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* From the Department of Medicine, Columbia University College of Physicians and Surgeons, New York, NY.
Correspondence to: Jane H. Morse, MD, Department of Medicine, Columbia-Presbyterian Medical Center, PH 8 East, Suite 101, 630 West 168th St, New York, NY 10032
Key Words: appetite suppressants • bone morphogenetic protein receptor 2 • genetics • primary pulmonary hypertension
This presentation on the genetic aspects of primary pulmonary
hypertension (PPH) concentrates on our experience at Columbia
University and also includes brief summaries of unpublished
observations on risk factors for PPH. Familial PPH (FPPH), is inherited
as an autosomal dominant with incomplete penetrance.1
2
3
Sporadic PPH occurs in one to two cases per million; the familial form
occurs in 6% of the 187 cases in the National Institutes of Health
registry.4
Microsatellite marker studies followed by
linkage analyses localized a gene for the FPPH to chromosome
2q31-322
3
and was designated PPH1.2
Subsequently, PPH1 was finely mapped to a more telomeric 3-centimorgan
region at 2q33.5
The existence of other PPH loci was not
systematically excluded. The two original gene scans applied 
220
autosomal on fewer than eight families (one family shared). A German
group, whose presentation follows, finds that stress echocardiograms
may identify a second chromosomal locus, more centromeric to PPH1 but
within the original approximately 25-centimorgan PPH1 locus, which they
call PPH2.
During the past year, four articles6 7 8 9 reported heterozygous mutations in bone morphogenetic protein receptor (BMPR) 2 cause PPH.6 7 8 9 The Columbia group found nine mutations in 19 FPPH families,6 and the International Consortium found seven mutations in 8 FPPH families.7 Subsequently, the International Consortium reported that approximately 50% (23 of 47 FPPH families) had BMPR2 mutations.8 Our unpublished observations found a total of 18 different mutations (20, including two additional families with the same single-site mutation) in exons 2, 3, 4, 6, 8, 9, 11, and 12. BMPR 2 mutations have also been reported in 26% (13 of 50 patients) with sporadic PPH.9 Qualitatively, the mutations are missense, nonsense, and frameshift. These mutations were not found in > 350 normal chromosomes.6 7 8 9
BMPR2, originally cloned by four independent laboratories in 1995,10 11 12 13 is a member of the transforming growth factor (TGF)-ß–receptor superfamily. It has 13 exons. Exons 1 to 3 encode an extracellular domain, exon 4 encodes a transmembrane domain, exons 5 to 11 encode the serine/threonine kinase domain, and exon 12 encodes a very large intracellular C-terminal domain of unknown function that is unique to BMPR2. The protein start codon is in exon 1, and the stop condon in exon 13. To date, mutations have been reported in all of the exons except 5, 10, and 13. Polymorphisms have been reported in exons 6, 8, and 12.8 As noted in the earlier mapping studies, each mutation has been private to a family. Although there can be the same nucleotide change in different families, the haplotypes constructed by microsatellite markers, were private to each family.
New mutations (mutations not present in either parent) were found in affected individuals from families with sporadic PPH9 and FPPH.6 In the FPPH family, an arginine was changed to glutamine in the proband. Importance was attached to this mutation because it was not found in her parents, suggesting that a new mutation in BMPR2 might cause sporadic cases of PPH.6 This finding was substantiated in the survey of 50 unrelated patients with sporadic PPH, where at least two families had mutations not present in either parent.9 The 26% frequency of BMPR2 mutations in sporadic PPH is higher than the 6% frequency found in the National Institutes of Health registry.4
Both groups noted that each of the amino-acid substitutions occurred in either a highly conserved or a functionally important site of the BMPR2 molecule. The potential to disrupt function depended whether the mutation affected the extracellular (exons 2, 3), transmembrane (exon 4), serine/kinase (exons 5 to 11) domains, or the cytoplasmic tail (exon 12). Certain of the mutations appeared to be in an amino acid, which is conserved in all known type II TGF-ß superfamily receptors. For example, arginine491 in exon 11, mutated in three Columbia families, is important because it is conserved in all known type II TGF-ß superfamily receptors. In addition to BMPR-II (man, mouse, Xenopus), these receptors include TGF-receptor II (man, mouse, rat), and other receptor type II proteins ActR-II, AMHR-II (rat), DAF-4 (Caenorhabditis elegans), and XSTK3 (Xenopus).
Of interest, mice homozygous for a mutation in the kinase domain of BMPR2 do not develop past gastrulation, whereas the heterozygous mouse appeared to be normal.14 This suggests that BMPR2 has varying roles during the normal life span of an animal. Mechanistically, haploinsufficiency has been found using BMPR2 mutations in in vitro expression systems.8 However, a dominant negative mechanism has not been formally excluded.
Both groups extrapolated from in vitro experiments of the TGF-ß pathway that BMPR2 mutations would interrupt the bone morphogenetic protein (BMP) signaling pathway by hindering ligand binding, eliminating kinase activity, or reducing heterodimeric dimer formation. The BMP/TGF-ß signaling pathway is extensively reviewed.15 16 Briefly, a BMP or TBF-ß ligand binds heteromeric complexes of type I and type II receptors. The type II receptors phosphorylate type I receptors, which in turn activate specific Smads by phosphorylation. The TGF-ß type I and type II receptors bind Smads 2 and 3, whereas the complex of BMP receptors, BMPR1a, BMPR1b, and BMPR2, bind and phosphorylate Smads 1, 5, and 8. The two types of complexes are translocated to the nucleus via Smad 4. The inhibitory Smads 6 and 7 can inhibit these interactions. Normally, the TGF-ß family of signaling molecules inhibits endothelial proliferation and modulates proteins involved in cell-cycle control and angiogenesis.15 16 17
The exact signaling scenario, and whether endothelial, smooth-muscle, and adventitial cells are all involved, remains to be determined for PPH. TGF-ß is upregulated in remodeling arteries in PPH.18 An insult such as shear stress can induce changes via a mechanotransduction pathway involving changes in K+ and Ca+ ion currents via the mediator TGF-ß1.19 In addition, Smads 6 and 7 are induced by shear stress and can modulate gene expression in response to both humoral and biomechanical stimulation and induce changes in vascular endothelium.20
The International Consortium also suggested that there could be a loss of the normal allele in the diseased tissue. By analogy, this resembles certain types of colon cancer, where there is microsatellite instability and a germline mutation in the kinase domain of TGF-ß type II predisposes to colorectal cancer. In fact, juvenile hereditary polyposis has mutations of Smad 421 and BMPR1a.22 Yeager et al23 reported microsatellite instability within the endothelial cells harvested from PPH plexiform lesions. Specifically, the isolated endothelial cells had microsatellite mutations and reduced protein expression of TGF-ß–receptor type II (6 of 19 lesions) and Bax 4 (4 of 19 lesions), akin to neoplasia. Geraci et al24 used gene microarray analyses to compare the oligonucleotide microarray expression patterns found in FPPH and PPH to those in normal lung. As anticipated, there was an imbalance between cell proliferation and apoptosis genes in pulmonary hypertension compared to normal lung. Surprisingly, alterations in the expression patterns of TGF-ß, BMP2, mitogen-activated protein kinase 5, receptor for activated C-kinase 1, apolipoprotein C-III, and laminin receptor I were overexpressed or underexpressed in tissues from sporadic PPH but not in FPPH tissues. These two articles did not state whether BMPR2 mutations were present in the FPPH and PPH histologic samples. Subsequent presentations by these two authors23 24 provided new information, especially on the differences between sporadic PPH and FPPH. BMPR-II and TGF-ß type 1 and type II receptors were localized immunohistologically to endothelial cells and myofibroblasts within the obliterative lesions of PPH lesions (10 sporadic PPH and 4 FPPH) by a British group.25
The failure to find BMPR2 mutations in all of the FPPH families, the 26% frequency of BMPR2 mutations in sporadic PPH, and the analogy to hereditary hemorrhagic telangiectasia (HHT) suggests that other genes remain to be found. As more genes are identified, classification by genetic profile is likely to supplant the present clinical classification.
The original reports also compared PPH to HHT or Osler-Rendu-Weber (ORW) syndrome, a rare autosomal dominant disorder, characterized by angiodysplastic lesions (telangiectasias and arteriovenous malformations) that affect many organs. Mutations in two related participants in the TGF-ß signaling pathway, endoglin26 and an activin receptor-like kinase (ALK) 127 were found previously in HHT. Endoglin, a TGF-ß type III receptor protein, is on chromosome 9q34 (ORW1); ALK1, a TGF-ß type 1 receptor is on chromosome 12 (ORW2). ORW families have a predisposition for the development of pulmonary arteriovenous malformations associated with pulmonary hypertension, usually mutations in endoglin. A recent abstract28 reported that three novel germline heterozygous ALK1 mutations were found in three HHT families who also had the clinical and histologic findings of PPH. These findings were published in more detail a month after the conference.29
The next section highlighted the results of the BMPR2 mutations found in several clinical entities with pulmonary hypertension and risk factors for PPH. The summaries are brief because the data are unpublished or have been recently submitted for publication. The World Health Organization 1998 Pulmonary Hypertension Symposium accepted two exogenous factors, HIV infection and the use of anorexic agents, as causes of PPH.30
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HIV Infection and PPH |
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 TOP HIV Infection and PPH Appetite-Suppressant PPHPulmonary Hypertension in...References |
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Appetite-Suppressant PPH |
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TOPHIV Infection and PPHAppetite-Suppressant PPHPulmonary Hypertension in...References |
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Pulmonary Hypertension in Gaucher’s Disease |
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TOPHIV Infection and PPHAppetite-Suppressant PPHPulmonary Hypertension in...References |
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Oral contraceptives and hormone replacement therapy (HRT) were not considered to be risk factors for PPH by the World Health Organization PPH Symposium Committee on Risk Factors and Associated Conditions,30 but this topic needs further study in those at risk for FPPH.31 32 We decided to reinvestigate these potential risk factors, because an asymptomatic postmenopausal known carrier of FPPH had PPH develop after 6 months of HRT. Findings on chest radiography, ECG, and echocardiography were normal 2 months prior to HRT. Eight months later, she was short of breath, and chest radiography, ECG, echocardiography, and right-heart catheterization confirmed PPH.32 As this could be an anecdotal case, we compared via a retrospective phone survey the use of these two agents in asymptomatic PPH1-positive gene carriers to those with PPH. PPH1 positivity was determined using microsatellite markers, as the survey predated the discovery that BMPR2 was PPH1. More patients with FPPH than asymptomatic gene-positive carriers used oral contraceptives and HRT. The comparisons were statistically significant despite the retrospective nature of the study, the small number of subjects, and the inability to quantitate drug preparation and duration of use.
Lastly, the PPH gene has been postulated to influence fertilization or development in utero, as suggested by the observation that fewer male patients are born in FPPH families than in the normal population.33 In our first 33 FPPH families, we also noted nine sets of fraternal twins in 7 families, three sets of boys, two sets of girls, and four boy/girl sets.31 Eight sets had at least one or more affected twin, and the ninth set had an asymptomatic gene-positive carrier (microsatellite markers). BMPR2 mutations were subsequently found in two of the seven FPPH families. The series was too small to make further clinical or genetic conclusions, and we have not systematically studied newer FPPH families. The BMP signaling pathway is involved in reproduction, but whether there is an increase in multiple births in FPPH and whether BMPR2 mutations are relevant remain to be investigated.
The presentation by Dr. Loyd discussed the value of genetic testing and counseling. We would like to emphasize that the identification of asymptomatic gene carriers is beginning to provide clinical and genetic information,34 and that the future may provide therapeutic modalities tailored to the individual’s genetic profile.
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Acknowledgements |
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Footnotes |
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Supported by grant HL-60056 from the National Institutes of Health-National Heart, Lung, and Blood Institute.
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References |
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TOPHIV Infection and PPHAppetite-Suppressant PPHPulmonary Hypertension in...References |
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This article has been cited by other articles:
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  G. Simonneau, N. Galie, L. J. Rubin, D. Langleben, W. Seeger, G. Domenighetti, S. Gibbs, D. Lebrec, R. Speich, M. Beghetti, et al. Clinical classification of pulmonary hypertension J. Am. Coll. Cardiol., June 16, 2004; 43(12_Suppl_S): 5S - 12S. [Abstract] [Full Text] [PDF]
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