Familial Pulmonary Arterial Hypertension Research Articles

S99 The anti-malarial drug and lysosomal inhibitor, chloroquine, increases cell surface expression of BMPR-II

Bone morphogenetic protein receptor type II (BMPR-II) is a member of the transforming growth factor ß (TGFß) receptor superfamily. Mutations in BMPR2 are the major cause of familial pulmonary arterial...

S102 The crosstalk of PDE inhibitor with BMP signalling pathway in human pulmonary arterial smooth muscle cells

RationalSildenafil, a potent PDE inhibitor, is an established treatment for PAH. However, the detailed mechanism of its effects on the proliferation of human pulmonary artery smooth muscle cells (hPASMCs) remains...

Pulmonary Hypertension Presenting With Apnea, Cyanosis, and Failure to Thrive in a Young Child

Pulmonary Hypertension Presenting With Apnea, Cyanosis, and Failure to Thrive in a Young Child

Does the Outcome Justify an Oral-First Treatment Strategy for Management of Pulmonary Arterial Hypertension?

Guidelines for the treatment of World Health Organization (WHO) functional class (FC) III pulmonary arterial hypertension (PAH) provide for oral (PO) therapy or parenteral prostacyclins at the discretion of expert physicians. The objective of this study was to assess the appropriateness of PO-first treatment in patients with WHO FC III PAH. This study was a retrospective analysis of 79 treatment-naive adult patients with idiopathic, familial, or anorexigen-associated PAH, referred to a single pulmonary hypertension center. Forty-eight received either PO therapy with an endothelin receptor antagonist, calcium channel blocker, or phosphodiesterase inhibitor (PO group) and 31 an IV or subcutaneous (SC) prostacyclin (IV/SC group). Patients in the IV/SC group had a significantly worse baseline hemodynamic profile; however, on univariate analysis, there was no association between hemodynamics and mortality. Initial treatment with PO vs IV/SC therapy was associated with a lower overall mortality (20.8% vs 45.2%, P = .02) and a lower 5-year mortality (14.6% vs 32.3%, P = .062). Based on the National Institutes of Health equation, actual survival for patients who received PO therapy was greater than predicted at 5 years. Finally, there were similar improvements between groups in 6-min walk distance (P = .38) at 6 to 12 months after initiation of treatment. For WHO FC III PAH that is idiopathic, familial, or anorexigen associated, the clinical decision for treatment with a PO-first strategy is associated with a high survival rate when patients are appropriately risk stratified prior to initiation of therapy. The more potent prostacyclins can be reserved for high-risk patients, those with evidence of disease progression, or those with treatment failure.

Paracrine Proliferative Signaling by Senescent Cells in World Health Organization Group 3 Pulmonary Hypertension

Paracrine Proliferative Signaling by Senescent Cells in World Health Organization Group 3 Pulmonary Hypertension

Epigenetic Mechanisms of Pulmonary Hypertension

Epigenetics refers to changes in phenotype and gene expression that occur without alterations in DNA sequence. Epigenetic modifications of the genome can be acquired de novo and are potentially heritable. This review focuses on the emerging recognition of a role for epigenetics in the development of pulmonary arterial hypertension (PAH). Lessons learned from the epigenetics in cancer and neurodevelopmental diseases, such as Prader-Willi syndrome, can be applied to PAH. These syndromes suggest that there is substantial genetic and epigenetic cross-talk such that a single phenotype can result from a genetic cause, an epigenetic cause, or a combined abnormality. There are three major mechanisms of epigenetic regulation, including methylation of CpG islands, mediated by DNA methyltransferases, modification of histone proteins, and microRNAs. There is substantial interaction between these epigenetic mechanisms. Recently, it was discovered that there may be an epigenetic component to PAH. In PAH there is downregulation of superoxide dismutase 2 (SOD2) and normoxic activation of hypoxia inducible factor (HIF-1α). This decrease in SOD2 results from methylation of CpG islands in SOD2 by lung DNA methyltransferases. The partial silencing of SOD2 alters redox signaling, activates HIF-1α) and leads to excessive cell proliferation. The same hyperproliferative epigenetic abnormality occurs in cancer. These epigenetic abnormalities can be therapeutically reversed. Epigenetic mechanisms may mediate gene-environment interactions in PAH and explain the great variability in susceptibility to stimuli such as anorexigens, virus, and shunts. Epigenetics may be relevant to the female predisposition to PAH and the incomplete penetrance of BMPR2 mutations in familial PAH.

ARIES‐3: Ambrisentan Therapy in a Diverse Population of Patients with Pulmonary Hypertension

Ambrisentan is an oral, once daily, endothelin receptor antagonist approved for treatment of pulmonary arterial hypertension (PAH). Previous studies of ambrisentan were limited to patients with Group 1 PAH and often excluded patients receiving other pulmonary hypertension (PH) therapies. ARIES-3 was an open-label study evaluating efficacy and safety of ambrisentan in patients with various PH etiologies and background PH medications. Patients received 5 mg ambrisentan once daily for 24 weeks. The primary endpoint was change from baseline in 6-minute walk distance (6MWD) at week 24. A total of 224 patients with PH due to idiopathic and familial PAH (31%), connective tissue disease (18%), chronic hypoxemia (22%), chronic thromboembolic disease (13%), or other etiologies (16%) were enrolled and 53% of patients received stable background PAH therapies. After 24 weeks of therapy, an increase in 6MWD (+21 m; 95% CI: 12-29) and a decrease in B-type natriuretic peptide (-26%; 95% CI: -34 to -16%) was observed in the overall population compared to baseline; however, increases in 6MWD were not observed in several non-Group 1 PH subpopulations. Peripheral edema, headache, and dyspnea were the most common adverse events. This study reconfirms the results of previous placebo-controlled studies, which demonstrate that ambrisentan is well tolerated and provides benefit in patients with PAH. Definitive conclusions regarding the safety and efficacy of ambrisentan in specific non-Group 1 PH etiologies cannot be determined and larger, controlled studies will be necessary to determine the efficacy and safety of ambrisentan in these populations.

Response to Letter Regarding Article, “Elevated Levels of Inflammatory Cytokines Predict Survival in Idiopathic and Familial Pulmonary Arterial Hypertension”

We appreciate the opportunity to discuss the issues raised by Dr Montani et al. The Kaplan-Meier survival curves were presented according to quartile values for each cytokine to determine whether there was a dose-response effect, which is standard practice for this form of analysis. When we dichotomized the data by median values, it also resulted in significant differences in survival, but the separation of curves given by the median values was not as good as that given by the optimum values stated in Table 4 of our article.1 Because it is our intention to validate the use of these cytokines as biomarkers in clinical practice, we believe that presentation of both quartiles and optimum values for each cytokine is the best and …

Pulmonary Arterial Hypertension: Insights from Genetic Studies

Familial pulmonary arterial hypertension (FPAH) was described 60 years ago, but real progress in understanding its origins and pathogenesis is just beginning. Germline mutations in bone morphogenetic protein receptor type 2 (BMPR2) are responsible for the disease in most families, and also in many sporadic cases of idiopathic PAH. Heritable PAH refers to patients with a positive family history, or with a responsible genetic mutation, and is an autosomal dominant disease that affects females disproportionately, may occur at any age, and is characterized by reduced penetrance and variable expressivity. These characteristics suggest that other endogenous or exogenous factors modify its expression. Several different factors have recently been demonstrated to modify the clinical expression of BMPR2 mutation, including estrogen metabolites and functional polymorphisms in transforming growth factor-β1 and CYP1B1. Furthermore, a linkage study recently identified modifier loci for BMPR2 clinical expression, which suggests an oligogenic model. Clinical testing for BMPR2 mutations is available for families with heritable and idiopathic PAH, and is an evolving model of personalized medicine. Variable age of onset and decreased penetrance confound genetic counseling, because the majority of carriers of a BMPR2 mutation will never develop PAH, but often transmit the risk to their progeny.

Hypoxia-inducible factors in human pulmonary arterial hypertension: a link to the intrinsic myeloid abnormalities

AbstractPulmonary arterial hypertension (PAH) is a proliferative vasculopathy characterized by high circulating CD34+CD133+ proangiogenic progenitors, and endothelial cells that have pathologic expression of hypoxia-inducible factor 1 α (HIF-1α). Here, CD34+CD133+ progenitor cell numbers are shown to be higher in PAH bone marrow, blood, and pulmonary arteries than in healthy controls. The HIF-inducible myeloid-activating factors erythropoietin, stem cell factor (SCF), and hepatocyte growth factor (HGF) are also present at higher than normal levels in PAH blood, and related to disease severity. Primary endothelial cells harvested from human PAH lungs produce greater HGF and progenitor recruitment factor stromal-derived factor 1 α (SDF-1α) than control lung endothelial cells, and thus may contribute to bone marrow activation. Even though PAH patients had normal numbers of circulating blood elements, hematopoietic alterations in myeloid and erythroid lineages and reticulin fibrosis identified a subclinical myeloproliferative process. Unexpectedly, evaluation of bone marrow progenitors and reticulin in nonaffected family members of patients with familial PAH revealed similar myeloid abnormalities. Altogether, the results show that PAH is linked to myeloid abnormalities, some of which may be related to increased production of HIF-inducible factors by diseased pulmonary vasculature, but findings in nonaffected family suggest myeloid abnormalities may be intrinsic to the disease process.

The Amiloride Derivative Phenamil Attenuates Pulmonary Vascular Remodeling by Activating NFAT and the Bone Morphogenetic Protein Signaling Pathway

Pulmonary artery hypertension (PAH) is characterized by elevated pulmonary artery resistance and increased medial thickness due to deregulation of vascular remodeling. Inactivating mutations of the BMPRII gene, which encodes a receptor for bone morphogenetic proteins (BMPs), are identified in ∼60% of familial PAH (FPAH) and ∼30% of idiopathic PAH (IPAH) patients. It has been hypothesized that constitutive reduction in BMP signal by BMPRII mutations may cause abnormal vascular remodeling by promoting dedifferentiation of vascular smooth muscle cells (vSMCs). Here, we demonstrate that infusion of the amiloride analog phenamil during chronic-hypoxia treatment in rat attenuates development of PAH and vascular remodeling. Phenamil induces Tribbles homolog 3 (Trb3), a positive modulator of the BMP pathway that acts by stabilizing the Smad family signal transducers. Through induction of Trb3, phenamil promotes the differentiated, contractile vSMC phenotype characterized by elevated expression of contractile genes and reduced cell growth and migration. Phenamil activates the Trb3 gene transcription via activation of the calcium-calcineurin-nuclear factor of activated T cell (NFAT) pathway. These results indicate that constitutive elevation of Trb3 by phenamil is a potential therapy for IPAH and FPAH.

Genetic counselling for pulmonary arterial hypertension: a matter of variable variability

We report three cases which highlight the complex considerations surrounding genetic counselling for pulmonary arterial hypertension (PAH). The first counselee developed PAH symptoms shortly after his daughter's death from PAH and was diagnosed with a delay of 1year. An early diagnosis of familial PAH was established in the second counselee. Oral therapy was initiated immediately, and her functional status has since remained stable. The third counselee was a healthy woman who struggled to cope with her risk for familial PAH, having lost two siblings from the disease. These cases show that incomplete penetrance and variable expression need particular attention during clinical assessment and genetic counselling of heritable PAH patients and family members.

Elevated Levels of Inflammatory Cytokines Predict Survival in Idiopathic and Familial Pulmonary Arterial Hypertension

Elevated Levels of Inflammatory Cytokines Predict Survival in Idiopathic and Familial Pulmonary Arterial Hypertension

Expression of mutant BMPR-II in pulmonary endothelial cells promotes apoptosis and a release of factors that stimulate proliferation of pulmonary arterial smooth muscle cells.

Mutations in the bone morphogenetic protein type II receptor gene (BMPR-II) are the major cause of heritable pulmonary arterial hypertension (PAH). Although both endothelial and smooth muscle cell BMPR-II dysfunction have been seen to contribute to pulmonary hypertension in vivo, little is known about the impact of BMPR-II mutation on the interaction between these two important cell types. We employed adenoviral vectors to overexpress wild type or mutant (kinase-deficient mutation, D485G) BMPR-II in human pulmonary arterial endothelial cells (PAECs). PAECs transfected with mutant BMPR-II demonstrated increased susceptibility to apoptosis. Conditioned media from PAECs transfected with mutant BMPR-II increased the proliferation of pulmonary arterial smooth muscle cells (PASMCs), when compared with conditioned media from PAECs transfected with wild-type BMPR-II. PAECs transfected with mutant BMPR-II released higher levels of TGF-β1 and FGF2 into the conditioned media than the wild-type BMPR-II-transfected cells. Conditioned media from PAECs expressing mutant BMPR-II also showed increased activation of luciferase activity in a TGF-β bioassay. The increased proliferation observed in PASMCs exposed to conditioned media from PAECs expressing mutant BMPR-II was inhibited by neutralizing the antibodies to TGF-β1, or small molecule inhibitors of ALK-5 (SD208) or FGFR1 (SU5402). We conclude that mutation in BMPR-II increases susceptibility to apoptosis of PAECs and leads to secretion of growth factors that stimulate the proliferation of PASMCs. These processes may contribute to the remodeling of pulmonary arteries observed in patients with familial or heritable PAH.

The Maze of Vasodilator Response Criteria

With great interest we read the article of Barst et al. [1]. We agree with their statement that the definition of a positive acute vasodilator response in patients with pulmonary arterial hypertension (PAH) remains controversial. We believe it is very important to minimize this controversy and use uniform criteria when studying the acute pulmonary vasodilator response in patients with PAH. In their article, Barst and coworkers defined acute response criteria based on a 20% or more decrease in mean pulmonary arterial pressure (mPAP) with no decrease in cardiac index, or on a 25% or more decrease in the pulmonary vascular resistance index (PVRI) with no decrease in cardiac index (the latter only in case of an unrestricted shunt at the ventricular or ductal level). We do not recognise these criteria from previously published reports. A review of the literature provides the following definitions of a positive response to acute pulmonary vasodilator tests: An mPAP decrease of 20% or more, no decrease in cardiac index and no change or a decrease in the pulmonary-to-systemic vascular resistance ratio [2, 3] A decrease of more than 20% in both mPAP and PVRI [4, 6] A fall in total pulmonary resistance of more than 30% [7] A decrease in mPAP of 10 mmHg or more, reaching an mPAP of 40 mmHg or less and a normal cardiac output [5]. To our knowledge, the criteria used in the current article have not been published previously. The use of different response criteria in different studies results in different prevalences of acute vasodilator response and different predictive values for identifying long-term responders to calcium-channel blocker treatment and patients with improved survival [5]. Furthermore, as a result, data from these studies cannot be compared directly. For example, the prevalence of acute responders among children with PAH (42%) is reported to be higher than among adults (12–26%) [2, 4, 5]. However, because the criteria used differed between children and adults, this comparison does not hold. We commend Barst and coworkers for their success in comparing the use of different vasodilating agents (inhaled nitric oxide, oxygen, or the combination of the two) for vasodilator testing in children. However, the introduction again of new response criteria prohibits interpretation of these data in the perspective of previously published data. It is not our intention to discourage the quest for new response criteria aimed at reaching better prediction of treatment effect or survival or at expanding its use to broader patient populations other than those with idiopathic PAH (iPAH) or familial PAH (FPAH). However, in view of the ongoing discussion on the optimal definition of acute response criteria, we strongly advocate presenting the use of new criteria in a manner comparative with the use of previously published criteria.

Effective treatments for PAH will require detailed knowledge of molecular aetiology

Therapies for idiopathic pulmonary arterial hypertension will need to address the underlying molecular defects, or they will be incapable of halting progression. Identification of the familial PAH gene, BMPR2, and determinants of penetrance in families with BMPR2 mutation, has provided a mechanism for discovering this underlying aetiology not just in those with BMPR2 mutation, but in all patients. These studies suggest that the fundamental defects are in cytoskeletal organization, energy metabolism, and inflammation, with mode of estrogen metabolism acting as the most significant modifier. A detailed understanding of these defects, and a determination of the correct points for intervention, will be required to produce effective treatments for idiopathic PAH.

Elevated Levels of Inflammatory Cytokines Predict Survival in Idiopathic and Familial Pulmonary Arterial Hypertension

Inflammation is a feature of pulmonary arterial hypertension (PAH), and increased circulating levels of cytokines are reported in patients with PAH. However, to date, no information exists on the significance of elevated cytokines or their potential as biomarkers. We sought to determine the levels of a range of cytokines in PAH and to examine their impact on survival and relationship to hemodynamic indexes. We measured levels of serum cytokines (tumor necrosis factor-alpha, interferon-gamma and interleukin-1beta, -2, -4, -5, -6, -8, -10, -12p70, and -13) using ELISAs in idiopathic and heritable PAH patients (n=60). Concurrent clinical data included hemodynamics, 6-minute walk distance, and survival time from sampling to death or transplantation. Healthy volunteers served as control subjects (n=21). PAH patients had significantly higher levels of interleukin-1beta, -2, -4, -6, -8, -10, and -12p70 and tumor necrosis factor-alpha compared with healthy control subjects. Kaplan-Meier analysis showed that levels of interleukin-6, 8, 10, and 12p70 predicted survival in patients. For example, 5-year survival with interleukin-6 levels of >9 pg/mL was 30% compared with 63% for patients with levels < or = 9 pg/mL (P=0.008). In this PAH cohort, cytokine levels were superior to traditional markers of prognosis such as 6-minute walk distance and hemodynamics. This study illustrates dysregulation of a broad range of inflammatory mediators in idiopathic and familial PAH and demonstrates that cytokine levels have a previously unrecognized impact on patient survival. They may prove to be useful biomarkers and provide insight into the contribution of inflammation in PAH.

Smurf1 ubiquitin ligase causes downregulation of BMP receptors and is induced in monocrotaline and hypoxia models of pulmonary arterial hypertension

Reduced bone morphogenetic protein (BMP) receptor (BMPR) expression and BMP signaling have been implicated in vascular cell proliferation and remodeling associated with pulmonary arterial hypertension (PAH). The low penetrance of the BMPR II disease gene in familial PAH suggests that additional genetic or environmental factors are involved in clinical manifestation of PAH. Smurf1 ubiquitin ligase, together with inhibitory SMAD 6/7, forms a negative feedback loop for the attenuation of BMP signals by downregulating BMPR and signaling molecules and, in addition, functions in the integration of MAPK/Ras mitogenic pathways. The present study found that Smurf1 was significantly elevated in pulmonary arteries of monocrotaline and hypoxia-induced PAH rats. In the pulmonary artery of hypoxia-exposed mice, elevation of Smurf1 and SMAD7 was correlated with reduced expression of BMPR II protein. Over-expression of Smurf1 in cultured cells induced ubiquitination and degradation of BMPR I and II whereas ligase-inactive Smurf1 reduced ubiquitination and elevated their protein levels, thus serving a dominant-negative function. Smurf1-induced receptor degradation was inhibited by both proteasomal and lysosomal inhibitors. Thus, Smurf1 reduces steady-state levels of BMPRs by ubiquitination and subsequent degradation involving proteasomes and lysosomes. Therefore, these results show that Smurf1 induction could be a key event for triggering downregulation of BMP signaling and causing vascular cell proliferation and remodeling in PAH and that abrogating Smurf1 function could be a strategy for PAH therapeutics.

Survival in Patients With Idiopathic, Familial, and Anorexigen-Associated Pulmonary Arterial Hypertension in the Modern Management Era

Novel therapies have recently become available for pulmonary arterial hypertension. We conducted a study to characterize mortality in a multicenter prospective cohort of patients diagnosed with idiopathic, familial, or anorexigen-associated pulmonary arterial hypertension in the modern management era. Between October 2002 and October 2003, 354 consecutive adult patients with idiopathic, familial, or anorexigen-associated pulmonary arterial hypertension (56 incident and 298 prevalent cases) were prospectively enrolled. Patients were followed up for 3 years, and survival rates were analyzed. For incident cases, estimated survival (95% confidence intervals [CIs]) at 1, 2, and 3 years was 85.7% (95% CI, 76.5 to 94.9), 69.6% (95% CI, 57.6 to 81.6), and 54.9% (95% CI, 41.8 to 68.0), respectively. In a combined analysis population (incident patients and prevalent patients diagnosed within 3 years before study entry; n=190), 1-, 2-, and 3-year survival estimates were 82.9% (95% CI, 72.4 to 95.0), 67.1% (95% CI, 57.1 to 78.8), and 58.2% (95% CI, 49.0 to 69.3), respectively. Individual survival analysis identified the following as significantly and positively associated with survival: female gender, New York Heart Association functional class I/II, greater 6-minute walk distance, lower right atrial pressure, and higher cardiac output. Multivariable analysis showed that being female, having a greater 6-minute walk distance, and exhibiting higher cardiac output were jointly significantly associated with improved survival. In the modern management era, idiopathic, familial, and anorexigen-associated pulmonary arterial hypertension remains a progressive, fatal disease. Mortality is most closely associated with male gender, right ventricular hemodynamic function, and exercise limitation.

Prognosis of Pulmonary Arterial Hypertension

The study of rare diseases is limited by just that, their infrequency. Pulmonary arterial hypertension (PAH), for example, has a prevalence of 15 cases per million.1 Although there has been an explosion in knowledge of and therapies for this life-threatening disease over the past decade, most of our insight is based on small studies. The first therapy that was approved by the Food and Drug Administration in 1995, intravenous epoprostenol, was based on the results of an 81-patient trial.2 The most recently approved therapy, inhaled treprostinil, in 2009, was based on the results of a 235-patient trial.3 Similarly, our understanding of the natural history of this disease is based on small observational series. Articles see pp 156 and 164 To further our comprehension of rare diseases, we often turn to “registries,” constructed as multicenter cohorts of patients who have the disease with longitudinal follow-up. Despite the inherent limitations of their observational and uncontrolled nature, which also represent strengths, these cohorts are useful to describe and compare patient characteristics, practice patterns, and outcomes. Observations from such registries can generate hypotheses that subsequently form the basis of further studies. Lastly, such cohorts facilitate the study of the prognostic profile of the disease via the derivation and validation of clinical prediction tools. In this issue of Circulation , data from the 2 of the most important present-day registries in PAH give us the opportunity to better understand the prognosis of PAH, its determinants, and outcomes in the current treatment era. Humbert and colleagues4 share data from the French National Registry, in which 354 consecutive idiopathic, heritable, and anorexigen-associated patients with PAH were enrolled from October 2002 to October 2003. They report 1-, 2-, and 3-year survival rates of 82.9%, 67.1%, and 58.2%, respectively. Sadly, despite the many advances in …