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 Richard
C Trembath,
MD
Professor of Medical Genetics
Division of Medical Genetics
Departments of Genetics adn Cardiovascular Science
University of Leicester
Leicester, United Kingdom
Introduction
Substantial progress in delineating the molecular and genetic
basis of pulmonary arterial hypertension has placed the TGFbeta
cell-signaling pathway as the centerpiece of contemporary
thinking about the pathogenesis of this disorder. Although identification
of deleterious mutations within the bone morphogenetic
protein receptor type II (BMPR2) gene provides a compelling
basis for implicating altered TGF-beta signaling in pulmonary
arterial hypertension, the precise molecular mechanisms
of disease development remain obscure. However, these
findings already present important clinical implications, not
least of which is the recognition of disorders in which pulmonary
arterial hypertension should now be considered a potential
and important complication. In this article, emerging features
of the association of pulmonary arterial hypertension and
the inherited vascular disorder known as hereditary hemorrhagic
telangiectasia (HHT) are described. Recognition of this relationship
not only serves to expand the clinical phenotype of
HHT,1but the findings emphasize the importance of clinical
observation in assisting the elucidation of the molecular basis
of human disease, in this case the enigmatic disorder we term
pulmonary arterial hypertension. Finally, the clinical association
of HHT and pulmonary arterial hypertension challenges cellular
biologists who now seek to provide the molecular detail of the
processes that alter normal structure and function of the pulmonary
vasculature.
HHT, or Rendu-Osler-Weber syndrome, describes an autosomal
dominant disorder characterized by vascular malformations
affecting both cutaneous and internal structures, which frequently
bleed (Figure 1).2 Hence, the typical symptomology of
HHT includes recurrent epitaxis, fatigue, and exertional dyspnea
due to chronic blood loss from the gut, together with more
complex complications arising from larger arteriovenous malformations
that may involve the pulmonary, cerebral, or hepatic
vasculature.
 Figure
1. A. Cutaneous telangiectasia characteristic
of hereditary
hemorrhagic telangiectasia (HHT). B. Electron microscopic appearance
of arteriovenous communication in HHT.
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By comparison to the paucity of information
regarding the development of the characteristic obstructive
and plexigenic
lesions of pulmonary arterial hypertension, substantial
detail of the emergence of the vascular lesions in HHT is
available. This includes systematic electron microscopy
studies of biopsied lesions during the evolution of the classic
cutaneous telangiectasia, which ultimately leads to loss
of the capillary bed intervening between arterioles and
venules (Figure 1B).3 Intriguingly, although focal, the initial
observation is one of venodilation, mononuclear (lymphocytic)
infiltration, and vessel wall thickening, due to pericyte
recruitment. Subsequently arteriole dilatation is observed
with a loss of the capillary bed, replaced by up to four direct
arteriovenous connections, with consequences for the local
flow dynamics. Although these changes appear to reflect
fundamental developmental defects, arteriovenous malformations,
and more generally telangiectasias, do not typically
appear clinically until early teenage life and hence may reflect
more prolonged exposure to hemodynamic influences.
HHT as a genetic syndrome and a vascular disorder has
risen onto the horizon of those interested in pulmonary arterial
hypertension. This has occurred because of the recognition
of a group of patients who develop classical features of
pulmonary arterial hypertension, including in some cases
histopathological evidence of both plexiform and concentric
lesions, against a background of clinical HHT or due to
mutations in the genes causative of HHT. These are
described in detail later in this article, but first it is appropriate
to describe some of the genetic and molecular features
of HHT, to enable comparison and contrast with
emerging views of the pathogenesis of pulmonary arterial
hypertension.
Molecular
Genetic Basis of HHT
Early linkage studies of extended HHT kindreds provided
compelling evidence for at least two distinct locations
for
HHT genes within the human genome. Using positional and
candidate gene cloning strategies, HHT1, which maps to
chromosome 9q33-34, was shown to encode the type III or
accessory receptor protein termed endoglin (ENG).4 The
HHT2 locus, which maps to chromosome 12q13, was shown
to be a gene that encodes a further member of the TGF-beta
superfamily of cytokines, known as activin A receptor, type
II-like kinase 1 or activin receptor-like kinase-1 (ACVRL1 or
ALK1).5 The more general features of TGF-beta mediated
signaling are described below, but of importance, transcripts
from both ENG and ALK1 are expressed predominantly in
endothelial cells. The majority of ENG and ALK1 mutations
appear to lead to loss of function.6 As an autosomal dominant
condition, the presence of a heterozygous (single)
mutant allele appears sufficient to predispose to the development
of the classic vascular anomalies; however, these are
site specific rather than generalized, suggesting that other
events, either genetic or environmental, are necessary to
switch the normal equilibrium of vascular cells from a maintenance
state. To date, no evidence for a second genetic hit
of either gene has been brought forward, although, and in
contrast to tumor formation, the vascular process of telangiectasia
formation and arteriovenous malformations is one of
remodeling rather than a predominant state of cell proliferation.
Molecular analysis of site-specific lesions in HHT
under such conditions may therefore be less than ideal for
looking, for example, for events such as loss of heterozygosity.
Interestingly, early studies suggested a phenotype-genotype
correlation with an increased predisposition to pulmonary
arteriovenous malformations among the HHT1
(ENG) group of subjects. Beyond this, precise functions for
either ENG or ALK1 in vascular processes remain unclear.
However, insight will further emerge through the detailedanalysis of animal models
harboring defects of these genes.
TGF-beta signaling in vascular disease
BMPR2, ENG, and ALK1 are membrane-bound receptor
members of the TGF-beta superfamily that form structurally
related polypeptides that regulate a number of biological
processes that include cell cycle control, embryogenesis,
growth, development, and differentiation of cell types. The
general and most studied model of TGF-beta signaling
requires ligand binding with a constitutively phosphylated
type II receptor. This process enables recruitment of the type
I receptor into the activated complex, with the intracellular
kinase domain of the type II receptor subsequently phosphorylating
the type II receptor at serine and threonine
residues, located in the cytoplasmic GS-domain. Once activated,
the type I receptor phosphylates members of the
SMAD family of intracellular mediators, which act as cell
type-specific transcription factors. Specificity for response is
at least in part determined by the components of the cascade
that are activated. For example, interaction of the type
I receptor ALK1 leads to phosphorylation of receptor SMAD
1, 5, and 8. Less well characterized SMAD independent
pathways, together with SMAD dependent processes, may
be further regulated by the expression of type III receptors
such as endoglin.7
 Figure
2. Line diagram of classic components of TGF-beta
mediated cell signaling pathway, illustrating the potential
cross-talk between the type II receptor BMPR2 and the type
I receptor ALK1, modulated by the accessory receptor endoglin.
The balance between activation of these pathways is considered
to regulate progression or resolution of angiogenesis,
which may contribute to the clinical features of vascular
remodeling.
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Despite the lack of evidence for a direct
interaction between BMPR2 (the familial pulmonary arterial
hypertension gene) and ALK1/endoglin complexes, there is
extensive cross-talk between intracellular signalling pathways activated
by these receptor complexes.8 For example,
signalling via ALK1 and BMP receptors occurs through the
same set of receptor-regulated SMAD proteins that complex
with the comediator SMAD 4. This allows transolaction into
the nucleus and transcriptional regulation of a restricted
range of genes in both a cell type
and context-specific manner
(Figure 2). Hence, we might anticipate that elucidation
of the impact of mutations in either BMPR2 or ALK1 on
these downstream pathways will point to molecular mechanisms critical
to the development of pulmonary arterial hypertension.
Pulmonary Arterial Hypertension in HHT
 Figure
3. Hemodynamic determinants of mean pulmonary
arterial
pressure (mPAP). A. Raised mPAP due to high flow through
large
arteriovenous malformations (AVMs) leading to raised
cardiac output
(CO) but typically normal pulmonary vascular resistance
(PVR) and
pulmonary capillary wedge pressure (PCWP). B. Contrast
to raised
mPAP due to raised PVR from obstructive pulmonary vascular
lesions
in HHT-related pulmonary arterial hypertension.
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Abnormalities of vascular pressure-flow dynamics have long
been recognised in HHT. The development of large arteriovenous
malformations has the potential to significantly
impact on cardiac output and peripheral resistance as well
as to generate significant shunting of blood flow, particularly
within the pulmonary vasculature. The hemodynamic consequences
of these clinical scenarios are illustrated in
Figure 3.
The association of HHT with familial pulmonary arterial
hypertension was first reported in detail in 2001.9 Six mutations
and 10 cases of pulmonary arterial hypertension
were identified in five families with HHT and one in one
individual with no available family history. In one of the families
with an extended HHT kindred, three young relatives
(<7 years age) each developed rapidly progressive and fatal
pulmonary arterial hypertension, the proband surviving a few
years following heart-lung transplantation (Figure
4).
Histology of the explanted lung revealed obliterature and
plexiform lesion, and investigations of this child and the
affected siblings revealed evidence of significantly raised
pulmonary vascular resistance (Figure 4). Intervening adult
relatives exhibited typical features of HHT, which was present
in only one of the three children with pulmonary arterial
hypertension. Hence, these subjects have HHT-related pulmonary
arterial hypertension, characterised by a significantly
raised pulmonary artery pressure, high pulmonary vascular
resistance, normal pulmonary arterial wedge pressure,
and a normal or low cardiac output. Molecular studies of this
and additional kindreds revealed pathogenic heterozygous
mutations of the ALK1 gene (Figure 5).10
 |
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Figure 4. Family A reported in detail
in Trembath et al.9 Pedigree
shows diversity of vascular phenotypes consequent upon
mutation of
ALK1 segregating within the kindred.
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Figure 5. Diagram of ALK1 cDNA with
exons encoding protein functional
domains. Location of previously reported mutations in classic
HHT (above) and all ALK1 mutations reported to date in
HHT-related
pulmonary arterial hypertension (below). Note potential
clustering
within the so-called NANDOR box, a region from codons 479
to 489
apparently necessary for TGF-beta signaling regulation.10
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Additional reports of patients with HHT-related
pulmonary arterial hypertension have subsequently been published.
Eight missense mutations of ALK1 were reported in
11 probands, one of which was observed in two families.11 Four of the eight mutations were novel. In addition, heterozygous
mutations of ENG were identified in two subjects,
each representing a novel frameshift mutation and predicted
to lead to premature truncation of the transcript. None of the
mutations was identified among appropriate matched control
groups. More recently, ALK1 mutations were identified
in four additional families with HHT.12 Three novel mutations
in exon 10 leading to truncated proteins were found.
In the fourth family, a missense mutation previously reported,
was identified.
The potential for the development of lesions characterised
by either vessel dilation or pulmonary hypertension
caused by concentric obstruction and abnormal remodeling,
seems at first sight counterintuitive. The recognition that the
genes involved in familial pulmonary arterial hypertension
and HHT are so closely related and may function in common
cell signaling pathways, likely holds the clues to the clinical
overlap. This report now implicates mutations of ENG in the
pulmonary arterial hypertension process; however, the clinical
consequences may be a little more complex. A germline
mutation of ENG was identified in a woman with HHT and
pulmonary arterial hypertension, who has also been exposed
to anorexic drugs, which are also known to predispose to the
development of pulmonary arterial hypertension. We have
observed that patients with pulmonary hypertension and
ENG mutations underlying HHT typically have substantially
reduced pulmonary vascular resistance due to arteriovenous
malformations and formation of high flow shunts. We have
recently reported a child with an ENG mutation who presented
at a very young age (<3 months old) with evidence of
substantially increased pulmonary vascular resistance. As
this child was followed over time, it was of considerable
interest to note that the pulmonary vascular resistance gradually
decreased, a change likely to be associated with the
subsequent development of microvascular pulmonary arteriovenous
malformations. In essence, this likely represents
the clinical transition from the hemodynamic response seen
in Figure 3B to the pressure flow response depicted in
Figure 3A.
Conclusion
Overall, the findings of germline mutations in the ALK1 and
rarely the ENG gene, in pulmonary arterial hypertension
raise key points. First, of clinical relevance, these recent
findings show that pulmonary arterial hypertension is an
uncommon yet serious presentation of the inherited disorder
HHT even in childhood and can be the initial presentation
of HHT within a kindred. Hence, detailed family history and
critical clinical examination of apparently healthy parents is
warranted, looking for the subtle manifestation of HHT. For
example, we have observed at least two subjects with no personal
or family history of HHT, who have presented with pulmonary
arterial hypertension due to de novo ALK1-mediated
disease.
In comparison, clinical features at presentation,
particularly in young children, failed to distinguish between
patients with mutations in the different genes or those without
identified mutations. Hence, clinicians need to consider
the possibility of germline mutation of TGF-beta receptor
genes in patients presenting with pulmonary arterial hypertension
across all age groups. In terms of molecular mechanisms,
it is already apparent that substantial overlap exists
between those mutations of ALK1 associated with HHT and
those that have been identified in patients with HHT-related
pulmonary arterial hypertension (Figure 5). We interpret
these findings as providing further evidence of a role for
additional genetic and/or environmental factors that influence
both the age of onset or presentation with pulmonary
arterial hypertension in subjects with ALK1 mutations.
Taken together, these recent genetic studies have demonstrated
that pulmonary arterial hypertension is an important
and not infrequent complication of ALK1-mediated disease.
HHT should also be considered, and if appropriate screened
for, in patients presenting with apparent idiopathic pulmonary
arterial hypertension.
Acknowledgements
Work described in this article has been undertaken with
financial support from the British Heart Foundation (RCT
Programme Grant with Dr Nick Morrell, Cambridge,
England, and Clinical Fellowship to Dr Rachel Harrison). I
wish to recognize the support and collaboration of many
clinical colleagues, who have generously introduced patients
to these research studies and provided clinical details following
informed consent.
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