Pulmonary Circulation

Pediatric pulmonary vascular disease (PVD) and pulmonary hypertension (PH) represent phenotypically and pathophysiologically diverse disease categories, contributing substantial morbidity and mortality to a complex array of pediatric conditions. Here, we review the multifactorial nature of pediatric PVD, with an emphasis on improved recognition, phenotyping, and endotyping strategies for pediatric PH. Novel tailored approaches to diagnosis and treatment in pediatric PVD, as well as the implications for long-term outcomes, are highlighted.

Pulmonary hypertension (PH) is characterized by severe exercise limitation mainly attributed to the impairment of right ventricular function resulting from a concomitant elevation of pulmonary vascular resistance and pressure. The unquestioned cornerstone in the management of patients with pulmonary arterial hypertension (PAH) is specific vasoactive medical therapy to improve pulmonary hemodynamics and strengthen right ventricular function. Nevertheless, evidence for a beneficial effect of exercise training (ET) on pulmonary hemodynamics and functional capacity in patients with PH has been growing during the past decade. Beneficial effects of ET on regulating factors, inflammation, and metabolism have also been described. Small case-control studies and randomized clinical trials in larger populations of patients with PH demonstrated substantial improvements in functional capacity after ET. These findings were accompanied by several studies that suggested an effect of ET on inflammation, although a direct link between this effect and the therapeutic benefit of ET in PH has not yet been demonstrated. On this background, the aim of the present review is to describe current concepts regarding the effects of exercise on the pulmonary circulation and pathophysiological limitations, as well as the clinical and mechanistic effects of exercise in patients with PH.

Pulmonary arterial hypertension (PH) and chronic kidney disease (CKD) both profoundly impact patient outcomes, whether as primary disease states or as co-morbid conditions. PH is a common co-morbidity in CKD and vice versa. A growing body of literature describes the epidemiology of PH secondary to chronic kidney disease and end-stage renal disease (ESRD) (WHO group 5 PH). But, there are only limited data on the epidemiology of kidney disease in group 1 PH (pulmonary arterial hypertension [PAH]). The purpose of this review is to summarize the current data on epidemiology and discuss potential disease mechanisms and management implications of kidney dysfunction in PAH. Kidney dysfunction, determined by serum creatinine or estimated glomerular filtration rate, is a frequent co-morbidity in PAH and impaired kidney function is a strong and independent predictor of mortality. Potential mechanisms of PAH affecting the kidneys are increased venous congestion, decreased cardiac output, and neurohormonal activation. On a molecular level, increased TGF-β signaling and increased levels of circulating cytokines could have the potential to worsen kidney function. Nephrotoxicity does not seem to be a common side effect of PAH-targeted therapy. Treatment implications for kidney disease in PAH include glycemic control, lifestyle modification, and potentially Renin-Angiotensin-Aldosterone System (RAAS) blockade.

Development of the pulmonary circulation is a complex process with a spatial pattern that is tightly controlled. This process is vulnerable for disruption by various events in the prenatal and early postnatal periods. Disruption of normal pulmonary vascular development leads to abnormal structure and function of the lung vasculature, causing neonatal pulmonary vascular diseases. Premature babies are especially at risk of the development of these diseases, including persistent pulmonary hypertension and bronchopulmonary dysplasia. Reactive oxygen species play a key role in the pathogenesis of neonatal pulmonary vascular diseases and can be caused by hyperoxia, mechanical ventilation, hypoxia, and inflammation. Besides the well-established short-term consequences, exposure of the developing lung to injurious stimuli in the perinatal period, including oxidative stress, may also contribute to the development of pulmonary vascular diseases later in life, through so-called “fetal or perinatal programming.” Because of these long-term consequences, it is important to develop a follow-up program tailored to adolescent survivors of neonatal pulmonary vascular diseases, aimed at early detection of adult pulmonary vascular diseases, and thereby opening the possibility of early intervention and interfering with disease progression. This review focuses on pathophysiologic events in the perinatal period that have been shown to disrupt human normal pulmonary vascular development, leading to neonatal pulmonary vascular diseases that can extend even into adulthood. This knowledge may be particularly important for ex-premature adults who are at risk of the long-term consequences of pulmonary vascular diseases, thereby contributing disproportionately to the burden of adult cardiovascular disease in the future.

The many types of pulmonary hypertension (PH) are so protean in their biological origin, histological expression, and natural history that it is difficult to create a summary picture of the disease, or to easily compare and contrast characteristics of one type of PH with another. For newcomers to the field, however, such a picture would facilitate a broad understanding of PH. In this paper, we suggest that four characteristics are fundamental to describing the nature of various types of PH, and that taken together they define a number of patterns of PH expression. These characteristics are histopathology, developmental origin, associated clinical conditions, and potential for resolution. The “snapshot” is a way to concisely display the ways that these signal characteristics intersect in select specific types of PH, and is an effort to summarize these patterns in a way that facilitates a “big picture” comprehension of this disease.

The primary aim was to explore the safety and tolerability of inhaled treprostinil when used in patients with pulmonary hypertension (PH) with concomitant chronic obstructive pulmonary disease (COPD). Patients with a diagnosis of pre-capillary PH (defined as pulmonary artery mean pressure of ≥ 25 mmHg and pulmonary artery wedge pressure or left ventricular end diastolic pressure of ≤ 15 mmHg) who were being initiated on inhaled treprostinil and had concomitant COPD (defined as FEV1/FVC ratio ≤ 70% with FEV1 ≥ 40% predicted) were considered for inclusion in this pilot study. Assessments included adverse events, physical exam, World Health Organization (WHO) functional class, 6-minute walk test (6MWT), modified Borg dyspnea score, and concomitant medication. At baseline and week 16 St. George’s Respiratory Questionnaire (SGRQ), arterial blood gas (ABG), and pulmonary function test (PFT) were assessed. The median age was 65 years (age range, 56–80 years) and five patients (56%) were men. Among the nine patients, a majority had an increase in 6MWT from baseline to week 16 (median change, 19 m). Only three of the nine patients (33%) had an increase in A-a gradient at week 16 (median change, –7). There was no difference in any of the following: arterial blood gases, WHO functional class, 6MWT results, or SGRQ scores from baseline to week 16. There was a statistically significant decline in several of the PFT measures, including FEV1 (median change, –0.18 L; P = 0.004; median change, –7% of predicted; P = 0.016), FVC (median change, –0.23 L; P = 0.027), and diffusion capacity for carbon monoxide (DLCO) (median change, –5% of predicted; P = 0.023). The small number of patients limits firm conclusions; however, inhaled treprostinil did not seem to adversely impact oxygenation in the majority of the study patients with pre-capillary PH and COPD. While there may have an adverse impact on some pulmonary function parameters, the clinical significance is unclear.

Alterations in the nitric oxide (NO) pathway play a major role in pulmonary arterial hypertension (PAH). L-arginine (LA) and tetrahydrobiopterin (BH₄) are main substrates in the production of NO, which mediates pulmonary vasodilation. Administration of either LA or BH₄ decrease pulmonary artery pressure (PAP). A combined administration of both may have synergistic effects in the therapy of PAH. In a telemetrically monitored model of unilateral pneumonectomy and monocrotaline-induced PAH, male Sprague-Dawley rats received either LA (300 mg/kg; n = 15), BH₄ (20 mg/kg; n = 15), the combination of LA and BH₄ (300 mg/kg, 20 mg/kg; n = 15), or vehicle (control group; n = 10) from day 28 after monocrotaline induction. Therapy was orally administered once daily over consecutive 14 days. LA, BH₄, or both equally lowered PAP, increased pulmonary vascular elasticity, restored spontaneous locomotoric activity, prevented body weight loss and palliated small vessel disease of severely pulmonary hypertensive rats. BH₄ substitution lowered asymmetric dimethylarginine levels sustainably at 60 min after administration and downregulated endothelial NO synthase mRNA expression. No significant survival, macro- and histomorphologic or hemodynamic differences were found between therapy groups at the end of the study period. Administration of LA and BH₄ both mediated a decrease of mean PAP, attenuated right ventricular hypertrophy and small vessel disease in monocrotaline-induced pulmonary hypertensive rats, though a combined administration of both substances did not reveal any synergistic therapy effects in our animal model.

Chronic hypoxia-induced pulmonary hypertension (PH) is characterized by increased pressure and resistance in the pulmonary vasculature and hypertrophy of the right ventricle (RV). The transcription factors, nuclear factor activated T-cells (NFAT), and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB/p65) contribute to RV hypertrophy (RVH). Because peroxisome proliferator-activated receptor gamma (PPARγ) activation attenuates hypoxia-induced PH and RVH, we hypothesized that PPARγ inhibits activation of RV hypertrophic transcriptional signaling mechanisms. C57BL/6J mice were exposed to normoxia (21% O₂) or hypoxia (10% O₂) for 21 days. During the final 10 days of exposure, selected mice were treated with the PPARγ ligand, pioglitazone. RV systolic pressure (RVSP) and RVH were measured, and NFATc2 and NF-kB/p65 protein levels were measured in RV and LV nuclear and cytosolic fractions. Cardiomyocyte hypertrophy was assessed with wheatgerm agglutinin staining. NFAT activation was also examined with luciferase reporter mice and analysis of protein levels of selected transcriptional targets. Chronic-hypoxia increased: (1) RVH, RVSP, and RV cardiomyocyte hypertrophy; (2) NFATc2 and NF-κB activation in RV nuclear homogenates; (3) RV and LV NFAT luciferase activity; and (4) RV protein levels of brain natriuretic peptide (BNP) and β-myosin heavy chain (β-MyHC). Treatment with pioglitazone attenuated hypoxia-induced increases in both RV and LV NFAT luciferase activity. Chronic hypoxia caused sustained RV NFATc2 and NF-κB activation. Pioglitazone attenuated PH, RVH, cardiomyocyte hypertrophy, and activation of RV hypertrophic signaling and also attenuated LV NFAT activation. PPARγ favorably modulates signaling derangements in the heart as well as in the pulmonary vascular wall.

Vasculopathies, characterized by the formation of fragile and abnormal microvessels, are associated with the severity of many chronic lung diseases, including pulmonary fibrosis, emphysema/chronic obstructive pulmonary disease, systemic sclerosis, and hypertension. However, the study of human lung vasculature has been limited by the ability to isolate generous quantities of microvascular endothelial cells (MVEC) free from mesenchymal contamination. Expansion and passaging of primary human MVEC in vitro typically results in loss of a traditional phenotype in favor of an intermediate mesenchymal one, as early as passage five. Here we provide a detailed protocol for the selection of large quantities of enriched primary human lung MVEC based upon differential adherence from mesenchyme and simple magnetic separation, which decreases the need for excessive passaging, in order to obtain sufficient cell numbers to successfully freeze stock cultures. Additional protocols are provided for Ac-di-LDL selection, characterization, and a sandwich angiogenesis method of functional tube formation. The complete protocol including cell isolation and characterization takes approximately six weeks to complete.

We have demonstrated that simvastatin and sphingosine 1−phosphate (S1P) both attenuate increased vascular permeability in preclinical models of acute respiratory distress syndrome. However, the underlying mechanisms remain unclear. As Krüppel-like factor 2 (KLF2) serves as a critical regulator for cellular stress response in endothelial cells (EC), we hypothesized that simvastatin enhances endothelial barrier function via increasing expression of the barrier-promoting S1P receptor, S1PR1, via a KLF2-dependent mechanism. S1PR1 luciferase reporter promoter activity in human lung artery EC (HPAEC) was tested after simvastatin (5 μM), and S1PR1 and KLF2 protein expression detected by immunoblotting. In vivo, transcription and expression of S1PR1 and KLF2 in mice lungs were detected by microarray profiling and immunoblotting after exposure to simvastatin (10 mg/kg). Endothelial barrier function was measured by trans-endothelial electrical resistance with the S1PR1 agonist FTY720-(S)-phosphonate. Both S1PR1 and KLF2 gene expression (mRNA, protein) were significantly increased by simvastatin in vitro and in vivo. S1PR1 promoter activity was significantly increased by simvastatin (P < 0.05), which was significantly attenuated by KLF2 silencing (siRNA). Simvastatin induced KLF2 recruitment to the S1PR1 promoter, and consequently, significantly augmented the effects of the S1PR1 agonist on EC barrier enhancement (P < 0.05), which was significantly attenuated by KLF2 silencing (P < 0.05). These results suggest that simvastatin upregulates S1PR1 transcription and expression via the transcription factor KLF2, and consequently augments the effects of S1PR1 agonists on preserving vascular barrier integrity. These results may lead to novel combinatorial therapeutic strategies for lung inflammatory syndromes.

Pulmonary arterial hypertension (PAH) is rare in children and few data are available in a pediatric general population. This study aims to calculate the annual incidence and prevalence of PAH and to describe these children in a large US population of patients aged under 18 years. Using the US MarketScan claims database we identified 695 children with PAH in 2010–2013. We calculated annual incidence rates and prevalence overall, by age and PAH type (idiopathic and non-idiopathic) using Byar’s method. We also described characteristics, co-morbidities, treatment patterns, and diagnostic procedures for these children. In 2010–2013, the annual incidence rates of PAH per 1,000,000 children-years was in the range of 4.8–8.1; 0.5–0.9 for idiopathic PAH and 4.3–7.3 for non-idiopathic PAH. The annual prevalence of PAH was in the range of 25.7–32.6 per 1,000,000 children; 4.4–6.0 for idiopathic PAH and 21.3–27.0 for non-idiopathic PAH. Incidence rates and prevalence were highest in children under age 2 years. Around 36% of affected children were born prematurely. Most (75%) had some type of congenital heart defect and 13% had Down’s syndrome. Most patients received PAH monotherapy (83%), while 13% received dual therapy. Phosphodiesterase type 5 inhibitors were the most commonly used treatments. Around 92% had at least one echocardiogram and 37% a right heart catheterization. PAH is very rare in children especially in the absence of etiological factors such as congenital heart defects. A large proportion of diagnoses in children seem to be based on echocardiography rather than right heart catheterization.

Patients with idiopathic pulmonary arterial hypertension (IPAH) and a reduced diffusion capacity of the lung for carbon monoxide (DLCO) have a worse survival compared to IPAH patients with a preserved DLCO. Whether this poor survival can be explained by unresponsiveness to pulmonary hypertension (PH)-specific vasodilatory therapy is unknown. Therefore, the aim of this study was to evaluate the hemodynamic and cardiac response to PH-specific vasodilatory therapy in patients with IPAH and a reduced DLCO. Retrospectively, we studied treatment naïve hereditary and IPAH patients diagnosed between January 1990 and May 2015 at the VU University Medical Center. After exclusion of participants without available baseline DLCO measurement or right heart catheterization data and participants carrying a BMPR2 mutation, 166 participants could be included in this study. Subsequently, hemodynamics, cardiac function, exercise capacity, and oxygenation at baseline and after PH-specific vasodilatory therapy were compared between IPAH patients with a preserved DLCO (DLCO >62%), IPAH patients with a moderately reduced DLCO (DLCO 43–62%), and IPAH patients with a severely reduced DLCO (DLCO <43%). Baseline hemodynamics and right ventricular function were not different between groups. Baseline oxygenation was worse in patients with IPAH and a severely reduced DLCO. Hemodynamics and cardiac function improved in all groups after PH-specific vasodilatory therapy without worsening of oxygenation at rest or during exercise. Patients with IPAH and a severely reduced DLCO show a similar response to PH-specific vasodilatory therapy in terms of hemodynamics, cardiac function, and exercise capacity as patients with IPAH and a moderately reduced or preserved DLCO.

Pulmonary hypertension (PH) complicating chronic obstructive pulmonary disease (COPD-PH) and interstitial lung disease (ILD-PH) (World Health Organization [WHO] Group III PH) increases medical costs and reduces survival. Despite limited data, many clinicians are using pulmonary arterial hypertension (PAH)-specific therapy to treat WHO Group III PH patients. To further investigate the utility of PAH-specific therapy in WHO Group III PH, we performed a systematic review and meta-analysis. Relevant studies from January 2000 through May 2016 were identified in the MEDLINE, EMBASE, and COCHRANE electronic databases and www.clinicaltrials.gov. Change in six-minute walk distance (6MWD) was estimated using random effects meta-analysis techniques. Five randomized controlled trials (RCTs) in COPD-PH (128 placebo or standard treatment and 129 PAH-medication treated patients), two RCTs in ILD-PH (23 placebo and 46 treated patients), and four single-arm clinical trials (50 patients) in ILD-PH were identified. Treatment in both COPD-PH and ILD-PH did not worsen hypoxemia. Symptomatic burden was not consistently reduced but there were trends for reduced pulmonary artery pressures and pulmonary vascular resistance with PAH-specific therapy. As compared to placebo, 6MWD was not significantly improved with PAH-specific therapy in the five COPD-PH RCTs (42.7 m; 95% confidence interval [CI], –1.0 – 86.3). In the four single-arm studies in ILD-PH patients, there was a significant improvement in 6MWD after PAH-specific treatment (46.2 m; 95% CI, 27.9–64.4), but in the two ILD-PH RCTs there was not an improvement (21.6 m; 95% CI, –17.8 – 61.0) in exercise capacity when compared to placebo. Due to the small numbers of patients evaluated and inconsistent beneficial effects, the utility of PAH-specific therapy in WHO Group III PH remains unproven. A future clinical trial that is appropriately powered is needed to definitively determine the efficacy of this widely implemented treatment approach.

Pulmonary hypertension (PH) results in significant morbidity and mortality. Chronic PH animal models may advance the study of PH’s mechanisms, evolution, and therapy. In this report, we describe the challenges and successes in developing three models of chronic PH in large animals: two models (one canine and one swine) utilized repeated infusions of ceramic microspheres into the pulmonary vascular bed, and the third model employed a surgical aorto-pulmonary shunt. In the canine model, seven dogs underwent microsphere infusions that resulted in progressive elevation of pulmonary arterial pressure over a few months. In this model, pulmonary endoarterial tissue was obtained for histology. In the aorto-pulmonary shunt swine model, 17 pigs developed systemic level pulmonary pressures after 2–3 months. In this model, pulmonary endoarterial tissue was sequentially obtained to assess for changes in gene and microRNA expression. In the swine microsphere infusion model, three pigs developed only a modest chronic increase in pulmonary arterial pressure, despite repeated infusions of microspheres (up to 40 in one animal). The main purpose of this model was for vasodilator testing, which was performed successfully immediately after acute microsphere infusions. Chronic PH in large animal models can be successfully created; however, a model’s characteristics need to match the investigational goals.

Oral treprostinil was recently labeled for treatment of pulmonary arterial hypertension. Similar to the period immediately after parenteral treprostinil was approved, there is a significant knowledge gap for practicing physicians who might prescribe oral treprostinil. Despite its oral route of delivery, use of the drug is challenging because of the requirement for careful titration and management of drug-related adverse effects. We aimed to create a consensus document combining available evidence with expert opinion to provide guidance for use of oral treprostinil. Following a methodology commonly used in business and social sciences (the ‘Delphi Process’), two investigators from the oral treprostinil (Freedom) studies created a series of statements based on available evidence and the package insert. The set of ‘best practice’ statements was circulated to nine other Freedom trial investigators. Their comments were incorporated into the document as new line items for further vote and comment. The subsequent document was put to vote line by line (scale of −5 to +5) and a final statement was drafted. Consensus recommendations include initial therapy with 0.125 mg for treatment na patients, three times daily dosing, aggressive use of antidiarrheal medication, and a strong preference for use of the drug in combination with other approved PAH therapies. This process was particularly valuable in providing guidance for the management of adverse events (where essentially no data is available). The Delphi process was useful to codify investigator experience and subsequently develop investigator consensus about practical issues for physicians who may wish to prescribe oral treprostinil.

Hypoxia-induced pulmonary hypertension (HPH) is a progressive disease that is mainly caused by chronic exposure to high altitude, chronic obstructive lung disease, and obstructive sleep apnea. The increased pulmonary vascular resistance and increased pulmonary arterial pressure result in increased right ventricular afterload, leading to right heart failure and increased morbidity. There are several clinical reports suggesting a link between PH and diabetes, insulin resistance, or obesity; however, it is unclear whether HPH is associated with diabetes as a progressive complication in diabetes. The major goal of this study is to examine the effect of diabetic “preconditioning” or priming effect on the progression of HPH and define the molecular mechanisms that explain the link between diabetes and HPH. Our data show that HPH is significantly enhanced in diabetic mice, while endothelium-dependent relaxation in pulmonary arteries is significantly attenuated in chronically hypoxic diabetic mice (DH). In addition, we demonstrate that mouse pulmonary endothelial cells (MPECs) isolated from DH mice exhibit a significant increase in mitochondrial reactive oxygen species (ROS) concentration and decreased SOD2 protein expression. Finally, scavenging mitochondrial ROS by mitoTempol restores endothelium-dependent relaxation in pulmonary arteries that is attenuated in DH mice. These data suggest that excessive mitochondrial ROS production in diabetic MPECs leads to the development of severe HPH in diabetic mice exposed to hypoxia.

Pulmonary arterial hypertension (PAH) is increasingly recognized as a systemic disease driven by alteration in the normal functioning of multiple metabolic pathways affecting all of the major carbon substrates, including amino acids. We found that human pulmonary hypertension patients (WHO Group I, PAH) exhibit systemic and pulmonary-specific alterations in glutamine metabolism, with the diseased pulmonary vasculature taking up significantly more glutamine than that of controls. Using cell culture models and transgenic mice expressing PAH-causing BMPR2 mutations, we found that the pulmonary endothelium in PAH shunts significantly more glutamine carbon into the tricarboxylic acid (TCA) cycle than wild-type endothelium. Increased glutamine metabolism through the TCA cycle is required by the endothelium in PAH to survive, to sustain normal energetics, and to manifest the hyperproliferative phenotype characteristic of disease. The strict requirement for glutamine is driven by loss of sirtuin-3 (SIRT3) activity through covalent modification by reactive products of lipid peroxidation. Using 2-hydroxybenzylamine, a scavenger of reactive lipid peroxidation products, we were able to preserve SIRT3 function, to normalize glutamine metabolism, and to prevent the development of PAH in BMPR2 mutant mice. In PAH, targeting glutamine metabolism and the mechanisms that underlie glutamine-driven metabolic reprogramming represent a viable novel avenue for the development of potentially disease-modifying therapeutics that could be rapidly translated to human studies.

Pulmonary endothelial cell (EC) barrier dysfunction and recovery is critical to the pathophysiology of acute respiratory distress syndrome. Cytoskeletal and subsequent cell membrane dynamics play a key mechanistic role in determination of EC barrier integrity. Here, we characterizAQe the actin related protein 2/3 (Arp 2/3) complex, a regulator of peripheral branched actin polymerization, in human pulmonary EC barrier function through studies of transendothelial electrical resistance (TER), intercellular gap formation, peripheral cytoskeletal structures and lamellipodia. Compared to control, Arp 2/3 inhibition with the small molecule inhibitor CK-666 results in a reduction of baseline barrier function (1,241 ± 53 vs 988 ± 64 ohm; p < 0.01), S1P-induced barrier enhancement and delayed recovery of barrier function after thrombin (143 ± 14 vs 93 ± 6 min; p < 0.01). Functional changes of Arp 2/3 inhibition on barrier integrity are associated temporally with increased intercellular gap area at baseline (0.456 ± 0.02 vs 0.299 ± 0.02; p < 0.05) and thirty minutes after thrombin (0.885 ± 0.03 vs 0.754 ± 0.03; p < 0.05). Immunofluorescent microscopy reveals reduced lamellipodia formation after S1P and during thrombin recovery in Arp 2/3 inhibited cells. Individual lamellipodia demonstrate reduced depth following Arp 2/3 inhibition vs vehicle at baseline (1.83 ± 0.41 vs 2.55 ± 0.46 µm; p < 0.05) and thirty minutes after S1P treatment (1.53 ± 0.37 vs 2.09 ± 0.36 µm; p < 0.05). These results establish a critical role for Arp 2/3 activity in determination of pulmonary endothelial barrier function and recovery through formation of EC lamellipodia and closure of intercellular gaps.

Quantifying metabolic derangements in pulmonary hypertension (PH) by plasma metabolomics could identify biomarkers useful for diagnosis and treatment. The objective of this paper is to test the hypotheses that circulating metabolites are differentially expressed in PH patients compared with controls and among different hemodynamic subtypes of PH associated with left heart disease. We studied patients enrolled in the CATHGEN biorepository with PH (right heart catheterization mPAP ≥ 25 mmHg; n = 280). Of these, 133 met criteria for postcapillary PH, 82 for combined precapillary and postcapillary PH (CpcPH), and 65 for precapillary PH. Targeted profiling of 63 metabolites (acylcarnitines, amino acids, and ketones) was performed using tandem flow injection mass spectrometry. Multivariable linear regression was used to determine differences in metabolite factors derived from a principal components analysis between PH cases, PH subtypes, and non-PH controls. In adjusted models, the metabolite factor loaded with long-chain acylcarnitines was higher in all PH cases versus non-PH controls (P = 0.00008), but did not discriminate between CpcPH and postcapillary PH (P = 0.56). In analyses of subtypes, CpcPH patients had lower levels of factors loaded with urea cycle amino acids and short chain acylcarnitines as compared to controls (P = 0.002 and P = 0.01, respectively) and as compared to postcapillary PH (P = 0.04 and P = 0.02, respectively). Compared to controls, PH was strongly associated with greater concentrations of long-chain acylcarnitines. Postcapillary PH and CpcPH were weakly associated with distinct metabolomic profiles. These findings suggest the presence of unique metabolic abnormalities in subtypes of PH and may reflect underlying pathophysiology.

Bone morphogenetic protein (BMP) signaling regulates vascular smooth muscle maturation, endothelial cell proliferation, and tube formation. The endogenous BMP antagonist Follistatin-like 1 (Fstl1) is highly expressed in pulmonary vascular endothelium of the developing mouse lung, suggesting a role in pulmonary vascular formation and vascular homeostasis. The aim of this study was to investigate the role of Fstl1 in the pulmonary vascular endothelium. To this aim, Fstl1 was conditionally deleted from endothelial and endothelial-derived cells using Tie2-cre driven Fstl1-KO mice (Fstl1-eKO mice). Endothelial-specific Fstl1 deletion was postnatally lethal, as ∼70% of Fstl1-eKO mice died at three weeks after birth. Deletion of Fstl1 from endothelium resulted in a reduction of right ventricular output at three weeks after birth compared with controls. This was associated with pulmonary vascular remodeling, as the percentage of actin-positive small pulmonary vessels was increased at three weeks in Fstl1-eKO mice compared with controls. Endothelial deletion of Fstl1 resulted in activation of Smad1/5/8 signaling and increased BMP/Smad-regulated gene expression of Jagged1, Endoglin, and Gata2 at one week after birth compared with controls. In addition, potent vasoconstrictor Endothelin-1, the expression of which is driven by Gata2, was increased in expression, both on the mRNA and protein levels, at one week after birth compared with controls. At three weeks, Jagged1 was reduced in the Fstl1-eKO mice whereas Endoglin and Endothelin-1 were unchanged. In conclusion, loss of endothelial Fstl1 in the lung is associated with elevated BMP-regulated genes, impaired small pulmonary vascular remodeling, and decreased right ventricular output.

17β-estradiol (E2) exerts complex and context-dependent effects in pulmonary hypertension. In hypoxia-induced pulmonary hypertension (HPH), E2 attenuates lung vascular remodeling through estrogen receptor (ER)-dependent effects; however, ER target genes in the hypoxic lung remain unknown. In order to identify the genome regulated by the E2-ER axis in the hypoxic lung, we performed a microarray analysis in lungs from HPH rats treated with E2 (75 mcg/kg/day) ± ER-antagonist ICI182,780 (3 mg/kg/day). Untreated HPH rats and normoxic rats served as controls. Using a false discovery rate of 10%, we identified a significantly differentially regulated genome in E2-treated versus untreated hypoxia rats. Genes most upregulated by E2 encoded matrix metalloproteinase 8, S100 calcium binding protein A8, and IgA Fc receptor; genes most downregulated by E2 encoded olfactory receptor 63, secreted frizzled-related protein 2, and thrombospondin 2. Several genes affected by E2 changed in the opposite direction after ICI182,780 co-treatment, indicating an ER-regulated genome in HPH lungs. The bone morphogenetic protein antagonist Grem1 (gremlin 1) was upregulated by hypoxia, but found to be among the most downregulated genes after E2 treatment. Gremlin 1 protein was reduced in E2-treated versus untreated hypoxic animals, and ER-blockade abolished the inhibitory effect of E2 on Grem1 mRNA and protein. In conclusion, E2 ER-dependently regulates several genes involved in proliferative and inflammatory processes during hypoxia. Gremlin 1 is a novel target of the E2-ER axis in HPH. Understanding the mechanisms of E2 gene regulation in HPH may allow for selectively harnessing beneficial transcriptional activities of E2 for therapeutic purposes.

Schistosomiasis (bilharzia) is a neglected parasitic disease caused by trematode flatworms of the genus Schistosoma which affects over 240 million people worldwide. It is characterized by the formation of inflammatory granulomas around deposited parasite eggs. Recent studies have revealed that immune and inflammatory responses play a crucial role in pathogenesis of schistosomiasis. The aim of this paper is to systematically evaluate the number and distribution of inflammatory cells in S. mansoni-infected mice at different doses and time points. Immunohistochemistry was performed on lung and liver tissue sections from Schistosoma-infected mice and uninfected healthy controls. Positively stained cells in whole-lung/liver tissue sections, surrounding the eggs, and in the different compartments of the tissues, were counted. We found a significant increase in the number of mast cells (toluidine blue⁺), CD3⁺ cells, CD14⁺ cells, CD68⁺ cells, and CD15⁺ cells in Schistosoma-infected tissues compared with untreated healthy controls (P ≤ 0.05 for all). Our findings revealed altered and enhanced immune cell infiltration in schistosomiasis. We suggest that these cells may contribute to the pathophysiology of Schistosoma resulting in pulmonary vascular remodeling.

Two new definitions of exercise-induced pulmonary hypertension (EIPH) have emerged. Both rely on measuring cardiac output (CO), yet this remains unstandardized. In our cohort of patients undergoing invasive cardiopulmonary exercise testing, we found that using thermodilution CO rather than direct Fick CO led to a significant excess of EIPH diagnoses.

Venous malformations have static venous lakes that predispose to spontaneous venous thrombosis within the malformation due to its low-flow static state. Thrombi of varying sizes can then embolize continually into the pulmonary arterial circulation, and occlude and narrow elastic pulmonary arteries causing chronic thromboembolic pulmonary hypertension (CTEPH). Pulmonary thromboendarterectomy (PTE) is potentially curative in CTEPH, but has not been previously reported in the setting of mediastinal and chest wall venous malformations. We report the case of a 21-year-old female with such a large malformation treated successfully with PTE. The patient underwent complete endovascular reconstruction of her subclavian vein system from the axillary vein to the innominate vein stump with covered stent grafts to exclude the malformations from causing recurrent pulmonary emboli. This was followed by embolization of the malformation to allow for the surgical approach. The series of events in this case serves as a novel approach in managing such rare patients.

Myeloproliferative neoplasia (MPN)-associated pulmonary hypertension (PH) is included in group five of the most recent clinical classification of PH. ¹ The MPNs are a heterogeneous group of disorders that includes disorders with primary expression of a myeloid phenotype and disorders characterized by expression of the Janus Kinase 2 (JAK2) mutation, p.V617F. The latter includes essential thrombocytosis, polycythemia vera, and idiopathic myelofibrosis. ² Pulmonary extra-medullary hematopoiesis (EMH) refers to the presence of hematopoietic precursor cells in the lung. It is a rare complication associated with myelofibrosis. Here we present a case series highlighting the clinical–pathological–radiological features of pulmonary EMH and PH from underlying polycythemia vera.