Familial primary pulmonary hypertension is a rare autosomal dominant disorder that has reduced penetrance and that has been mapped to a 3-cM region on chromosome 2q33 (locus PPH1). The phenotype is characterized by monoclonal plexiform lesions of proliferating endothelial cells in pulmonary arterioles. These lesions lead to elevated pulmonary-artery pressures, right-ventricular failure, and death. Although primary pulmonary hypertension is rare, cases secondary to known etiologies are more common and include those associated with the appetite-suppressant drugs, including phentermine-fenfluramine. We genotyped 35 multiplex families with the disorder, using 27 microsatellite markers; we constructed disease haplotypes; and we looked for evidence of haplotype sharing across families, using the program TRANSMIT. Suggestive evidence of sharing was observed with markers GGAA19e07 and D2S307, and three nearby candidate genes were examined by denaturing high-performance liquid chromatography on individuals from 19 families. One of these genes (BMPR2), which encodes bone morphogenetic protein receptor type II, was found to contain five mutations that predict premature termination of the protein product and two missense mutations. These mutations were not observed in 196 control chromosomes. These findings indicate that the bone morphogenetic protein-signaling pathway is defective in patients with primary pulmonary hypertension and may implicate the pathway in the nonfamilial forms of the disease.

We report a large monocentric case series of 82 patients with human immunodeficiency virus-associated pulmonary arterial hypertension (PAH). No germline mutations of the PPH1 gene (bone morphogenetic protein receptor-II) were found in any of the 19 patients tested. PAH was the direct cause of death in 72% of cases. Survival rates of the overall population at 1, 2, and 3 years were 73, 60, and 47%, respectively. Survival was significantly poorer in patients in New York Heart Association functional class III-IV at the time of diagnosis, as compared with those in functional class I-II with respective rates of 60, 45, and 28% versus 100, 90, 84% at 1, 2, and 3 years (p < 0.0001). Subsequently, we analyzed prognostic factors in patients in functional class III-IV. Univariate analysis indicated that CD4 lymphocyte count of more than 212 cells mm(-3), the use of combination antiretroviral therapy (CART), and epoprostenol infusion were related with a better survival. On multivariate analysis only CD4 lymphocyte count was an independent predictor of survival, presumably because CART and epoprostenol infusion were strongly linked in our patient population. These results suggest that patients with severe human immunodeficiency virus-associated PAH should be considered for long-term epoprostenol infusion in association with CART.

This study investigated whether patients developing pulmonary arterial hypertension (PAH) after exposure to the appetite suppressants fenfluramine and dexfenfluramine have mutations in the bone morphogenetic protein receptor 2 (BMPR2) gene, as reported in primary pulmonary hypertension.BMPR2 was examined for mutations in 33 unrelated patients with sporadic PAH, and in two sisters with PAH, all of whom had taken fenfluramine derivatives, as well as in 130 normal controls. The PAH patients also underwent cardiac catheterisation and body mass determinations.Three BMPR2 mutations predicting changes in the primary structure of the BMPR-II protein were found in three of the 33 unrelated patients (9%), and a fourth mutation was found in the two sisters. No BMPR2 mutations were identified in the 130 normal controls. This difference in frequency was statistically significant. Moreover, the mutation-positive patients had a somewhat shorter duration of fenfluramine exposure before illness than the mutation-negative patients, a difference that was statistically significant when the two sisters were included in the analysis.In conclusion, the present authors have detected bone morphogenetic protein receptor 2 mutations that appear to be rare in the general population but may combine with exposure to fenfluramine derivatives to greatly increase the risk of developing severe pulmonary arterial hypertension.

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In a previous report on the effects of the leukocytosis-and lymphocytosispromoting factor (LPF) 1 ofBordetella pertussis on cyclic adenosine 3':5'-monophosphate (cAMP) metabolism, a method for isolation of highly active LPF was presented briefly (1). In the present communication, the results of further studies on the isolation and chemical, immunochemical, and biological properties of LPF are described. It has been found that a single moiety is responsible for the induction in mice of leukocytosis with both lymphocytosis and granulocytosis, histamine sensitization, and hypoglycemia and unresponsiveness to the hyperglycemic effects of epinephrine. In confirmation of our preliminary report (2), LPF has been found to be clearly distinct from the hemagglutinating pili of B. pertussis, the properties of which are also described. Materials and MethodsB. pertussis Cultures. Strain NIH 114 (3779B), which has been utilized in continuing studies in this laboratory was also employed in the present investigations. Lyophilizod samples, obtained through the courtesy of Dr. Charles Manclark, Bureau of Biologics, Bethesda, Md., were reconstituted in sterile distilled H~O and plated on Bordet-Gengou medium (Clinical Sciences Inc., Whippany, N. J.). The 2-day growth at 36°C on one Petri dish was harvested in 10 ml of 2% casamino acids (technical grade; Difco Laboratories, Detroit, Mich.), and 1 ml of the suspension was inoculated into 100 ml of liquid medium in i liter Blake bottles. After 2 days of incubation at 36°C, 10 ml of the seed culture (approximately 5 × 10 x° organJma) was inoculated into 1 liter of liquid medium in Povitsky bottles which were then incubated at 35-36°C. In order to provide maximum surface area for aeration, all bottles were placed in the horizontal position during incubation. In the case of the 100 ml cultures, the depth of the medium was 1.0 cm and the surface area 218 cm2; the values for the 1-1iter cultures were 2.5 cm and 580 cm 2, respectively.The liquid medium was that previously described (3) except for the omission of an anion exchange resin which was found to be unnecessary for either optimal growth, maintenance of the Phase I state, or maximal production of LPF. Culture density was assessed with the National Institutes of Health opacity standard, and the criteria utilized to determine whether the organisms were Phase I included: a homogenous population of small, gram-negative coccobacilli and

Primary pulmonary hypertension (PPH) is a rare but severe and progressive disease characterised by obstructive lesions of small pulmonary arteries. Patients with PPH often have mutations in the bone morphogenetic protein receptor type II (BMPR2) gene, whereas some carry mutations in the activin receptor-like kinase 1 (ALK-1) gene, generally associated with hereditary haemorrhagic telangiectasia (HHT) type 2, a vascular dysplasia affecting multiple organs. The aim of this study was to determine whether members of families with PPH and confirmed or probable HHT had ALK-1 mutations.ALK-1 and BMPR2 mutation analysis was performed on deoxyribonucleic acid from affected members of four families with PPH and confirmed or suspected HHT.ALK-1 mutations were identified in all four families and three novel mutations found in exon 10, leading to truncated proteins. In the fourth family, a missense mutation, previously reported in four independent HHT families, was detected in exon 8. Analysis of the BMPR2 gene revealed no exonic mutations in the probands with both PPH and HHT.The present data bring to 10 the number of reported families with primary pulmonary hypertension and hereditary haemorrhagic telangiectasia type 2, representing 16% of the 61 families with known activin receptor-like kinase 1 mutations. Such mutations might predispose to primary pulmonary hypertension, and specialists should be aware of the potential link between these two disorders. Primary pulmonary hypertension (PPH) is a rare disease with an estimated incidence of 1-2 cases per million population [1]. The presenting symptoms usually include fatigue, anorexia and shortness of breath, which, if left untreated, lead to a progressive increase in pulmonary arterial pressure, right ventricular failure and death [2]. The affected small pulmonary arteries and arterioles are characterised by intimal proliferation, medial hypertrophy, concentric fibrosis and the presence of plexiform lesions composed of both vascular smooth muscle cells and endothelial cells [3]. Monoclonal endothelial cell proliferation is found in the plexiform lesions of PPH but not in secondary pulmonary hypertension [4].Recently, PPH has been found to be caused by mutations in either of two genes: the bone morphogenetic protein receptor type II gene (BMPR2) [5][6][7] and the activin receptor-like kinase 1 gene (ACVRL1 or ALK-1) [8], both members of the transforming growth factor-b (TGF-b) receptor superfamily. A recent abstract reported an endoglin gene (ENG) mutation in a patient with HHT and dexfenfluramine-associated PPH [9]. ALK-1 and ENG are the two genes associated with hereditary haemorrhagic telangiectasia (HHT). Mutations in ALK-1 lead to HHT type 2 (HHT2) [10,11], whereas mutations in ENG are responsible for HHT type 1 (HHT1) [12]. HHT is an autosomal dominant vascular disorder that occurs at an incidence of w1 in 10,000 population. It is characterised by vascular dysplasia with the formation of mucocutaneous telangiectases and arteriovenous malformations (AVMs) in ...

Mutations of bone morphogenetic protein receptor type II (BMPR-II) have been associated with familial and idiopathic pulmonary arterial hypertension (PAH). BMPR-II is a member of the transforming growth factor-␤ receptor superfamily. It consists of extracellular, transmembrane, and kinase domains, and a unique C-terminus with mostly unknown function. However, a number of PAH-causing mutations are predicted to truncate the C-terminus, suggesting that this domain plays an important role in the homeostasis of pulmonary vessels. In this study, we sought to elucidate the functional role of this C-terminus by seeking its interacting partners. Using yeast two-hybrid screening, we identified c-Src tyrosine kinase as a binding partner of this C-terminus. In vitro co-immunoprecipitation confirmed their interaction. Mutations truncating the C-terminus disrupted their interaction, while missense mutation within kinase domain reduced their interaction. In addition, BMPR-II and c-Src tyrosine kinase colocalized within intracellular aggregates when overexpressed in HEK293 cells. Moreover, mutations truncating the C-terminus disrupted their colocalization, whereas missense mutation within kinase domain had no effect on their colocalization. Keywords: bone morphogenetic protein receptor type II; c-Src tyrosine kinase; pulmonary arterial hypertension Bone morphogenetic protein receptor type II (BMPR-II) is a member of the transforming growth factor-␤ (TGF-␤) receptor superfamily. Members of the TGF-␤ receptor superfamily include type I and type II receptor proteins, and play a complex and multifunctional role in the regulation of cell proliferation, differentiation, and apoptosis during embryogenesis and throughout adult life (1). Homozygous BMPR-II knockout mice die very early in development during gastrulation (2), while mice that express a mutant BMPR-II, lacking half of the ligandbinding domain, undergo normal gastrulation, but die at midgestation with cardiovascular and skeletal defects (3).

Cosegregation of this region with disease in different ethnic groups suggests that we mapped an important locus in familial PPH. Careful study of additional families and sporadic cases will be required to confirm this localization of PPH1 and characterize its overall role.