia. Therefore, we conclude that the cyclin-dependent kinase inhibitor p27, but not p21, is required for the inhibition of hypoxic pulmonary vascular remodeling by heparin.

Genetic mapping of quantitative trait loci affecting susceptibility in chicken to develop pulmonary hypertension syndrome

Pulmonary hypertension syndrome (PHS), also referred to as ascites syndrome, is a growth-related disorder of chickens frequently observed in fast-growing broilers with insufficient pulmonary vascular capacity at low temperature and/or at high altitude. A cross between two genetically different broiler dam lines that originated from the White Plymouth Rock breed was used to produce a three-generation population. This population was used for the detection and localization of quantitative trait loci (QTL) affecting PHS-related traits. Ten full-sib families consisting of 456 G2 birds were typed with 420 microsatellite markers covering 24 autosomal chromosomes. Phenotypic observations were collected on 4202 G3 birds and a full-sib across family regression interval mapping approach was used to identify QTL. There was statistical evidence for QTL on chicken chromosome 2 (GGA2), GGA4 and GGA6. Suggestive QTL were found on chromosomes 5, 8, 10, 27 and 28. The most significant QTL were located on GGA2 for right and total ventricular weight as percentage of body weight (%RV and %TV respectively). A related trait, the ratio of right ventricular weight as percentage to total ventricular weight (RATIO), reached the suggestive threshold on this chromosome. All three QTL effects identified on GGA2 had their maximum test statistic in the region flanked by markers MCW0185 and MCW0245 (335-421 cM).

Direct and maternal genetic effects for ascites-related traits in broilers

The objective of the present study was to estimate heritabilities for ascites-related traits in broilers and to assess the importance of maternal genetic effects for these traits. Several traits related to ascites were measured on more than 4,000 broilers kept under cold conditions. Heritabilities were estimated using an animal model with a direct genetic effect and a model with direct and maternal genetic effects. Estimated heritabilities from the direct genetic effects model were 0.46 for hematocrit value, 0.42 for BW, 0.47 for right ventricular weight, 0.46 for total ventricular weight, 0.45 for ratio of right ventricular weight to the total ventricular weight, 0.32 for total mortality, and 0.18 for fluid accumulation in the heart sac. Maternal effects significantly influenced the traits BW, total ventricular weight, and total mortality. Direct and maternal heritabilities, respectively, for BW were 0.21 and 0.04, for total ventricular weights were 0.29 and 0.03, and for total mortality were 0.16 and 0.05. The heritability estimates for ascites-related traits and the significance of maternal genetic effects for most of these traits indicate that direct and maternal genetic effects play an important role in the development of the ascites syndrome.