Pulmonary Hypertension In Bronchopulmonary Dysplasia Research Articles

Navigating Diagnostic and Treatment Challenges of Pulmonary Hypertension in Infants with Bronchopulmonary Dysplasia.

Advances in perinatal intensive care have significantly enhanced the survival rates of extremely low gestation-al-age neonates but with continued high rates of bronchopulmonary dysplasia (BPD). Nevertheless, as the survival of these infants improves, there is a growing awareness of associated abnormalities in pulmonary vascular development and hemodynamics within the pulmonary circulation. Premature infants, now born as early as 22 weeks, face heightened risks of adverse development in both pulmonary arterial and venous systems. This risk is compounded by parenchymal and airway abnormalities, as well as factors such as inflammation, fibrosis, and adverse growth trajectory. The presence of pulmonary hypertension in bronchopulmonary dysplasia (BPD-PH) has been linked to an increased mortality and substantial morbidities, including a greater susceptibility to later neurodevelopmental challenges. BPD-PH is now recognized to be a spectrum of disease, with a multifactorial pathophysiology. This review discusses the challenges associated with the identification and management of BPD-PH, both of which are important in minimizing further disease progression and improving cardiopulmonary morbidity in the BPD infant.

Factors in the Neonatal Period Associated With Pulmonary Hypertension at 28 Days of Life in Broncho-Pulmonary Dysplasia.

Objectives. To identify factors associated with pulmonary hypertension (PH) at 28 days of life in preterm infants with bronchopulmonary dysplasia (BPD). Methods. This observational study included 128 premature infants with BPD between January 2022 and February 2023 from the neonatal intensive care unit of Vietnam National Children's Hospital. Results. PH was observed using echocardiography in 29 patients (22.66%). The prevalence of severe BPD in the PH group (62.07%) was significantly higher than that in the non-PH group (18.18%). The multivariate logistic regression showed 2 predictors of PH in BPD: invasive mechanical ventilation up to 28 days of life (odds ratio [OR]:9.440; 95% confidence interval [CI]: 3.090-28.833; P < .001) and history of shock (OR: 2.962; 95% CI: 1.067-8.225; P = .037). Conclusion. We found 2 predictors of PH at 28 days of life in BPD: invasive mechanical ventilation up to 28 days of life and history of shock.

Current diagnosis and treatment practice for pulmonary hypertension in bronchopulmonary dysplasia-A survey study in Germany (PUsH BPD).

Pulmonary hypertension (PH) is the most severe complication in preterm infants with bronchopulmonary dysplasia (BPD) and associated with significant mortality. Diagnostic and treatment strategies, however, still lack standardization. By the use of a survey study (PH in BPD), we assessed clinical practice (diagnosis, treatment, follow-up) in preterm infants with early postnatal persistent pulmonary hypertension of the newborn(PPHN) as well as at risk for or with established BPD-associated PH between 06/2018 and 10/2020 in two-thirds of all German perinatal centers with >70 very low birthweight infants/year including their cardiology departments and outpatient units. Data wereanalyzed descriptively by measures of locations and distributional shares. In routine postnatal care, clinical presentation and echocardiography were reported as the main diagnostic modalities to screen for PPHN in preterm infants, whereas biomarkers brain natriuretic peptide/N-terminal pro b-type natriuretic peptide were infrequently used. For PPHN treatment, inhaled nitric oxidewas used in varying frequency. The majority of participants agreed to prescribe diuretics and steroids (systemic/inhaled) for infants at risk for or with established BPD-associated PH and strongly agreed on recommending respiratory syncytial virus immunization and the use of home monitoring upon discharge. Reported oxygen saturation targets, however, varied in these patients in in- and outpatient care. The survey reveals shared practices in diagnostic and therapeutic strategies for preterms with PPHN and BPD-associated PH in Germany. Future studies are needed to agree on detailed echo parameters and biomarkers to diagnose and monitor disease next to a much-needed agreement on the use of pulmonary vasodilators, steroids, and diuretics as well as target oxygen saturation levels.

Bronchopulmonary dysplasia – a new perspective on prevention and treatment

Introduction &#x0D; Premature birth is one of the most common problems in neonatal care and affects from 6 to 14% of pregnancies (frequency depends on geographical factors) [1]. Prematurity is the cause of almost half of deaths of children younger than 5 years old [3]. The most frequent and one of the most severe risks of preterm births is bronchopulmonary dysplasia (also called a chronic lung disease in some sources). &#x0D; Aim of the study &#x0D; The aim of this study is to collect data on both new and traditional ways of prevention and treatment of chronic lung disease in a population of infants born preterm (&lt;37 weeks of gestational age). &#x0D; Materials and methods &#x0D; The work was created based on the articles available in PubMed and Google Scholar databases and available scientific literature. Publications were searched in English and Polish using the following keywords: bronchopulmonary dysplasia, chronic lung disease of the newborns, pulmonary hypertension in bronchopulmonary dysplasia, stem cells in bronchopulmonary dysplasia. &#x0D; Results&#x0D; Although bronchopulmonary dysplasia is a common problem among children born preterm, there is still a limited amount of possibilities of prevention and treatment of this condition. Despite efforts of scientists, many children with this condition suffer from both short-term and long-term consequences, such as frequent infections, asthma and impaired growth and neurodevelopment. &#x0D; Summary &#x0D; In the last twenty years the survival rate of children born preterm rapidly increased thank to the development of neonatal care. Up to 20% of children born with body weight less than 1500g will develop bronchopulmonary dysplasia. That is why there is a need for further research on the topic of bronchopulmonary dysplasia management, especially when it comes to biomarkers, use of mesenchymal stem cells and treatment of pulmonary hypertension. &#x0D;

The role of rhIGF-1/BP3 in the prevention of pulmonary hypertension in bronchopulmonary dysplasia and its underlying mechanism

BackgroundThis study aimed to determine whether postnatal treatment with recombinant human IGF-1 (rhIGF-1)/binding peptide 3 (BP3) ameliorates lung injury and prevents pulmonary hypertension (PH) in bronchopulmonary dysplasia (BPD) models.MethodsWe used two models of BPD in this study: one model that was associated with chorioamnionitis (CA), stimulated by intra-amniotic fluid and exposure to lipopolysaccharide (LPS), whereas the other was exposed to postnatal hyperoxia. Newborn rats were treated with rhIGF-1/BP3 (0.2 mg/Kg/d) or saline via intraperitoneal injection. The study endpoints included the wet/dry weight (W/D) ratio of lung tissues, radial alveolar counts (RACs), vessel density, right ventricular hypertrophy (RVH), lung resistance, and lung compliance. Hematoxylin and eosin (H&E) and Masson staining were used to evaluate the degree of lung injury and pulmonary fibrosis. IGF-1 and eNOS expression were detected using western blotting or quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). The levels of SP-C, E-cadherin, N-cadherin, FSP1, and Vimentin in the lung tissues were detected by immunofluorescence.ResultsLPS and hyperoxia treatment increased lung injury and pulmonary fibrosis, enhanced RVH and total respiratory resistance, and decreased RAC, pulmonary vascular density and pulmonary compliance in young mice (all p < 0.01). Simultaneously, LPS and hyperoxia induced an increase in epithelial-mesenchymal transition (EMT) in airway epithelial cells. However, rhIGF-1/BP3 treatment reduced lung injury and pulmonary fibrosis, decreased RVH and total respiratory resistance, and enhanced RAC, pulmonary vascular density and pulmonary compliance, as well as inhibited EMT in airway epithelial cells in LPS and hyperoxia treated mice.ConclusionPostnatal rhIGF-1/BP3 treatment relieved the effects of LPS or hyperoxia on lung injury and prevented RVH, providing a promising strategy for the treatment of BPD.

Recovery of hyperoxia‐induced DDAH1 suppression with FXR agonist GW4064 in human pulmonary microvascular endothelial cells

ObjectivePulmonary hypertension (PH) predicts mortality in patients with bronchopulmonary dysplasia (BPD), and chronic hyperoxia exposure contributes to BPD‐PH disease progression. BPD‐PH is characterized by aberrant pulmonary vascular wall proliferation/remodeling, which occludes the microvascular lumen, increasing pulmonary vascular resistance. Endogenous nitric oxide (NO) production by pulmonary vascular endothelial cells promotes apoptosis of pulmonary vascular endothelial and smooth muscle cells. In vitro studies of human pulmonary microvascular endothelial cells (hPMVEC) have shown that dimethylarginine dimethylaminohydrolase‐1 (DDAH1) promotes NO‐mediated apoptosis. DDAH1 metabolizes the majority of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of endothelial NO synthase. Therefore, our hypothesis is that pharmacologically enhancing DDAH1 expression in hPMVEC will restore endogenous endothelial NO production.MethodshPMVEC were grown at 37 ̊C to ~70% confluence. Farnesoid X receptor (FXR) agonist, GW4064 (1.25 µM) was used to bind FXR element (FXRE) of the DDAH1 promoter to enhance transcription. hPMVEC were pre‐treated with FXR agonist for 24 hours and then exposed to 85%O2 or 21%O2 for an additional 24 hours. DDAH1 protein expression was analyzed by Western blot. Groups were compared by 1‐way ANOVA.ResultsIn hPMVEC, hyperoxia exposure resulted in lower DDAH1 protein levels compared to hPMVEC grown in normoxia (N=6 in each group, p=0.003). Treatment with GW4064 prior to exposing to hyperoxia resulted in 4.3‐fold greater DDAH1 protein levels than in vehicle/85%O2 hPMVEC (N=6, p=0.044), and restored DDAH1 protein levels to 72% of that seen in the vehicle/21%O2exposed hPMVEC.ConclusionsChronic hyperoxia suppresses DDAH1 levels, and FXR agonist promotes DDAH1 recovery in hPMVEC. We speculate that promoting endogenous DDAH1 expression may prevent aberrant proliferation in the pulmonary vascular wall and thereby attenuate or prevent the vascular remodeling characteristic of pulmonary hypertension in BPD.

DDAH1 SNP rs480414 that protects against the development of pulmonary hypertension in bronchopulmonary dysplasia results in lower nitric oxide production in neonatal cord blood-derived lymphoblastoid cell lines.

Bronchopulmonary dysplasia (BPD) is chronic lung disease of prematurity and pulmonary hypertension (PH) is a major contributor to morbidity and mortality in BPD patients. Nitric oxide (NO) is a vasodilator and apoptotic mediator made by NO synthase (NOS). NOS is inhibited by asymmetric dimethylarginine (ADMA), and dimethylarginine dimethylaminohydrolase (DDAH) hydrolyzes ADMA. Previously, in a BPD patient cohort, we identified single nucleotide polymorphism (SNP) DDAH1 rs480414 (G > A) that was protective against developing PH. This study aims to determine functional consequences of the DDAH1 SNP in lymphoblastoid cell lines (LCLs) derived from neonatal cord blood. We tested the hypothesis that DDAH1 SNP (AA) results in DDAH1 gain of function, leading to greater NO-mediated apoptosis compared to DDAH1 wild-type (GG) in LCLs. LCLs were analyzed by Western blot (DDAH1, cleaved and total caspase-3 and -8, and β-actin), and RT-PCR (DDAH1, iNOS). Cell media assayed for nitrites with chemiluminescence NO analyzer, and conversion of ADMA to L-citrulline was measured by spectrophotometry. LCLs with DDAH1 SNP had similar levels of DDAH1 protein and mRNA expression, as well as DDAH activity, compared to DDAH1 WT LCLs. There were also no changes in cleaved caspase-3 and -8 protein levels. LCLs with DDAH1 SNP had similar iNOS mRNA expression. Nitrite levels in media were lower for DDAH1 SNP LCLs compared to DDAH1 WT LCLs (p < 0.05). Contrary to our hypothesis, we found that NO production was lower in DDAH1 SNP LCLs, indicative of a loss of function phenotype.

Current and Future Applications of Thoracic Dual-Energy CT in Children: Pearls and Pitfalls of Technique and Interpretation

Dual-energy computer tomography (DECT) technology has experienced rapid growth in recent years, now allowing for the collection of 2 CT data sets and opening the potential for functional data acquisition. Data from a single postcontrast phase are deconstructed and Iodine can be subtracted to create a virtual noncontrast image, or selectively represented as a contrast map that allows for the qualification and quantification of lung perfusion. Virtual monoenergetic images can also be used to reduce beam-hardening artifact from concentrated contrast or metal implants. In children, DECT is of particular interest because it has been shown to be dose neutral in most applications, dose-reducing in multiphase studies, and to increase the contrast to noise ratio in suboptimal studies. We review the basics of acquisition, postprocessing, and thoracic applications of DECT with a focus on pulmonary blood volumes as a surrogate for perfusion imaging. The discussed applications include pulmonary embolism, hypoplastic lung, pulmonary hypertension in bronchopulmonary dysplasia, and pediatric lung masses.

DDAH1 Overexpression in Human Pulmonary Vascular Endothelial Cells Co‐Cultured with Human Pulmonary Artery Smooth Muscle Cells

Bronchopulmonary dysplasia (BPD) is the most common chronic lung disease of preterm infants. Pulmonary hypertension (PH) develops in 25–40% of BPD patients as a result of pathologic vascular remodeling, and PH is an independent predictor of death in BPD patients. Nitric oxide (NO) is a potent vasodilator and apoptotic mediator made in the pulmonary endothelium by nitric oxide synthase (eNOS). Asymmetric dimethylarginine (ADMA) is an endogenous competitive NOS inhibitor. Dimethylarginine dimethylaminohydrolase (DDAH) metabolizes the majority of ADMA in vivo and there are two isoforms, DDAH1 and DDAH2. Translational studies from our group analyzed a cohort of BPD patients and identified a potential biomarker, DDAH1 single nucleotide polymorphism (rs480414), that decreased the risk of PH in BPD by 61%. In the present study, we hypothesized that overexpression of DDAH1 in human pulmonary microvascular endothelial cells (hPMVEC) would lead to greater apoptosis and lower proliferation of human pulmonary artery smooth muscle cells (hPASMC) via effects in hPMVEC. We utilized a co‐culture system where hPMVEC were transfected with AdDDAH1 or AdGFP control for 24h and then incubated in co‐culture with hPASMC for 24h. Analysis included 1) western blot analysis for cleaved and total caspase‐3, ‐8, ‐9, p21, PCNA, and β‐actin and 2) trypan blue exclusion for viable cell numbers. Transfection of hPMVEC with AdDDAH1 resulted in greater DDAH1 protein expression (n=6, p=0.003) as compared to control. After AdDDAH1 transfection of hPMVEC and co‐culture with hPASMC, we found greater cleaved caspase‐3 (n=6, p=0.01), greater cleaved caspase‐8 (n=6, p=0.03), similar cleaved caspase‐9 (n=6, p=0.94), similar p21 (n=6, p=0.7), and similar PCNA (n=3, p=0.82) protein expression as compared to control. After AdDDAH1 transfection of hPMVEC and co‐culture with hPASMC, we found lower hPASMC viable cell numbers (n=3, p=0.008) as compared to control. After AdDDAH1 transfection of hPMVEC and co‐culture with hPASMC treated with DETA NONOate 250μM or DETA NONOate 250μM + Hemoglobin 100μM, the hPASMC viable cell numbers were lower compared to control (n=3, p=0.001; n=3, p=0.003, respectively). There was no difference detected between DETA NONOate 250μM and DETA NONOate 250μM + Hemoglobin 100μM groups (n=3, p=0.429). In conclusion, DDAH1 overexpression in hPMVEC resulted in greater apoptosis and lower proliferation in hPASMC, consistent with our hypothesis. We propose that DDAH1 is a potential new therapy for BPD‐PH via relieving pulmonary vascular endothelial ADMA‐inhibition of eNOS, resulting in greater endogenous NO production, greater pulmonary vascular wall apoptosis, and potentially less pathologic vascular remodeling. Future studies will determine if DDAH1 endothelial knockout mice have worse lung function as compared to wild‐type, after exposure to BPD‐PH model of LPS and hyperoxia.Support or Funding InformationK08 HL129080 (PI: Trittmann)

Bronchopulmonary dysplasia-associated pulmonary hypertension

Chronic respiratory sequelae are a major health problem in surviving premature infants. Despite immense improvement in the care of premature infants, the rate of bronchopulmonary dysplasia (BPD) and pulmonary morbidities have remained the same or even slightly increased over the past decades. BPD results from the impairment of lung development following premature birth, and the resultant dysregulated pulmonary vasculature leads to chronic pulmonary hypertension (PH). It is seen more commonly in the setting of moderate to severe BPD. A high index of suspicion is necessary to diagnose PH in BPD. Diagnosis is most commonly done by echocardiogram, but cardiac catheterization may be required to delineate the cardiac structure, cardio-pulmonary vasculature and myocardial function. Once the diagnosis is made, a multidisciplinary approach for management of these patients is suggested, aiming to optimize management of BPD and use of various pulmonary vasodilators. The various therapeutic agents available help reduce pulmonary vascular resistance by acting on the nitric oxide (NO), prostacyclin and/or endothelin pathways. Serial echocardiograms and measurement of biomarkers are used to evaluate the response to therapy. Currently there is not much evidence to support optimal management approaches, but the recent guidelines published by the Pediatric Pulmonary Hypertension Network (PPHNet) provide practical recommendations for screening and management of these infants.

Death or resolution: the "natural history" of pulmonary hypertension in bronchopulmonary dysplasia.

The primary objective was to describe the early "natural history" of pulmonary hypertension (PH) in the premature population. The secondary objective was to describe factors associated with poor outcomes in the premature population with PH at 36 weeks post-menstrual age (PMA). Retrospective chart review of patients followed at our institution from 2000 to 2017 with echocardiographic (ECHO) evidence of PH at 36 weeks PMA, and born ≤ 32 weeks estimated gestational age (GA). Cox regression was used for survival analysis. Sixty-one patients with PH (26.5 ± 1.5 weeks at birth) were included. All PH patients had bronchopulmonary dysplasia (BPD), with 89% considered severe; 38% were small for gestational age. Necrotizing enterocolitis requiring surgery was common (25%). Use of post-natal steroids (HR 11.02, p = 0.01) and increased severity of PH (HR 1.05, p < 0.001) were associated with mortality. Pulmonary vein stenosis (PVS) was documented in 26% of the PH cohort, but not associated with increased mortality. ECHO estimation of pulmonary artery pressure (PAP) was available in 84%. PAP was higher in those who died (sPAP/sBP ratio 1.09 ± 27 vs 0.83 ± 20 %, p = 0.0002). At follow-up (mean 250 ± 186 weeks PMA), 72% of the PH cohort was alive. Most survivors (66%) had resolution of their PH on their most recent ECHO; 31% remained on PH therapy. PH resolved in most survivors in this study population. Mortality in those with BPD-PH was associated with male sex, post-natal steroid use, and increased severity of PH, but not with PVS.

Using clinical and genetic data to predict pulmonary hypertension in bronchopulmonary dysplasia.

Pulmonary hypertension significantly increases morbidity and mortality in infants with bronchopulmonary dysplasia. The frequency of single nucleotide polymorphisms in arginase-1 (ARG1 rs2781666) and dimethylarginine dimethylaminohydrolase-1 (DDAH1 rs480414) genes has been found to differ in a cohort of bronchopulmonary dysplasia patients with pulmonary hypertension (cases) and without pulmonary hypertension (controls). Therefore, we tested the hypothesis that combining these genotypes with phenotypic data would better predict pulmonary hypertension in bronchopulmonary dysplasia patients. Bronchopulmonary dysplasia patients (n=79) born at <35weeks gestation were studied. Pulmonary hypertension was diagnosed by echocardiographic criteria (n=20). ROC curves to predict pulmonary hypertension in bronchopulmonary dysplasia were generated from genotype and/or clinical data. Cases were born at an earlier gestation and weighed less at birth than did controls. ROC curves for rs2781666 had an AUC of 0.61, while rs480414 had an AUC of 0.66. Together, the AUC was 0.70. When clinical data were added to the genetic model, AUC was 0.73. These findings demonstrate that ROC predictive modelling of pulmonary hypertension in bronchopulmonary dysplasia improves with inclusion of both genotypic and phenotypic data. Further refinement of these types of models could facilitate the implementation of precision medicine approaches to pulmonary hypertension in bronchopulmonary dysplasia.

Morphological characterization of pulmonary microvascular disease in bronchopulmonary dysplasia caused by hyperoxia in newborn mice.

Pulmonary microvascular injury is associated with the pathogenesis of bronchopulmonary dysplasia (BPD). To characterize the mechanisms of pulmonary vascular disease resulting from BPD, we studied the ultrastructural changes affecting pulmonary microvasculature. Newborn ICR mice were exposed to 85% hyperoxia or normoxia for 14days, and then normal air replacement conditions for the following 7days. At postnatal day (P)14 and P21, lungs were harvested for ultrastructural examination and assessment of pulmonary hypertension. The ultrastructure of pulmonary microvasculature in the hyperoxia-exposed lungs revealed a collapsed capillary lumen. This was due to the abnormal morphology of endothelial cells (ECs) characterized by heterogeneously thick cytoplasm. Compared to normal air controls, the specimens displayed also remarkably thick blood-air barriers (BABs), most of which were occupied by EC layer components. Structural changes were accompanied by increased pulmonary artery medial thickness and right ventricular hypertrophy (RVH). Moreover, abnormalities in ECs persisted even after exposure to 7days of normal air replacement conditions. Results were confirmed by morphometric quantification. Our results suggest that the abnormal morphology of capillary ECs and thick BABs correlates with pulmonary artery remodeling and RVH. These ultrastructural changes might represent possible mechanisms of secondary pulmonary hypertension in BPD.

Practices surrounding pulmonary hypertension and bronchopulmonary dysplasia amongst neonatologists caring for premature infants.

Pulmonary hypertension (PH) is associated with bronchopulmonary dysplasia (BPD). Screening strategies, a thorough investigation of co-morbidities, and multidisciplinary involvement prior to anti-PH medications have been advocated by recent guidelines. We sought to evaluate current practices of neonatologists caring for premature infants with PH. Electronic survey of American Academy of Pediatrics neonatology members. Among 306 neonatologist respondents, 38% had an institutional screening protocol for patients with BPD; 83% screened at 36 weeks for premature neonates on oxygen/mechanical ventilation. In those practicing more than 5 years, 54% noted increasing numbers of premature infants diagnosed with PH. Evaluation for PH in BPD patients included evaluations for micro-aspiration (41%), airways anomalies (29%), and catheterization (10%). Some degree of acquired pulmonary vein stenosis was encountered in 47%. A majority (90%) utilized anti-PH medications during the neonatal hospitalization. Screening for PH in BPD, and subsequent evaluation and management is highly variable.

Bronchopulmonary dysplasia and pulmonary hypertension: a meta-analysis.

Pulmonary hypertension (PH) is a complication of bronchopulmonary dysplasia (BPD) but the true impact of PH in patients with BPD remains unclear. We sought to systematically review and meta-analyze incidence of PH in BPD and compare clinical outcomes of BPD patients with PH to those without PH in preterm infants. Medline, Embase, PsychINFO and CINAHL were searched from January 2000 through December 2015. Cohort, case-control and randomized studies were included. Case-reports, case-series and letters to editors and studies with high risk of bias were excluded. Study design, inclusion/exclusion criteria, diagnostic criteria for BPD and PH and outcomes were extracted independently by two co-authors. The pooled incidence of PH in patients with BPD (any severity) was 17% (95% confidence interval (CI) 12 to 21; 7 studies) and 24% (95% CI 17 to 30; 9 studies) in moderate-severe BPD. Patients with BPD have higher unadjusted odds of developing PH compared to those without BPD (odds ratio (OR) 3.00; 95% CI 1.18 to 7.66; 4 studies). Patients with BPD and PH were at higher odds of mortality (OR 5.29; 95% CI 2.07 to 13.56; 3 studies) compared with BPD without PH, but there was no significant difference in duration of initial hospitalization, duration of supplemental oxygen requirement or need for home oxygen. No studies included in this review reported on long-term pulmonary or neurodevelopmental outcomes. PH occurs in one out of 4 to 5 preterm neonates with BPD. Patients with BPD and PH may have higher odds of mortality; however, there is urgent need for high quality studies that control for confounders and provide data on long-term outcomes.

An arginase-1 SNP that protects against the development of pulmonary hypertension in bronchopulmonary dysplasia enhances NO-mediated apoptosis in lymphocytes.

Arginase and nitric oxide synthase (NOS) share a common substrate, l‐arginine, and have opposing effects on vascular remodeling. Arginase is the first step in polyamine and proline synthesis necessary for cellular proliferation, while NO produced from NOS promotes apoptosis. Previously, we identified a single nucleotide polymorphism (SNP) in the arginase‐1 (ARG1) gene, rs2781666 (T‐allele) that was associated with a decreased risk for developing pulmonary hypertension (PH) in a cohort of infants with bronchopulmonary dysplasia (BPD). In this study, we utilized lymphocytes from neonates (the only readily available cells from these patients expressing the two genotypes of interest) with either the rs2781666 SNP (TT) or wild type (GG) to test the hypothesis that the protection of the ARG1 SNP against the development of PH in BPD would involve augmented NO production leading to more apoptosis. Lymphocytes were stimulated with IL‐4, IL‐13, and phorbol myristate acetate (PMA). We found that TT lymphocytes had similar levels of arginase I and arginase II expression, but there was a tendency for lower urea production (a surrogate marker of arginase activity), than in the GG lymphocytes. The TT lymphocytes also had significantly greater NO production than did GG lymphocytes despite no differences in iNOS expression between genotypes. Furthermore, the TT lymphocytes had lower numbers of viable cells, and higher levels of cleaved caspase‐3 than did GG lymphocytes. Inhibiting NOS activity using N ω‐Nitro‐l‐arginine methyl ester hydrochloride (l‐NAME) significantly decreased cleaved caspase‐3 levels in the TT lymphocytes. These data demonstrate that the TT genotype results in greater levels of NO production leading to more apoptosis, which is consistent with the concept that BPD patients with the TT genotype are protected against the development of PH by producing greater basal levels of endogenous NO.

Endothelin-1-Rho kinase interactions impair lung structure and cause pulmonary hypertension after bleomycin exposure in neonatal rat pups.

Bronchopulmonary dysplasia (BPD) is the chronic lung disease associated with premature birth, characterized by impaired vascular and alveolar growth. In neonatal rats bleomycin decreases lung growth and causes pulmonary hypertension (PH), which is poorly responsive to nitric oxide. In the developing lung, through Rho kinase (ROCK) activation, ET-1 impairs endothelial cell function; however, whether ET-1-ROCK interactions contribute to impaired vascular and alveolar growth in experimental BPD is unknown. Neonatal rats were treated daily with intraperitoneal bleomycin with and without selective ETA (BQ123/BQ610) and ETB (BQ788) receptor blockers, nonselective ET receptor blocker (ETRB) (bosentan), or fasudil (ROCK inhibitor). At day 14, lungs were harvested for morphometrics, and measurements of Fulton's index (RV/LV+S), medial wall thickness (MWT), and vessel density. Lung ET-1 protein and ROCK activity (phospho-MYPT-1:total MYPT-1 ratio) were also measured by Western blot analysis. Bleomycin increased lung ET-1 protein expression by 65%, RV/LV+S by 60%, mean linear intercept (MLI) by 212%, and MWT by 140% and decreased radial alveolar count (RAC) and vessel density by 40 and 44%, respectively (P < 0.01 for each comparison). After bleomycin treatment, fasudil and bosentan partially restored RAC and vessel density and decreased MLI, RV/LV+S, and MWT to normal values. Bleomycin increased ROCK activity by 120%, which was restored to normal values by bosentan but not selective ETRB. We conclude that ET-1-ROCK interactions contribute to decreased alveolar and vascular growth and PH in experimental BPD. We speculate that nonselective ETRB and ROCK inhibitors may be effective in the treatment of infants with BPD and PH.

Sildenafil, pulmonary hypertension and bronchopulmonary dysplasia

Pulmonary hypertension (PH) secondary to bronchopulmonary dysplasia (BPD) in infants remains a serious concern and continues to cause significant morbidity despite improvements in both quality of life and survival for patients. One of the potential agents that might help is sildenafil citrate, a phosphodiesterase-V inhibitor used a first line therapy for idiopathic PH. However, only limited evidence exists for its use as either monotherapy or part of a combination approach towards the management of PH in BPD. The evidence and current knowledge is presented for sildenafil alone and in combination with other disease modifying agents to treat PH in the presence of BPD. We have previously suggested that sildenafil appears to be safe and possibly effective in this condition. We present the evidence that if continued until PH resolution, there might be reduced mortality in this debilitating disease.

A single nucleotide polymorphism in the dimethylarginine dimethylaminohydrolase gene is associated with lower risk of pulmonary hypertension in bronchopulmonary dysplasia.

Pulmonary hypertension (PH) develops in 25-40% of bronchopulmonary dysplasia (BPD) patients, substantially increasing mortality. We have previously found that asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide (NO) production, is elevated in patients with BPD-associated PH. ADMA is metabolised by N(ᴳ) ,N(ᴳ) -dimethylarginine dimethylaminohydrolase (DDAH). Presently, we test the hypothesis that there are single nucleotide polymorphisms (SNPs) in DDAH1 and/or DDAH2 associated with the development of PH in BPD patients. BPD patients were enrolled (n = 98) at Nationwide Children's Hospital. Clinical characteristics and 36 SNPs in DDAH1 and DDAH2 were compared between BPD-associated PH patients (cases) and BPD-alone patients (controls). In BPD patients, 25 (26%) had echocardiographic evidence of PH (cases). In this cohort, DDAH1 wild-type rs480414 was 92% sensitive and 53% specific for PH in BPD, and the DDAH1 SNP rs480414 decreased the risk of PH in an additive model of inheritance (OR = 0.39; 95% CI [0.18-0.88], p = 0.01). The rs480414 SNP in DDAH1 may be protective against the development of PH in patients with BPD. Furthermore, the DDAH1 rs480414 may be a useful biomarker in developing predictive models for PH in patients with BPD.

Bronchopulmonary Dysplasia‐associated Pulmonary Hypertension and Mutations in the DDAH1 Gene

Pulmonary hypertension (PH) is the most common comorbidity in bronchopulmonary dysplasia (BPD). Nitric oxide (NO) is produced by endothelial NO synthase (eNOS) in pulmonary endothelial cells and is a potent vasodilator. Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of eNOS. Our group has recently published that ADMA levels are elevated in patients with BPD‐associated PH. ADMA is degraded by the enzyme Nᴳ,Nᴳ‐ dimethylarginine dimethylaminohydrolase (DDAH). We tested the hypothesis that there are single nucleotide polymorphisms (SNPs) in DDAH1 associated with PH in BPD. Neonates diagnosed with BPD were enrolled at Nationwide Children's Hospital. DNA was assayed for 34 SNPs in DDAH1. Calculated minor allele frequency (MAF) for cases and controls were compared using chi‐square. Assuming recessive inheritance, logistic regression was used to estimate the odds ratio. Among 157 patients with BPD, 53 (34%) also had echocardiographic evidence of PH. Gestational age, birth weight, gender, and ethnicity were not different between groups. DDAH1 rs11161637 SNP had a significantly (p=0.04) lower MAF in cases than in controls (0.25 vs. 0.34). DDAH1 rs11161637 was associated with a decreased risk (OR=0.27) of PH in this BPD cohort, although this did not reach statistical significance (p‐value=0.06). These results indicate that rs11161637 SNP of DDAH1 may be protective against the development of PH in patients with BPD. We speculate that the DDAH1 SNP may be a useful biomarker in developing predictive models for PH in patients with BPD.