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Mouse Model for Hereditary Hemorrhagic Telangiectasia Has a Generalized Vascular Abnormality
Abstract
Mutations in endoglin or activin like kinase-1, both involved in the endothelial transforming growth factor-beta signaling pathway, cause the autosomal dominant bleeding disorder hereditary hemorrhagic telangiectasia. We and others have reported mouse models for this disease that share the characteristic phenotype of dilated vessels and sporadic hemorrhage. The reasons for the variable phenotype in hereditary hemorrhagic telangiectasia are not understood.
After a detailed immunohistochemical analysis of 129/Ola mice, which are heterozygous for a targeted deletion in the endoglin gene, we observed intrinsic abnormalities in the vascular walls throughout the cutaneous vasculature. Postcapillary venules were dilated, and up to 70% of the vascular wall had no smooth muscle cells. The supporting layers of collagens and elastin were irregular, with thin areas, adding to the fragility of these vessels. A variable hemorrhagic phenotype was observed in which local bleeding is associated not only with fragile vessels but also with regions of inflammation.
These findings have relevance to our understanding of the molecular basis of vascular integrity in a wide range of diseases.
... A three-event hypothesis suggesting concurrence of two additional, regionally confined triggers, is mainly based on insights gained from animal studies. First, mice that were heterozygous in either one of the HHT causing genes, presented very mild HHT-like features, at a penetrance below 50% [40][41][42]. This suggested that loss of one functional allele in HHT-genes is not enough for the development of AVMs. ...
... One of the additional triggers proposed was a local complete loss of protein, or loss of heterozygosity. Mice with blepharitis that were heterozygous in the HHT genes endoglin (ENG) had more pronounced hemorrhagic lesions in the eyelids [42]. In an inflammatory environment, the extracellular domain of endoglin is cleaved off and released to bind to circulating ligands [43]. ...
... Subsequently, mice with one functional and one deleted allele in either one of the above gene loci were generated [40][41][42]. Resulting heterozygous Eng +/− and Alk1 +/− mice developed HHT-like vascular lesions between seven and twenty months, and three and twenty-four months, respectively. However, these studies reveal that lesions occurred in less than half of the animals, and in an unpredictable fashion. ...
Arteriovenous malformations are a vascular anomaly typically present at birth, characterized by an abnormal connection between an artery and a vein (bypassing the capillaries). These high flow lesions can vary in size and location. Therapeutic approaches are limited, and AVMs can cause significant morbidity and mortality. Here, we describe our current understanding of the pathogenesis of arteriovenous malformations based on preclinical and clinical findings. We discuss past and present accomplishments and challenges in the field and identify research gaps that need to be filled for the successful development of therapeutic strategies in the future.
... Mice that are heterozygous for mutations in Acvrl1 or Eng develop vascular abnormalities that resemble the clinical pathology of HHT (Bourdeau et al. 1999;Srinivasan et al. 2003;Torsney et al. 2003). These phenotypes are, like in patients, quite variable and mild, occur with a late onset, and depend on the genetic background (observed in 129/Ola background only), suggesting that additional factors are required for HHT development, such as genetic modifiers or environmental triggers (Bourdeau et al. 1999;Torsney et al. 2003). ...
... Mice that are heterozygous for mutations in Acvrl1 or Eng develop vascular abnormalities that resemble the clinical pathology of HHT (Bourdeau et al. 1999;Srinivasan et al. 2003;Torsney et al. 2003). These phenotypes are, like in patients, quite variable and mild, occur with a late onset, and depend on the genetic background (observed in 129/Ola background only), suggesting that additional factors are required for HHT development, such as genetic modifiers or environmental triggers (Bourdeau et al. 1999;Torsney et al. 2003). Angiogenesis has been proposed to act as an additional trigger or secondary hit, because Eng þ/2 and Acvrl1 þ/2 mice injected with VEGF-expressing adenoviral particles show increased vascular dysplasia (Xu et al. 2004;Hao et al. 2010). ...
... Finally, chronic lipopolysaccharide administration to Bmpr2 þ/2 mice induces an exacerbated inflammatory response and pulmonary hypertension develop-ment with elevated IL-6 levels in the lungs and circulation (Soon et al. 2015). Inflammation has also been proposed to be a trigger for HHT, because Eng þ/2 mice with blepharitis have more pronounced bleeding lesions (Torsney et al. 2003). Lipopolysaccharide was also shown to trigger the formation of skin AVMs ). ...
It is well established that control of vascular morphogenesis and homeostasis is regulated by vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF), Delta-like 4 (Dll4), angiopoietin, and ephrin signaling. It has become clear that signaling by bone morphogenetic proteins (BMPs), which have a long history of studies in bone and early heart development, are also essential for regulating vascular function. Indeed, mutations that cause deregulated BMP signaling are linked to two human vascular diseases, hereditary hemorrhagic telangiectasia and pulmonary arterial hypertension. These observations are corroborated by data obtained with vascular cells in cell culture and in mouse models. BMPs are required for normal endothelial cell differentiation and for venous/arterial and lymphatic specification. In adult life, BMP signaling orchestrates neo-angiogenesis as well as vascular inflammation, remodeling, and calcification responses to shear and oxidative stress. This review emphasizes the pivotal role of BMPs in the vascular system, based on studies of mouse models and human vascular disorders.
... These genotypes were lethal at mid-gestation, due to abnormal yolk sac vasculature, angiogenesis, and cardiac development. The heterozygous Eng +/- [80,83] and Alk1 +/- [84] mice share the phenotype of epistaxis, mucocutaneous telangiectasias, and AVMs with HHT patients. ...
... An important observation in the Eng +/mice was the decrease in vascular smooth muscle cells (VSMCs) and abnormal extracellular collagen and elastin matrix [83]. This is consistent with previous studies of the Alk1 -/embryos, in which abnormal recruitment and differentiation of VSMCs were observed [82]. ...
Vascular malformations may arise in any of the vascular beds present in the human body. These lesions vary in location, type, and clinical severity of the phenotype. In recent years, the genetic basis of several vascular malformations has been elucidated. This review will consider how the identification of the genetic factors contributing to different vascular malformations, with subsequent functional studies in animal models, has provided a better understanding of these factors that maintain vascular integrity in vascular beds, as well as their role in the pathogenesis of vascular malformations.
... This suggests that additional triggers are needed for AVM development. [11][12][13] Local homozygous loss of Eng or Acvrl1 gene expression, neoangiogenesis, inflammation, and wounding have been implicated triggering arteriovenous shunt formation, in accordance with secondary triggers acting as underlying mechanisms. Ectopic expression of vascular endothelial growth factor, the prime angiogenic growth factor, using adeno-associated viruses has been shown to induce cerebrovascular dysplasia in both HHT1 and HHT2 mouse models. ...
... 14,15 Inflammation induces endoglin protein null locally 16 that may increase the risk of vascular abnormalities in Eng +/− mice. 13 Postnatal homologous loss of the Eng or Acvrl1 gene in endothelial cells leads to the formation of arteriovenous shunts resembling those seen in HHT individuals only in sites where angiogenesis is active, supporting an hypothesis that at least 3 hits-the loss of both Eng and Acvrl1 alleles combined with environmental proangiogenic triggers-are necessary for AVM development. 10 How mutations in the Eng and Acvrl1 genes lead to AVM formation is still poorly understood, although recent findings indicate that aberrant angiogenesis may account for the development of such vessel abnormalities. ...
Objective:
To determine the role of Gja5 that encodes for the gap junction protein connexin40 in the generation of arteriovenous malformations in the hereditary hemorrhagic telangiectasia type 2 (HHT2) mouse model.
Approach and results:
We identified GJA5 as a target gene of the bone morphogenetic protein-9/ALK1 signaling pathway in human aortic endothelial cells and importantly found that connexin40 levels were particularly low in a small group of patients with HHT2. We next took advantage of the Acvrl1(+/-) mutant mice that develop lesions similar to those in patients with HHT2 and generated Acvrl1(+/-); Gja5(EGFP/+) mice. Gja5 haploinsufficiency led to vasodilation of the arteries and rarefaction of the capillary bed in Acvrl1(+/-) mice. At the molecular level, we found that reduced Gja5 in Acvrl1(+/-) mice stimulated the production of reactive oxygen species, an important mediator of vessel remodeling. To normalize the altered hemodynamic forces in Acvrl1(+/-); Gja5(EGFP/+) mice, capillaries formed transient arteriovenous shunts that could develop into large malformations when exposed to environmental insults.
Conclusions:
We identified GJA5 as a potential modifier gene for HHT2. Our findings demonstrate that Acvrl1 haploinsufficiency combined with the effects of modifier genes that regulate vessel caliber is responsible for the heterogeneity and severity of the disease. The mouse models of HHT have led to the proposal that 3 events-heterozygosity, loss of heterozygosity, and a proangiogenic-are necessary for arteriovenous malformation formation. Here, we present a novel 3-step model in which pathological vessel caliber and consequent altered blood flow are necessary events for arteriovenous malformation development.
... HHTmutations crosstalk even with the VEGF, PTEN, PI3K/AKT, and MAPK/ERK activity, which regulates the physiological mechanism of angiogenesis [53][54][55]. This evidence, also supported by studies on genetic mouse models, supports the hypothetical role of the aberrant pathways underlying the etiology and pathogenesis of hereditary bAVMs [56][57][58]. ...
Introduction:
microRNAs (miRNAs) are a class of non-coding RNAs playing a myriad of important roles in regulating gene expression. Of note, recent work demonstrated a critical role of miRNAs in the genesis and progression of brain arteriovenous malformations (bAVMs). Accordingly, here we examine miRNA signatures related to bAVMs and associated gene expression. In so doing we expound on the potential prognostic, diagnostic, and therapeutic significance of miRNAs in the clinical management of bAVMs.
Methods:
A PRISMA-based literature review was performed using PubMed/Medline database with the following search terms: "brain arteriovenous malformations", "cerebral arteriovenous malformations", "microRNA", and "miRNA". All preclinical and clinical studies written in English, regardless of date, were selected. For our bioinformatic analyses, miRWalk and miRTarBase machine learning algorithms were employed; the Kyoto Encyclopedia of Genes and Genomes (KEGG) database was quired for associated pathways/functions.
Results:
four studies were ultimately included in the final analyses. Sequencing data consistently revealed the decreased expression of miR-18a in bAVM-endothelial cells, resulting in increased levels of vascular endodermal growth factor (VEGF), Id-1, matrix metalloproteinase, and growth signals. Our analyses also suggest that the downregulation of miR-137 and miR-195* within vascular smooth muscle cells (VSMCs) may foster the activation of inflammation, aberrant angiogenesis, and phenotypic switching. In the peripheral blood, the overexpression of miR-7-5p, miR-629-5p, miR-199a-5p, miR-200b-3p, and let-7b-5p may contribute to endothelial proliferation and nidus development. The machine learning algorithms employed confirmed associations between miRNA-related target networks, vascular rearrangement, and bAVM progression.
Conclusion:
miRNAs expression appears to be critical in managing bAVMs' post-transcriptional signals. Targets of microRNAs regulate canonical vascular proliferation and reshaping. Although additional scientific evidence is needed, the identification of bAVM miRNA signatures may facilitate the development of novel prognostic/diagnostic tools and molecular therapies for bAVMs.
... Of note, the HHT phenotype of these animals is highly dependent on the mouse strain. Thus, Eng +/− and Alk1 +/− animals with 129/Ola background develop highly penetrant and frequent clinical manifestations similar to those of HHT, such as dilated vessels, nosebleeds, telangiectases and vascular lesions in the liver, the nail bed, ears, intestine and skin (24,51,52). By contrast, in Eng +/− and Alk1 +/− animals with a C57BL/6J background the penetrance of the HHT phenotype is incomplete and its frequency is negligible (24,51). ...
Hereditary Hemorrhagic Telangiectasia (HHT) is an autosomal-dominant genetic disorder involving defects in two predominant genes known as endoglin ( ENG ; HHT-1) and activin receptor-like kinase 1 ( ACVRL1/ALK1 ; HHT-2). It is characterized by mucocutaneous telangiectases that, due to their fragility, frequently break causing recurrent epistaxis and gastrointestinal bleeding. Because of the severity of hemorrhages, the study of the hemostasis involved in these vascular ruptures is critical to find therapies for this disease. Our results demonstrate that HHT patients with high bleeding, as determined by a high Epistaxis Severity Score (ESS), do not have prolonged clotting times or alterations in clotting factors. Considering that coagulation is only one of the processes involved in hemostasis, the main objective of this study was to investigate the overall mechanisms of hemostasis in HHT-1 ( Eng +/− ) and HHT-2 ( Alk1 +/− ) mouse models, which do not show HHT vascular phenotypes in the meaning of spontaneous bleeding. In Eng +/− mice, the results of in vivo and in vitro assays suggest deficient platelet-endothelium interactions that impair a robust and stable thrombus formation. Consequently, the thrombus could be torn off and dragged by the mechanical force exerted by the bloodstream, leading to the reappearance of hemorrhages. In Alk1 +/− mice, an overactivation of the fibrinolysis system was observed. These results support the idea that endoglin and Alk1 haploinsufficiency leads to a common phenotype of impaired hemostasis, but through different mechanisms. This contribution opens new therapeutic approaches to HHT patients' epistaxis.
... Previous studies have demonstrated that TGF-β1/ALK1/ENG signaling leads to an increase in EC proliferation (51) but not in SMCs (52). Moreover, increased EC proliferation with decreased SMCs is observed in malformed vessels of HHT and in mouse models (15). Consistent with these observations, TGF-β1 stimulation showed a minimal impact on cell proliferation (18% increase) with an increase in cell death, as assessed by increased cleaved caspase-3/7 activity in SMCs isolated from Nrp1 SM22KO compared with WT sex-matched littermates. ...
Hereditary hemorrhagic telangiectasia (HHT) patients have arteriovenous malformations (AVMs) with genetic mutations involving the activin-A receptor like type 1 (ACVRL1 or ALK1) and endoglin (ENG). Recent study showed that Neuropilin-1 (NRP-1) inhibits ALK1. We investigated the expression of NRP-1 in livers of patients with HHT and found that there was a significant reduction in NRP-1 in perivascular smooth muscle cells (SMCs). We used Nrp1SM22KO mice (Nrp1 was ablated in SMCs) and found hemorrhage, increased immune cell infiltration with decrease in SMCs and pericyte lining in lungs and liver in adult mice. Histologic examination revealed lung AVFs with enlarged liver vessels. Evaluation of the retina vessels at post-natal day 5 from Nrp1SM22KO mice demonstrated dilated capillaries with a reduction of pericytes. In inflow artery of surgical AVFs from the Nrp1SM22KO vs. WT mice, there was a significant decrease in Tgfb1, Eng and Alk1 expression, phosphorylated-SMAD (pSMAD)1/5/8, with an increase in apoptosis. TGF-β1 stimulated aortic SMCs from Nrp1SM22KO vs. WT mice have decreased pSMAD1/5/8 upon and increased apoptosis. Coimmunoprecipitation experiments revealed that NRP-1 interacts with ALK1 and ENG in SMCs. In sum, NRP-1 deletion in SMCs leads to reduced ALK1, ENG, pSMAD1/5/8 signaling, and cell death associated with AVM formation.
... We next examined various organs in endoglin heterozygous mice (Eng +/− ) that are the closest genetic model of HHT1 in terms of genotype. These mice exhibited an intermediate phenotype with a widespread abnormality of the vascular walls and development of agedependent vascular lesions similar to those seen in HHT patients [42][43][44]. As expected, endoglin levels were reduced by half in all analyzed organs ( Figure 3A). ...
Hereditary Hemorrhagic Telangiectasia type 1 (HHT1) is an autosomal dominant inherited disease characterized by arteriovenous malformations and hemorrhage. HHT1 is caused by mutations in ENDOGLIN, which encodes an ancillary receptor for Transforming Growth Factor-β/Bone Morphogenetic Protein-9 expressed in all vascular endothelial cells. Haploinsufficiency is widely accepted as the underlying mechanism for HHT1. However, it remains intriguing that only some, but not all, vascular beds are affected, as these causal gene mutations are present in vascula-ture throughout the body. Here, we have examined the endoglin expression levels in the blood vessels of multiple organs in mice and in humans. We found a positive correlation between low basal levels of endoglin and the general prevalence of clinical manifestations in selected organs. Endoglin was found to be particularly low in the skin, the earliest site of vascular lesions in HHT1, and even undetectable in the arteries and capillaries of heterozygous endoglin mice. Endoglin levels did not appear to be associated with organ-specific vascular functions. Instead, our data revealed a critical endoglin threshold compatible with the haploinsufficiency model, below which endothelial cells independent of their tissue of origin exhibited abnormal responses to Vascular Endothelial Growth Factor. Our results support the development of drugs promoting endoglin expression as potentially protective.
... These Bmp9-KO mice also showed a marked fat accumulation and liver steatosis which had not been described previously 32 . Interestingly, we identified a strain-dependent effect for Bmp9 deletion with a stronger phenotype in the 129/Ola background versus C57BL/6 and BALB/c background 99 as previously shown in Eng-KO mice 100,101 . The reason for this strain specificity is not yet understood but clearly supports the notion of genetic modifiers in BMP9 signaling. ...
Bone morphogenetic proteins (BMPs) are dimeric transforming growth factor ß (TGFß) family cytokines that were first described in bone and cartilage formation but have since been shown to be involved in many pleiotropic functions. In human, there are 15 BMP ligands, which initiate their cellular signaling by forming a complex with two copies of type I receptors and two copies of type II receptors, both of which are transmembrane receptors with an intracellular serine/threonine kinase domain. Within this receptor family, ALK1 (activin receptor‐like kinase 1), which is a type I receptor mainly expressed on endothelial cells, and BMPRII (BMP Receptor type II), a type II receptor also highly expressed on endothelial cells, have been directly linked to two rare vascular diseases: hereditary hemorrhagic telangiectasia (HHT), and pulmonary arterial hypertension (PAH), respectively. BMP9 (gene name GDF2) and BMP10, two close members of the BMP family, are the only known ligands for the ALK1 receptor. This specificity gives them a unique role in physiological and pathological angiogenesis and tissue homeostasis. The aim of this current review is to present an overview of what is known about BMP9 and BMP10 on vascular regulation with a particular emphasis on recent results and the many questions that remain unanswered regarding the roles and specificities between BMP9 and BMP10.
... Strong evidence from mouse models of HHT1 and HHT2 also supports this two-hit mechanism where complete endothelial loss of ENG or ALK1 protein is a pre-requisite for robust vascular lesion formation in response to angiogenic stimuli [7]. In contrast, mice that are heterozygous for mutations (Eng+/− or Alk1+/−) have low penetrance of disease [8,9]. ...
IntroductionEndoglin (ENG) forms a receptor complex with ALK1 in endothelial cells (ECs) to promote BMP9/10 signalling. Loss of function mutations in either ENG or ALK1 genes lead to the inherited vascular disorder hereditary haemorrhagic telangiectasia (HHT), characterised by arteriovenous malformations (AVMs). However, the vessel-specific role of ENG and ALK1 proteins in protecting against AVMs is unclear. For example, AVMs have been described to initiate in arterioles, whereas ENG is predominantly expressed in venous ECs. To investigate whether ENG has any arterial involvement in protecting against AVM formation, we specifically depleted the Eng gene in venous and capillary endothelium whilst maintaining arterial expression, and investigated how this affected the incidence and location of AVMs in comparison with pan-endothelial Eng knockdown.Methods
Using the mouse neonatal retinal model of angiogenesis, we first established the earliest time point at which Apj-Cre-ERT2 activity was present in venous and capillary ECs but absent from arterial ECs. We then compared the incidence of AVMs following pan-endothelial or venous/capillary-specific ENG knockout.ResultsActivation of Apj-Cre-ERT2 with tamoxifen from postnatal day (P) 5 ensured preservation of arterial ENG protein expression. Specific loss of ENG expression in ECs of veins and capillaries led to retinal AVMs at a similar frequency to pan-endothelial loss of ENG. AVMs occurred in the proximal as well as the distal part of the retina consistent with a defect in vascular remodelling during maturation of the vasculature.Conclusion
Expression of ENG is not required in arterial ECs to protect against AVM formation.
... Strong evidence from mouse models of HHT1 and HHT2 also supports this two hit mechanism where complete endothelial loss of ENG or ALK1 protein is a pre-requisite for robust vascular lesion formation in response to angiogenic stimuli [7]. In contrast, mice that are heterozygous for mutations (Eng+/-or Alk1+/-) have low penetrance of disease [8,9]. ...
Endoglin (ENG) forms a receptor complex with ALK1 in endothelial cells (ECs) to promote BMP9/10 signalling. Loss of function mutations in either ENG or ALK1 genes lead to the inherited vascular disorder Hereditary Haemorrhagic Telangiectasia (HHT), characterised by arteriovenous malformations (AVMs). However, the vessel-specific role of ENG and ALK1 proteins in protecting against AVMs is unclear. For example, AVMs have been described to initiate in arterioles, whereas ENG is predominantly expressed in venous ECs. To investigate whether ENG has any arterial involvement in protecting against AVM formation we specifically depleted the Eng gene in venous and capillary endothelium whilst maintaining arterial expression, and investigated how this affected the incidence and location of AVMs in comparison with pan-endothelial Eng knockdown.
Methods: Using the mouse neonatal retinal model of angiogenesis, we first established the earliest time point at which Apj-Cre-ERT2 activity was present in venous and capillary ECs but absent from arterial ECs. We then compared the incidence of AVMs following pan-endothelial or venous/capillary-specific ENG knockout.
Results: Activation of Apj-Cre-ERT2 with tamoxifen from postnatal day (P) 5 ensured preservation of arterial ENG protein expression. Specific loss of ENG expression in ECs of veins and capillaries led to retinal AVMs at a similar frequency to pan-endothelial loss of ENG. AVMs occurred in the proximal as well as the distal part of the retina consistent with a defect in vascular remodelling during maturation of the vasculature.
Conclusion: Expression of ENG is not required in arterial ECs to protect against AVM formation.
... Genetic mouse models of bAVM have been mostly developed by manipulating Eng or Alk1 genes. Eng +/− or Alk1 +/− heterozygous knockout (KO) mice are viable and show HHT phenotypes during adulthood, while Eng −/− or Alk1 −/− homozygous KO mice are embryonic lethal [38,[54][55][56][57][58][59]. bAVMs, including AV shunts and niduses of dilated vessels, occurred in only 30% of Eng +/− mice aged 25 to 40 weeks with incomplete penetrance [56]. ...
Arteriovenous malformations (AVMs) are abnormal connections of vessels that shunt blood directly from arteries into veins. Rupture of brain AVMs (bAVMs) can cause life-threatening intracranial bleeding. Even though the majority of bAVM cases are sporadic without a family history, some cases are familial. Most of the familial cases of bAVMs are associated with a genetic disorder called hereditary hemorrhagic telangiectasia (HHT). The mechanism of bAVM formation is not fully understood. The most important advances in bAVM basic science research is the identification of somatic mutations of genes in RAS-MAPK pathways. However, the mechanisms by which mutations of these genes lead to AVM formation are largely unknown. In this review, we summarized the latest advance in bAVM studies and discussed some pathways that play important roles in bAVM pathogenesis. We also discussed the therapeutic implications of these pathways.
... Genetic mouse models of bAVM have been mostly developed by manipulating Eng or Alk1 genes. Eng +/− or Alk1 +/− heterozygous knockout (KO) mice are viable and show HHT phenotypes during adulthood, while Eng -/-or Alk1 -/-homozygous KO mice are embryonic lethal [38,[54][55][56][57][58][59]. bAVMs, including AV shunts and niduses of dilated vessels, occurred in only 30% of Eng +/− mice aged 25 to 40 weeks with incomplete penetrance [56]. ...
Arteriovenous malformations (AVMs) are abnormal connections of vessels that shunt blood directly from arteries into veins. Rupture of brain AVMs (bAVMs) can cause life-threatening intracranial bleeding. Even though the majority of bAVM cases are sporadic without a family history, some cases are familial. Most of the familial cases of bAVMs are associated with a genetic disorder called hereditary hemorrhagic telangiectasia (HHT). The mechanism of bAVM formation is not fully understood. The most important advances in bAVM basic science research is the identification of somatic mutations of genes in RAS-MAPK pathways. However, the mechanisms by which mutations of these genes lead to AVM formation are largely unknown. In this review, we summarized the latest advance in bAVM studies and discussed some pathways that play important roles in bAVM pathogenesis. We also discussed the therapeutic implications of these pathways.
... Heterozygous mice for Acvrl1 or Eng reproduce some HHT-like lesions but with a low frequency [22]. Interestingly, these HHT-like lesions have been shown to be more frequent in the 129/Ola than in the C57BL/6 genetic background, suggesting again that genetic modifiers might play a role in susceptibility to the HHT disease [23,24]. ...
The aim of the present work was to address the role of BMP9 in different genetic backgrounds (C57BL/6, BALB/c, and 129/Ola) of mice deleted for Bmp9. We found that Bmp9 deletion led to premature mortality only in the 129/Ola strain. We have previously shown that Bmp9 deletion led to liver sinusoidal endothelial cells (LSEC) capillarization and liver fibrosis in the 129/Ola background. Here, we showed that this is not the case in the C57BL/6 background. Analysis of LSEC from Wild-type (WT) versus Bmp9-KO mice in the C57BL/6 background showed no difference in LSEC fenestration and in the expression of differentiation markers. Comparison of the mRNA expression of LSEC differentiation markers between WT C57BL/6 and 129/Ola mice showed a significant decrease in Stabilin2, Plvap, and CD209b, suggesting a more capillary-like phenotype in WT C57BL/6 LSECs. C57BL/6 mice also had lower BMP9 circulating concentrations and hepatic Vegfr2 mRNA levels, compared to the 129/Ola mice. Taken together, our observations support a role for BMP9 in liver endothelial cell fenestration and prevention of fibrosis that is dependent on genetic background. It also suggests that 129/Ola mice are a more suitable model than C57BL/6 for the study of liver fibrosis subsequent to LSEC capillarization.
... Tu et al. proposed that BMP9 confers a vasoconstrictive effect by inducing factors such as endothelin-1 and suppressing vasodilation factors such as adrenomedullin and apelin, whereas our report has shown extensively that exogenous BMP9 exerts anti-apoptotic and anti-proliferative effects on vascular endothelial cells, thereby reversing PAH by correcting endothelial dysfunction [7]. Nevertheless, the findings by Tu et al. are of interest and, if confirmed, provide a starting point to examine whether the chronic loss of BMP9 induces vascular changes typical of HHTwith reduced vascular tone and reduced muscularisation of arterioles, as seen in mice that are heterozygous for a targeted deletion in endoglin gene [37]. Such changes would protect rodents from pulmonary vasoconstrictive stimuli, but chronic inhibition of BMP9 might be a hazardous approach in PAH patients, given the strong human genetic evidence that loss-of-function in the BMP9 signalling axis promotes the development of PAH [5]. ...
Bone morphogenetic protein 9 (BMP9), a member of the transforming growth factor β (TGFβ) superfamily, is a circulating vascular quiescence and endothelial protective factor, accounting for the majority of BMP activities in plasma. BMP9 and BMP10 bind preferentially to the high-affinity type I receptor activin receptor-like kinase 1 on vascular endothelial cells. Recently, many reports have highlighted the important roles of BMP9 in cardiovascular disease, particularly pulmonary arterial hypertension. In vivo, BMP9 activity and specificity are determined by tightly regulated protein-protein recognition with cognate receptors and a co-receptor, and may also be influenced by other proteins present on the endothelial cell surface (such as low-affinity receptors) and in circulation (such as TGFβ family ligands competing for the same receptors). In this review, we summarise recent findings on the role and therapeutic potential of BMP9 in cardiovascular disease and review the current understanding of how the extracellular protein-protein interaction milieu could play a role in regulating endothelial BMP9 signalling specificity and activity.
... This prompted us to analyze the effects of BMP9 gene deletion on hepatic functions. For this, as HHT symptoms were demonstrated to be more frequent in the 129/Ola background than in the C57BL/6 background, (20,21) we generated Bmp9-KO 129/Ola mice and observed that they presented strong hepatic perisinusoidal fibrosis. We report here the phenotypic and mechanistic characterizations of these liver alterations, which lend us to propose a key role for BMP9 in liver homeostasis. ...
Bone morphogenetic protein 9 (BMP9) is a circulating factor produced by hepatic stellate cells that plays a critical role in vascular quiescence via its endothelial receptor ALK1. Mutations in the gene encoding ALK1 cause HHT2 (Hereditary Hemoragic Telangiectasia type 2), a rare genetic disease presenting hepatic vessel malformations. Variations of both the circulating levels and the hepatic mRNA levels of BMP9 have been recently associated with various forms of hepatic fibrosis. However, the molecular mechanism that links BMP9 with liver diseases is still unknown. Here, we report that Bmp9 gene deletion in 129/Ola mice triggers hepatic perisinusoidal fibrosis that was detectable from 15 weeks of age. An inflammatory response appeared within the same time frame as fibrosis, whereas sinusoidal vessel dilation developed later on. Proteomic and mRNA analyses of primary liver sinusoidal endothelial cells (LSECs) both revealed that the expression of the LSEC‐specifying transcription factor GATA4 was strongly reduced in Bmp9‐KO mice as compared to wild type mice. LSECs from Bmp9‐KO mice also lost the expression of several terminal differentiation markers (Lyve1, Stab1, Stab2, Ehd3, Cd209b, eNos, Maf, Plvap). They gained CD34 expression and deposited a basal lamina, indicating that they were capillarized. Another main characteristic of differentiated LSECs is the presence of permeable fenestrae. LSECs from Bmp9‐KO mice had a significantly reduced number of fenestrae. This was already observable in 2‐week‐old pups. Moreover, we could show that addition of BMP9 to primary cultures of LSECs prevented the loss of their fenestrae and maintained the expression levels of Gata4 and Plvap. Taken together, our observations show that BMP9 is a key paracrine regulator of liver homeostasis, controlling LSEC fenestration and protecting from perivascular hepatic fibrosis. This article is protected by copyright. All rights reserved.
... So, one hypothesis that could explain why the loss of BMP9 protects from PH development is that BMP9 could act as a potent modulator of vasoconstriction in the pulmonary circulation. This hypothesis is consistent with the dilation phenotype observed in Alk1 +/-or Eng +/-mice 29,30 or Alk1 mutants, vbg, in zebrafish 31 . This hypothesis is further supported by the fact that we found that Bmp9 -/-pups presented a partial reopening of the ductus arteriosus just after birth that could due to a defect in vasoconstriction 32 . ...
Rationale: Although many familial cases of pulmonary arterial hypertension (PAH) exhibit an autosomal dominant mode of inheritance with the majority having mutations in essential constituents of the bone morphogenetic protein (BMP) signaling, the specific contribution of the long-term loss of signal transduction triggered by the type 2 BMP receptor (BMPR2) remains poorly characterized.
Objective: To investigate the role of BMP9, the main ligand of ALK1/BMPR2 heterocomplexes, in pulmonary hypertension (PH).
Method and Results: The absence of BMP9 in Bmp9-/- mice and its inhibition in C57BL/6 mice using neutralizing anti-BMP9 antibodies substantially prevent against chronic hypoxia induced PH judged by
right ventricular systolic pressure (RVSP) measurement, right ventricular hypertrophy, and pulmonary distal arterial muscularization. In agreement with these observations, we found that the BMP9/BMP10 ligand trap ALK1ECD administered in monocrotaline (MCT) or Sugen/hypoxia (SuHx) rats substantially attenuate proliferation of pulmonary vascular cells, inflammatory cell infiltration and regresses established PH in rats. Our data obtained in human pulmonary endothelial cells derived from controls and PAH patients
indicate that BMP9 can affect the balance between endothelin-1, apelin and adrenomedullin. We reproduced these in vitro observations in mice chronically exposed to hypoxia, with Bmp9-/- mice
exhibiting lower mRNA levels of the vasoconstrictor peptide endothelin (ET)-1 and higher levels of the two potent vasodilator factors apelin and adrenomedullin (ADM) compared with Bmp9 +/+ littermates.
Conclusion: Taken together, our data indicate that the loss of BMP9, by deletion or inhibition, has beneficial effects against PH onset and progression.
... Many studies like 4,3 have been demonstrated that COPD patients of PAH were play a vital role in PAH which associated with decrement of BMPR2 that inversely correlated with endoglin protein level.The results in figure (9) shows a significant increase (P 0.05) between different grades severe, moderate and mild of secondary and primary patients group with the pulmonary arterial hypertension. The endoglin in mice increased pulmonary macrophages infiltration and enhanced inflammation and severity of disease have been described as a vascular bleeding 30 and have important roles in the pathogenesis of PAH 31 . The highly expression of TGFpathway in lung tissue and pulmonary endothelial cells (PECs) and pulmonary artery smooth muscle cells (PASMC) from patients of IPAH may be related severity of disease 32 . ...
Abstract
Background and Objective: The study was done at the Cardiac Centre of Al-Sader Medical City Hospital in Al-Najaf Al-Ashraf City during
the period from December 2016 to May 2017 to investigate the level of biomarker of endoglin for patients with pulmonary arterial
hypertension as predictive indicators of the disease for detection, diagnosis and early treatment of patients.
Materials and Methods:The study was applied on 88 people aged 30-69 years; 67 patients group with pulmonary arterial hypertension (PAH)
and 21 healthy group. The group of patients was divided into subgroups based on gender, age, body mass index (BMI), waist circumference,
smokers and non-smokers, type of primary and secondary arterial hypertension, types of secondary arterial hypertension, and disease grade.
The healthy group was divided according to gender and age.
Results :The study protruded a significant increase (p <0.05) in the level of Endoglin in the group of patients with pulmonary arterial
hypertension compared to healthy, the female group compared to males, Smokers compared to non-smokers, and the group of two types of
PAH disease found a significant increase in secondary compared to the primary PAH , the grade of disease group found increase in the severe
level compared to moderate and mild and the age group at the age of 30 to 69 , but there is no significant increase between ages 30 to 49.
While the level of Endoglin in the healthy group divided by gender and age have no significant differences that are almost constant. In
comparing secondary PAH types, there was a significant increase (p<0.05) in the level of Endoglin for both Chronic obstructive pulmonary
disease (COPD) and Left Heart Disease systolic dysfunction or diastolic dysfunction (compared with Valvular, Congenital and Pulmonary
embolism. While there was no significant difference in the level of Endoglin between the Valvular, Congenital and Pulmonary embolism.The
study explained that no significant difference in the level of Endoglin for the group of patients with pulmonary arterial hypertension divided by
BMI group (normal weight, over weight, obese weight) and waist circumference group (70-80 cm , 81-90 cm, 91-100 cm, 101-110 cm, 111-
120 cm).
... Many studies like 4,3 have been demonstrated that COPD patients of PAH were play a vital role in PAH which associated with decrement of BMPR2 that inversely correlated with endoglin protein level.The results in figure (9) shows a significant increase (P 0.05) between different grades severe, moderate and mild of secondary and primary patients group with the pulmonary arterial hypertension. The endoglin in mice increased pulmonary macrophages infiltration and enhanced inflammation and severity of disease have been described as a vascular bleeding 30 and have important roles in the pathogenesis of PAH 31 . The highly expression of TGFpathway in lung tissue and pulmonary endothelial cells (PECs) and pulmonary artery smooth muscle cells (PASMC) from patients of IPAH may be related severity of disease 32 . ...
Background and Objective: The study was done at the Cardiac Centre of Al-Sader Medical City Hospital in Al-Najaf Al-Ashraf City during the period from December 2016 to May 2017 to investigate the level of biomarker of endoglin for patients with pulmonary arterial hypertension as predictive indicators of the disease for detection, diagnosis and early treatment of patients. Materials and Methods:The study was applied on 88 people aged 30-69 years; 67 patients group with pulmonary arterial hypertension (PAH) and 21 healthy group. The group of patients was divided into subgroups based on gender, age, body mass index (BMI), waist circumference, smokers and non-smokers, type of primary and secondary arterial hypertension, types of secondary arterial hypertension, and disease grade. The healthy group was divided according to gender and age. Results:The study protruded a significant increase (p <0.05) in the level of Endoglin in the group of patients with pulmonary arterial hypertension compared to healthy, the female group compared to males, Smokers compared to non-smokers, and the group of two types of PAH disease found a significant increase in secondary compared to the primary PAH, the grade of disease group found increase in the severe level compared to moderate and mild and the age group at the age of 30 to 69, but there is no significant increase between ages 30 to 49. While the level of Endoglin in the healthy group divided by gender and age have no significant differences that are almost constant. In comparing secondary PAH types, there was a significant increase (p<0.05) in the level of Endoglin for both Chronic obstructive pulmonary disease (COPD) and Left Heart Disease systolic dysfunction or diastolic dysfunction (compared with Valvular, Congenital and Pulmonary embolism. While there was no significant difference in the level of Endoglin between the Valvular, Congenital and Pulmonary embolism.The study explained that no significant difference in the level of Endoglin for the group of patients with pulmonary arterial hypertension divided by BMI group (normal weight, over weight, obese weight) and waist circumference group (70-80 cm, 81-90 cm, 91-100 cm, 101-110 cm, 111-120 cm). Conclusion: The biomarker Endoglin consider warning sign as a prognostic marker for diagnosis PAH patients.
... So, one hypothesis that could explain why the loss of BMP9 protects from PH development is that BMP9 could act as a potent modulator of vasoconstriction in the pulmonary circulation. This hypothesis is consistent with the dilation phenotype observed in Alk1 +/-or Eng +/-mice 29,30 or Alk1 mutants, vbg, in zebrafish 31 . This hypothesis is further supported by the fact that we found that Bmp9 -/-pups presented a partial reopening of the ductus arteriosus just after birth that could due to a defect in vasoconstriction 32 . ...
... These mice die during gestation likely from severe impairments in vascular development ( Li et al. 1999, Urness et al. 2000. Alterations in a single copy of endoglin or ALK-1 in mice result in spontaneous vascular dysplasia, telangiectasia and random hemorrhages ( Bourdeau et al. 1999, Satomi et al. 2003, Srinivasan et al. 2003, Torsney et al. 2003, Park et al. 2008). Small vessel malformations are observed in these mice together with disappearance of vascular smooth muscle cells. ...
... The importance of Eng as a crucial contributor for correct vascular development has been shown in knockout mouse studies. Eng -/mouse embryos die during embryogenesis due to severe vascular and cardiac defects [29,30]. Li et al. found that Eng -/mice present an absence of vascular organisation, atretic major vessels, a failure of endothelial remodelling and poor vascular SMC Fig. 1 qRT-PCR revealed significantly decreased relative mRNA expression levels of Eng, (**p \ 0.005) Tgfbr1 (*p \ 0.05) and Tgfbr2 (***p \ 0.001) lungs of the CDH group compared to control lung tissue Fig. 2 Western blot results demonstrated a decreased pulmonary expression of Eng, Tgfbr1 and Tgfbr2 in the CDH group compared to control animals. ...
Aim of the study:
Pulmonary hypertension (PH) remains a therapeutical challenge in neonates born with congenital diaphragmatic hernia (CDH). Endoglin (Eng), an auxiliary receptor component of the transforming growth factor β (TGFβ) signalling pathway, is expressed mainly by endothelial cells and has been found to be involved in angiogenesis and vascular remodelling. Genetic studies have linked TGFβ and Eng mutations to human arterial PH and other cardiovascular syndromes. Eng interacts with the TGFβ receptors 1 and 2 (Tgfβr1, Tgfβr2). We designed this study to investigate the hypothesis that Eng is altered in the pulmonary vasculature of rats with nitrofen-induced CDH subjected to its interdependency with Tgfβr1 and Tgfβr2.
Methods:
After ethical approval (Rec 913b), time-pregnant Sprague-Dawley rats received either nitrofen or olive oil on gestational day (D9). The foetuses (n = 22) were sacrificed and divided into CDH and control group on D21. Gene and protein expressions of Eng, Tgfβr1 and Tgfβr2 were assessed via qRT-PCR and western blotting. Immunofluorescence staining for Eng was combined with CD34 to evaluate Eng expression in the pulmonary vasculature.
Main results:
Relative mRNA levels of Eng, Tgfβr1 and Tgfβr2 were significantly downregulated in CDH lungs compared to controls (Eng CDH 0.341 ± 0.022, Eng Ctrl 0.471 ± 0.031, p = 0.0015; Tgfβr1 CDH 0.161 ± 0.008, Tgfβr1 Ctrl 0.194 ± 0.01, p = 0.0114; Tgfβr2 CDH 0.896 ± 0.099, Tgfβr2 Ctrl 1.379 ± 0.081, p = 0.0006) Western blotting confirmed the reduced pulmonary protein expression of these three proteins in the CDH lungs. A markedly diminished endothelial expression of Eng in the pulmonary vasculature of nitrofen-exposed foetuses compared to controls was seen in laser scanning confocal-microscopy.
Conclusion:
This study demonstrates for the first time a reduced expression of Endoglin in the pulmonary vasculature of nitrofen-induced CDH. Abnormal Eng/Tgfβr1/Tgfβr2 signalling may contribute to impaired vascular remodelling and development of PH in this CDH animal model.
... Another animal that had an ear telangiectasis showed an AVM nidus that resembled those in human and canine brain [70]. Despite the fact that Eng +/í and Acvrl1 +/í mice are the best available HHT models at present, past studies do not show the HHT features at high frequency [69,72,73]. Although the two studies could be criticised as not being true AVM models and for their small sample size, they may contribute to research on the abnormal phenotype of AVMs in humans [66]. ...
Arteriovenous malformations (AVMs) are congenital lesions that cause brain haemorrhage in children and young adults. Current treatment modalities include surgery, radiosurgery and embolization. These treatments are generally effective only for small AVMs. Over one third of AVMs cannot be treated safely and effectively with existing options. Several animal models have been developed with the aims of understanding AVM pathophysiology and improving treatment. No animal model perfectly mimics a human AVM. Each model has limitations and advantages. Models contribute to the understanding of AVMs and hopefully to the development of improved therapies. This paper reviews animal models of AVMs and their advantages and disadvantages.
... Surgical AVM samples are a diverse mix of vascular cells (endothelial cells, pericytes, and smooth muscle cells), circulating cells (lymphocytes), glial cells, and fibroblasts, with the exact pathological cell population being unknown ( Table 2). [39][40][41][42] Therefore, if the potential causative mutation occurs in only a single cell type (ie, endothelium), the purity of the sample analyzed is low, and the chance of finding that mutation in any single reading of the DNA sequence is proportionally low. Next-generation sequencing overcomes this limitation by reading DNA sequences many times over, with "deep sequencing" of several hundred-fold coverage being used to effectively identify low-frequency events in heterogeneous samples. ...
... Smad7 Smad7 transmembrane receptors, endoglin or activin-like kinase-1, which ultimately impair TGFb signaling in endothelial cells, compromising microvasculature and inducing hemorrhage (35). Such off-target effects were reported in clinical trials of an ActRIIB ligand trap (36) and are evident in mouse models of HHT. ...
Patients with advanced cancer often succumb to complications arising from striated muscle wasting associated with cachexia. Excessive activation of the type IIB activin receptor (ActRIIB) is considered an important mechanism underlying this wasting, where circulating procachectic factors bind ActRIIB and ultimately lead to the phosphorylation of SMAD2/3. Therapeutics that antagonize the binding of ActRIIB ligands are in clinical development, but concerns exist about achieving efficacy without off-target effects. To protect striated muscle from harmful ActRIIB signaling, and to reduce the risk of off-target effects, we developed an intervention using recombinant adeno-associated viral vectors (rAAV vectors) that increase expression of Smad7 in skeletal and cardiac muscles. SMAD7 acts as an intracellular negative regulator that prevents SMAD2/3 activation and promotes degradation of ActRIIB complexes. In mouse models of cachexia, rAAV:Smad7 prevented wasting of skeletal muscles and the heart independent of tumor burden and serum levels of procachectic ligands. Mechanistically, rAAV:Smad7 administration abolished SMAD2/3 signaling downstream of ActRIIB and inhibited expression of the atrophy-related ubiquitin ligases MuRF1 and MAFbx. These findings identify muscle-directed Smad7 gene delivery as a potential approach for preventing muscle wasting under conditions where excessive ActRIIB signaling occurs, such as cancer cachexia.
... Selectively expressed by endothelial cells undergoing vasculogenesis, angiogenesis, and inflammation Promotes endothelial cell proliferation, migration, and tube formation during early development and in tumors [73][74][75][76][77][78] Fibroblast growth factor receptor (FGFR) Bind to fibroblast growth factor family members, having diverse roles in organogenesis, proliferation, pro-survival cell signaling and cellular migration FGF/FGFR system contributes to both adult angiogenesis and tumor angiogenesis FGFR signaling is implicated in escape of tumor vasculature from VEGF inhibitor treatment [61,[79][80][81][82] Integrin αvβ3; CD51/CD61 An extracellular matrix adhesion receptor that coordinates endothelial cell responses during angiogenesis Expressed in neo-angiogenic vessels; therefore, it is upregulated in the vasculature of solid tumors, while expression is low in quiescent vasculature [83][84][85][86][87] Tumor endothelial marker 1 (TEM1); endosialin or CD248 One of the most abundantly expressed proteins on tumor vasculature and tumor stroma of human tumors Expression is variable among tumor types; however, this protein can reportedly be expressed on endothelial cells, pericytes, and/or fibroblasts of human and animal tumors [67,[88][89][90][91][92][93] See Tables 3, 4 and 5. Submit your next manuscript to BioMed Central and we will help you at every step: These Terms are supplementary and will apply in addition to any applicable website terms and conditions, a relevant site licence or a personal subscription. These Terms will prevail over any conflict or ambiguity with regards to the relevant terms, a site licence or a personal subscription (to the extent of the conflict or ambiguity only). ...
The mechanisms underlying discrimination between “self” and “non-self”, a central immunological principle, require careful consideration in immune oncology therapeutics where eliciting anti-cancer immunity must be weighed against the risk of autoimmunity due to the self origin of tumors. Whole cell vaccines are one promising immunotherapeutic avenue whereby a myriad of tumor antigens are introduced in an immunogenic context with the aim of eliciting tumor rejection. Despite the possibility collateral damage to healthy tissues, cancer immunotherapy can be designed such that off target autoimmunity remains limited in scope and severity or completely non-existent. Here we provide an immunological basis for reconciling the safety of cancer vaccines, focusing on tumor endothelial cell vaccines, by discussing the following topics: (a) Antigenic differences between neoplastic and healthy tissues that can be leveraged in cancer vaccine design; (b) The layers of tolerance that control T cell responses directed against antigens expressed in healthy tissues and tumors; and, (c) The hierarchy of antigenic epitope selection and display in response to whole cell vaccines, and how antigen processing and presentation can afford a degree of selectivity against tumors. We conclude with an example of early clinical data utilizing ValloVax™, an immunogenic placental endothelial cell vaccine that is being advanced to target the tumor endothelium of diverse cancers, and we report on the safety and efficacy of ValloVax™ for inducing immunity against tumor endothelial antigens.
... Selectively expressed by endothelial cells undergoing vasculogenesis, angiogenesis, and inflammation Promotes endothelial cell proliferation, migration, and tube formation during early development and in tumors [73][74][75][76][77][78] Fibroblast growth factor receptor (FGFR) Bind to fibroblast growth factor family members, having diverse roles in organogenesis, proliferation, pro-survival cell signaling and cellular migration FGF/FGFR system contributes to both adult angiogenesis and tumor angiogenesis FGFR signaling is implicated in escape of tumor vasculature from VEGF inhibitor treatment [61,[79][80][81][82] Integrin αvβ3; CD51/CD61 An extracellular matrix adhesion receptor that coordinates endothelial cell responses during angiogenesis Expressed in neo-angiogenic vessels; therefore, it is upregulated in the vasculature of solid tumors, while expression is low in quiescent vasculature [83][84][85][86][87] Preliminary clinical data demonstrate that vaccination of cancer patients with ValloVax ™ is not only well-tolerated but also elicits humoral immunity against an array of tumor endothelial cell-associated antigens with known pathological roles in tumor angiogenesis. ...
... So, one hypothesis that could explain why the loss of BMP9 protects from PH development is that BMP9 could act as a potent modulator of vasoconstriction in the pulmonary circulation. This hypothesis is consistent with the dilation phenotype observed in Alk1 +/-or Eng +/-mice 29,30 or Alk1 mutants, vbg, in zebrafish 31 . This hypothesis is further supported by the fact that we found that Bmp9 -/-pups presented a partial reopening of the ductus arteriosus just after birth that could due to a defect in vasoconstriction 32 . ...
... The major phenotype observed in these mutant mice is spontaneous PAH, associated with rarefaction of the peripheral lung vasculature and secondary right ventricular hypertrophy (RVH) [21,22]. Overall, these findings have led to the concept that mutations in ENG and ALK1 genes are associated with dysregulated angiogenesis, predisposing to HHT and/or PAH [12,23]. ...
Hereditary hemorrhagic telangiectasia (HHT) is a vascular dysplasia associated with dysregulated angi-ogenesis and arteriovascular malformations. The disease is caused by mutations in endoglin (ENG; HHT1) or activin receptor-like kinase 1 (ALK1; HHT2) genes, coding for transforming growth factor b (TGF-b) superfamily receptors. Vascular endothelial growth factor (VEGF) has been implicated in HHT and beneficial effects of anti-VEGF treatment were recently reported in HHT patients. To investigate the systemic angiogenic phenotype of Endoglin and Alk1 mutant mice and their response to anti-VEGF therapy, we assessed microvessel density (MVD) in multiple organs after treatment with an antibody to mouse VEGF or vehicle. Lungs were the only organ showing an angiogenic defect, with reduced peripheral MVD and secondary right ventricular hypertrophy (RVH), yet distinctly associated with a fourfold increase in thrombo-spondin-1 (TSP-1) in Eng ?/-versus a rise in angiopoietin-2 (Ang-2) in Alk1 ?/-mice. Anti-VEGF treatment did reduce lung VEGF levels but interestingly, led to an increase in peripheral pulmonary MVD and attenuation of RVH; it also normalized TSP-1 and Ang-2 expression. Hepatic MVD, unaffected in mutant mice, was reduced by anti-VEGF therapy in heterozygous and wild type mice, indicating a liver-specific effect of treatment. Contrast-enhanced micro-ultrasound demonstrated a reduction in hepatic microvascular perfusion after anti-VEGF treatment only in Eng ?/-mice. Our findings indicate that the mechanisms responsible for the angiogenic imbalance and the response to anti-VEGF therapy differ between Eng and Alk1 heterozygous mice and raise the need for systemic monitoring of anti-angiogenic therapy effects in HHT patients.
... Eng or Alk1 knockouts exhibit embryonic vascular defects, including dilated and fused artery-vein pairs and die in utero. 42,43 Eng +/− or Alk1 +/− heterozygous mice are viable and develop characteristics of HHT during adulthood; [10][11][12][13] however, features of BAVM, including AV shunts, niduses of dilated vessels, and rounded, misaligned EC nuclei, occur in 30% of Eng +/− mice aged 25-40 weeks, similar to BAVM incidence in HHT1 patients. 12 Thus, loss of one allele of Eng or Alk1 is sufficient to induce BAVM in adult mice, but with incomplete penetrance. ...
Arteriovenous (AV) malformation (AVM) is a vascular anomaly capable of both hemorrhagic and ischemic insults, leading to seizures, headaches, stroke, and even death.1 BAVM prevalence is estimated at 0.05%,2 often occurring in young people between 20 and 40 years of age.3 BAVMs account for 50% of hemorrhagic stroke in children4 and 1% to 2% of all strokes in the population.5 Brain AVMs (BAVMs) can cause life-threatening intracerebral hemorrhage (Figure 1).6 Fifty percent of patients are first diagnosed on intracerebral hemorrhage,1 with 1% and 5% annual hemorrhage rate for previously unruptured and ruptured AVMs, respectively.7,8 After BAVM rupture, reported mortality rates range from to 15% to 29%,7 and long-term morbidity rates range from 16% to 56%.1,9 Thus, BAVM is defined by vascular features and accompanying neurological deficits.1
Figure 1.
Features of human brain arteriovenous malformation (AVM). A , An AVM is visualized on the lateral temporal surface of a human brain. 5, 6 are landmarks placed by surgeon; 40 shows Broca's area; 48 shows Wernicke's area. B , Left internal carotid artery (ICA) angiography (lateral view) reveals a left lateral temporal AVM with a large feeding artery and draining vein. C , Cartoon of this subtype (lateral view), indicating feeding arteries and draining veins. ATA indicates anterior temporal artery. Reprinted from Lawton6 with permission of the publisher. Copyright © 2014, Thieme Medical Publishers.
AVM is characterized by high-flow AV connections that shunt blood directly from arteries to veins, displacing intervening capillaries with a nidus of enlarged and tortuous vessels. BAVM clinical characteristics include (1) AV shunting, the presence of direct connections between arteries and veins, displacing intervening capillaries; (2) abnormally high blood flow through the feeding artery, AV …
Hereditary hemorrhagic telangiectasia (HHT) is a genetic disorder characterized by vessel dilatation, such as telangiectasia in skin and mucosa and arteriovenous malformations (AVM) in internal organs such as the gastrointestinal tract, lungs, and brain. AVMs are fragile and tortuous vascular anomalies that directly connect arteries and veins, bypassing healthy capillaries. Mutations in transforming growth factor β (TGFβ) signaling pathway components, such as ENG (ENDOGLIN), ACVRL1 (ALK1), and SMAD4 (SMAD4) genes, account for most of HHT cases. 10–20% of HHT patients develop brain AVMs (bAVMs), which can lead to vessel wall rupture and intracranial hemorrhages. Though the main mutations are known, mechanisms leading to AVM formation are unclear, partially due to lack of animal models. Recent mouse models allowed significant advances in our understanding of AVMs. Endothelial-specific deletion of either Acvrl1, Eng or Smad4 is sufficient to induce AVMs, identifying endothelial cells (ECs) as primary targets of BMP signaling to promote vascular integrity. Loss of ALK1/ENG/SMAD4 signaling is associated with NOTCH signaling defects and abnormal arteriovenous EC differentiation. Moreover, cumulative evidence suggests that AVMs originate from venous ECs with defective flow-migration coupling and excessive proliferation. Mutant ECs show an increase of PI3K/AKT signaling and inhibitors of this signaling pathway rescue AVMs in HHT mouse models, revealing new therapeutic avenues. In this review, we will summarize recent advances and current knowledge of mechanisms controlling the pathogenesis of bAVMs, and discuss unresolved questions.
Endoglin (ENG) is expressed on the surface of endothelial cells (ECs) where it efficiently binds circulating BMP9 and BMP10 ligands to initiate activin A receptor like type 1 (ALK1) protein signalling to protect the vascular architecture. Patients heterozygous for ENG or ALK1 mutations develop the vascular disorder known as hereditary haemorrhagic telangiectasia (HHT). Many patients with this disorder suffer from anaemia, and are also at increased risk of stroke and high output heart failure. Recent work using animal models of HHT has revealed new insights into cellular and molecular mechanisms causing this disease. Loss of the ENG (HHT1) or ALK1 (HHT2) gene in ECs leads to aberrant arteriovenous connections or malformations (AVMs) in developing blood vessels. Similar phenotypes develop following combined EC specific loss of SMAD1 and 5, or EC loss of SMAD4. Taken together these data point to the essential role of the BMP9/10-ENG-ALK1-SMAD1/5-SMAD4 pathway in protecting the vasculature from AVMs. Altered directional migration of ECs in response to shear stress and increased EC proliferation are now recognised as critical factors driving AVM formation. Disruption of the ENG/ALK1 signalling pathway also affects EC responses to vascular endothelial growth factor (VEGF) and crosstalk between ECs and vascular smooth muscle cells. It is striking that the vascular lesions in HHT are both localised and tissue specific. Increasing evidence points to the importance of a second genetic hit to generate biallelic mutations, and the sporadic nature of such somatic mutations would explain the localised formation of vascular lesions. In addition, different pro-angiogenic drivers of AVM formation are likely to be at play during the patient’s life course. For example, inflammation is a key driver of vessel remodelling in postnatal life, and may turn out to be an important driver of HHT disease. The current wealth of preclinical models of HHT has led to increased understanding of AVM development and revealed new therapeutic approaches to treat AVMs, and form the topic of this review.
Endothelial cells (ECs) line the luminal surface of blood and lymphatic vessels, where they serve as a selective barrier between blood and lymphatic circulation and tissue parenchyma throughout the body. In all tissues, ECs support the movement and exchange of oxygen, nutrients, paracrine signals, lymph, and immune cells; however, ECs also serve distinct tissue-specific functions. To accomplish these diverse functions, ECs must acquire heterogeneous phenotypes and unique structural features during organogenesis and maintain these properties in adult organs. Heterogeneity begins early in development when primordial ECs are specified into arterial, venous, capillary, lymphatic, or hemogenic fates. In some large vessels, initial EC fate specification is genetically determined; however, increasing evidence reveals the importance of the immediate microenvironment in this process. Tissue- and vessel-specific environmental factors, such as hemodynamic forces, hypoxia, and metabolic state, coordinately promote a broad range of EC phenotypes throughout the body. Epigenetic regulation and noncoding RNAs also play important roles in establishing and maintaining this phenotypic diversity. In this chapter, we briefly discuss the signaling pathways that regulate EC specification, and highlight the contribution of environmental factors and epigenetic regulation during organogenesis and in adult organs. Elucidating these governing modalities of EC phenotypic diversification will confer a more complete understanding of vascular development and function that will ultimately provide insights needed to improve clinical treatments for vascular disorders.
Vascular anomalies are developmental defects of the vasculature and encompass a variety of disorders. The identification of genes mutated in the different malformations provides insight into the etiopathogenic mechanisms and the specific roles the associated proteins play in vascular development and maintenance. A few familial forms of vascular anomalies exist, but most cases occur sporadically. It is becoming evident that somatic mosaicism plays a major role in the formation of vascular lesions. The use of Next Generating Sequencing for high throughput and "deep" screening of both blood and lesional DNA and RNA has been instrumental in detecting such low frequency somatic changes. The number of novel causative mutations identified for many vascular anomalies has soared within a 10-year period. The discovery of such genes aided in unraveling a holistic overview of the pathogenic mechanisms, by which in vitro and in vivo models could be generated, and opening the doors to development of more effective treatments that do not address just symptoms. Moreover, as many mutations and the implicated signaling pathways are shared with cancers, current oncological therapies could potentially be repurposed for the treatment of vascular anomalies.
Rationale: Hereditary hemorrhagic telangiectasia (HHT) is a genetic disease caused by mutations in ENG, ALK1, or SMAD4. Since proteins from all three HHT genes are components of signal transduction of TGF-β family members, it has been hypothesized that HHT is a disease caused by defects in the ENG-ALK1-SMAD4 linear signaling. However, in vivo evidence supporting this hypothesis is scarce.
Objective: We tested this hypothesis and investigated the therapeutic effects and potential risks of induced-ALK1 or -ENG overexpression for HHT.
Methods and Results: We generated a novel mouse allele (ROSA26 Alk1 ) in which HA-tagged ALK1 and bicistronic eGFP expression are induced by Cre activity. We examined whether ALK1-overexpression (OE) using the ROSA26 Alk1 allele could suppress the development of AVMs in wounded adult skin and developing retinas of Alk1- and Eng-inducible knockout (iKO) mice. We also used a similar approach to investigate whether ENG-OE could rescue AVMs. Biochemical and immunofluorescence analyses confirmed the Cre-dependent overexpression of the ALK1-HA transgene. We could not detect any pathological signs in ALK1-OE mice up to 3 months after induction. ALK1-OE prevented the development of retinal AVMs and wound-induced skin AVMs in Eng-iKO as well as Alk1-iKO mice. ALK1-OE normalized expression of SMAD and NOTCH target genes in ENG-deficient endothelial cells (ECs) and restored the effect of BMP9 on suppression of phosphor-AKT levels in these ECs. On the other hand, ENG-OE could not inhibit the AVM development in Alk1-iKO models.
Conclusions: These data support the notion that ENG and ALK1 form a linear signaling pathway for the formation of a proper arteriovenous network during angiogenesis. We suggest that ALK1 overexpression or activation can be an effective therapeutic strategy for HHT1 and HHT2 in Alk1- and Eng-inducible knockout (iKO) mice. Further research is required to study whether this therapy could be translated into treatment for humans.
Vascular calcification is associated with atherosclerosis, chronic kidney disease, and diabetes, and results from processes resembling endochondral or intramembranous ossification, or from processes that are distinct from ossification. Bone morphogenetic proteins (BMP), as well as other ligands, receptors, and regulators of the transforming growth factor beta (TGFβ) family regulate vascular and valvular calcification by modulating the phenotypic plasticity of multipotent progenitor lineages associated with the vasculature or valves. While osteogenic ligands BMP2 and BMP4 appear to be both markers and drivers of vascular calcification, particularly in atherosclerosis, BMP7 may serve to protect against calcification in chronic kidney disease. BMP signaling regulators such as matrix Gla protein and BMP-binding endothelial regulator protein (BMPER) play protective roles in vascular calcification. The effects of BMP signaling molecules in vascular calcification are context-dependent, tissue-dependent, and cell-type specific. Here we review the current knowledge on mechanisms by which BMP signaling regulates vascular calcification and the potential therapeutic implications.
Background:
While prior studies reporting de novo arteriovenous malformations (AVMs) after birth are not uncommon, reports of ectopic recurrence of AVM after radiotherapy are scarce.
Case description:
Here, we present an 8-year-old girl with a ruptured cerebellar AVM treated with stereotactic radiosurgery (SRS). The nidus was completely obliterated 17 months after the SRS; however, 20 years later she had a rupture of a recurrent nidus that occurred adjacent to the original nidus, accompanied with a radiation-induced cyst. A surgical resection was performed, and a subsequent pathological examination revealed two different compartments: a typical nidus and a post-irradiated degenerated nidus with an encapsulated hematoma. Immunohistochemical stains showed an increased expression of Ki-67 and decreased expression of endoglin in endothelial cells in the recurrent nidus.
Conclusions:
Depletion in endoglin may lead to the formation of vessel malformations in the presence of angiogenic stimuli. This case serves as a strong reminder of the importance of long-term follow-up after SRS, especially for pediatric cases.
Hereditary hemorrhagic telangiectasia (HHT) is a genetic disorder characterized by multi-systemic vascular dysplasia affecting 1 in 5000 people worldwide. Individuals with HHT suffer from many complications including nose and gastrointestinal bleeding, anemia, iron deficiency, stroke, abscess, and high-output heart failure. Identification of the causative gene mutations and the generation of animal models have revealed that decreased transforming growth factor-β (TGF-β)/bone morphogenetic protein (BMP) signaling and increased vascular endothelial growth factor (VEGF) signaling activity in endothelial cells are responsible for the development of the vascular malformations in HHT. Perturbations in these key pathways are thought to lead to endothelial cell activation resulting in mural cell disengagement from the endothelium. This initial instability state causes the blood vessels to response inadequately when they are exposed to angiogenic triggers resulting in excessive blood vessel growth and the formation of vascular abnormalities that are prone to bleeding. Drugs promoting blood vessel stability have been reported as effective in preclinical models and in clinical trials indicating possible interventional targets based on a normalization approach for treating HHT. Here, we will review how disturbed TGF-β and VEGF signaling relates to blood vessel destabilization and HHT development and will discuss therapeutic opportunities based on the concept of vessel normalization to treat HHT.
Activin A receptor like type 1 (ALK1) is a transmembrane serine/threonine receptor kinase in the transforming growth factor-beta receptor family that is expressed on endothelial cells. Defects in ALK1 signaling cause the autosomal dominant vascular disorder, hereditary hemorrhagic telangiectasia (HHT), which is characterized by development of direct connections between arteries and veins, or arteriovenous malformations (AVMs). Although previous studies have implicated ALK1 in various aspects of sprouting angiogenesis, including tip/stalk cell selection, migration, and proliferation, recent work suggests an intriguing role for ALK1 in transducing a flow-based signal that governs directed endothelial cell migration within patent, perfused vessels. In this review, we present an updated view of the mechanism of ALK1 signaling, put forth a unified hypothesis to explain the cellular missteps that lead to AVMs associated with ALK1 deficiency, and discuss emerging roles for ALK1 signaling in diseases beyond HHT.
BMPs regulate multiple essential processes contributing to the formation and homeostasis of the vascular system. Here we describe the impact of BMP signaling in mesoderm formation, vasculogenesis, arteriovenous differentiation, sprouting angiogenesis, endothelial-to-mesenchymal transition (EndMT), and the barrier function of the endothelium.
Aberrant signaling during vascular morphogenesis as well as morphogenic processes, such as endothelial-to-mesenchymal transition (EndMT), have been implicated in several pathological conditions, including tumor neovascularization, hereditary hemorrhagic telangiectasia (HHT), cerebral cavernous malformation (CCM), and fibrodysplasia ossificans progressiva (FOP). We emphasize the molecular mechanisms underlying BMP-dependent regulations of endothelial cell functions and highlight possible applications in the treatment of vascular diseases.
This chapter discusses arteriovenous patterning in the vascular system. One of the primary distinctions in the mammalian vasculature is its division into arterial and venous systems. These simple diagrams underscore the functional, anatomical and structural differences that exist between arteries and veins. While both vessel types are lined by a thin inner layer of endothelial cells, arteries have a thicker vessel wall with more elastic fibers and vascular smooth muscle cells to support the higher blood pressure in arteries. Veins, on the other hand, contain valves to prevent retrograde flow of blood. Acquisition of these morphological and structural differences has long been attributed to the physiological factors that differ between these two vessel types. Recent work has established that, contrary to the historic view, genetic prepatterning prior to the onset of circulation is a primary determinant in regulating the differentiation of arteries and veins. However, this genetic prepattern is plastic, and is modified and optimized by environmental cues such as hemodynamic flow.
Members of the transforming growth factor-β (TGF-β) family exert their effect virtually on all cell types in the body, producing diverse and complex cellular responses. TGF-β signaling is deregulated and hyperactive in many malignant conditions, making it an appealing target in the combat of cancer disease. The predominantly endothelial TGF-β receptors, ALK1 and endoglin, which are activated during neoangiogenesis both during development and pathological conditions, pose attractive modulating opportunities to impair tumor veβel formation and cancer progreβion. However, the precise function of TGF-β family signaling in ECs is difficult to predict, as it appears highly context dependent due to the many ligands and receptors influencing the final outcome. Furthermore, TGF-β is involved in autocrine and intricate dynamic paracrine signaling events in the context of the tumor microenvironment. Pharmacological inhibitors for ALK1, endoglin, and TGF-β or its receptors have been developed and will facilitate more comprehensive studies on the exact function of the TGF-β family in the endothelium, and more specifically in tumor angiogenesis. Here, we will summarize the current knowledge on TGF-β signaling in the regulation of the formation and function of the vascular network.
Bone morphogenetic proteins (BMPs) are growth factors of the TGFβ family. Among them, BMP9 and BMP10 have been shown to bind with high affinity to a receptor expressed only on blood and lymphatic endothelial cells, ALK1 (activin receptor-like kinase 1), suggesting an important role for BMP9, BMP10, and ALK1 in vascular development. Indeed, mutations of ALK1 have been linked to two vascular diseases, the Rendu-Osler-Weber syndrome, also known as hemorrhagic hereditary telangiectasia, and pulmonary arterial hypertension. BMP9 is expressed by the liver, while BMP10 is mainly expressed by the heart and both are present in blood. The current working model is that BMP9 and BMP10 via ALK1 maintain a quiescent endothelial state. BMP9 and BMP10 have been recently shown to play a critical role in blood vessel development in an interchangeable manner. On the other hand, preliminary works seem to indicate that BMP9 plays a specific role in lymphatic development. Although the cellular functions of ALK1 are not completely understood, therapeutic treatment blocking ALK1 have already been developed. Two different approaches are ongoing: one, using the extracellular domain of ALK1 (ALK1ECD) that will trap any ligand able to bind to this soluble form of ALK1 and, the second, using a blocking anti-ALK1 antibody. Preliminary data from phase 1 clinical trials indicate no major toxicity with these two compounds and phase 2 clinical trials have started in patients suffering from different cancers. Taken together, these results clearly demonstrate that the BMP9/BMP10/ALK1 is an emerging vascular signaling pathway with potential therapeutic applications.
Hereditary hemorrhagic telangiectasia (HHT) is an autosomal-dominant, highly penetrant and variable multisystem disorder due to maldevelopment of capillaries. The condition used to be called Osler–Weber–Rendu syndrome. The major features are epistaxis; mucocutaneous telangiectasias; and visceral arteriovenous malformations (especially in the brain, gastrointestinal tract, liver and lung). The causes defined thus far are in genes involved in signaling through the transforming growth factor β/bone morphogenetic protein pathway in endothelial cells. Early diagnosis, clinical screening and aggressive management can greatly reduce morbidity and mortality.
Hereditary haemorrhagic telangiectasia (HHT) is an autosomal dominant disorder characterized by multisystemic vascular dysplasia and recurrent haemorrhage. Linkage for some families has been established to chromosome 9q33-q34. In the present study, endoglin, a transforming growth factor beta (TGF-beta) binding protein, was analysed as a candidate gene for the disorder based on chromosomal location, expression pattern and function. We have identified mutations in three affected individuals: a C to G substitution converting a tyrosine to a termination codon, a 39 base pair deletion and a 2 basepair deletion which creates a premature termination codon. We have identified endoglin as the HHT gene mapping to 9q3 and have established HHT as the first human disease defined by a mutation in a member of the TGF-beta receptor complex.
We studied 10 cutaneous telangiectatic lesions of hereditary hemorrhagic telangiectasia (HHT), ranging in size from pinpoint to 2 mm, by light and electron microscopy. Four representative lesions were reconstructed by computer from serial 1- or 2-mm plastic embedded sections. The earliest clinically detectable lesion of HHT is a focal dilatation of postcapillary venules, which continue to enlarge and eventually connect with dilated arterioles through capillaries. As the vascular lesion increases in size, the capillary segments disappear and a direct arterio-venous communication is formed. This entire sequence of morphologic events is associated with a perivascular mononuclear cell infiltrate in which the majority of cells are lymphocytes and the minority are monocytes/macrophages by ultrastructure. Comparison of these findings with the telangiectatic mats of scleroderma and cherry angiomas revealed that the former, previously shown to be composed of dilated postcapillary venules, are also associated with perivascular infiltrates, but the latter, which are produced by capillary loop aneurysms, are not.
Vascular smooth muscle cells (SMCs) produce the bulk of the connective tissue of major arteries, including collagen types I, III, and V. Recently, we have shown, they also have the capacity to synthesize the alpha 1 chain of type XI, a collagen related to type V (Brown, K., Lawrence, R., and Sonenshein, G. (1991) J. Biol. Chem. 266, 23268-23273). Furthermore, expression of types V and XI collagen were coordinately regulated with respect to serum deprivation and cell density in a fashion distinct from that for types I and III. To begin to determine the factors that influence vascular SMC production of types V/XI collagen, we have examined the effects of transforming growth factor (TGF)-beta 1, a major modulator of connective tissue expression. In serum-deprived confluent cultures of bovine pulmonary artery SMCs, TGF-beta 1 treatment increased the steady-state levels of the mRNAs of collagen types V and XI, as well as of types I and III, elastin and fibronectin. The largest increase was seen for alpha 2(V) procollagen. The increase in alpha 2(V) mRNA was detectable by 12 h after addition of 2 ng/ml TGF-beta 1, and concentrations as little as 0.5 ng/ml were effective. A similar increase in alpha 2(V) mRNA levels was observed with SMCs derived from the aortic arch and carotid artery. Type V collagen protein was found to be elevated by TGF-beta 1 treatment in both the conditioned media and the cell layer associated fraction of pulse-labeled cultures. A slight decrease in SMC proliferation as judged by DNA content, [3H]thymidine incorporation, and steady-state levels of histone H3.2 mRNA resulted from TGF-beta 1 treatment. These results suggest that the elevated levels of TGF-beta 1 in the vessel wall during atherosclerosis may be, in part, responsible for the increase in type V collagen that typifies advanced fibrotic lesions.
We aimed to determine if and how endothelial cells (EC) recruit precursors of smooth muscle cells and pericytes and induce their differentiation during vessel formation. Multipotent embryonic 10T1/2 cells were used as presumptive mural cell precursors. In an under-agarose coculture, EC induced migration of 10T1/2 cells via platelet-derived growth factor BB. 10T1/2 cells in coculture with EC changed from polygonal to spindle-shaped, reminiscent of smooth muscle cells in culture. Immunohistochemical and Western blot analyses were used to examine the expression of smooth muscle (SM)-specific markers in 10T1/2 cells cultured in the absence and presence of EC. SM-myosin, SM22alpha, and calponin proteins were undetectable in 10T1/2 cells cultured alone; however, expression of all three SM-specific proteins was significantly induced in 10T1/2 cells cocultured with EC. Treatment of 10T1/2 cells with TGF-beta induced phenotypic changes and changes in SM markers similar to those seen in the cocultures. Neutralization of TGF-beta in the cocultures blocked expression of the SM markers and the shape change. To assess the ability of 10T1/2 cells to contribute to the developing vessel wall in vivo, prelabeled 10T1/2 cells were grown in a collagen matrix and implanted subcutaneously into mice. The fluorescently marked cells became incorporated into the medial layer of developing vessels where they expressed SM markers. These in vitro and in vivo observations shed light on the cell-cell interactions that occur during vessel development, as well as in pathologies in which developmental processes are recapitulated.
Endoglin (CD105) is a transmembrane glycoprotein that binds transforming growth factor (TGF)-β1 and -β3, and coprecipitates
with the Ser/Thr kinase signaling receptor complex by affinity labeling of endothelial and leukemic cells. The present study
shows that in addition to TGF-β1 and -β3, endoglin interacts with activin-A, bone morphogenetic protein (BMP)-7, and BMP-2
but requires coexpression of the respective ligand binding kinase receptor for this association. Endoglin cannot bind ligands
on its own and does not alter binding to the kinase receptors. It binds TGF-β1 and -β3 by associating with the TGF-β type
II receptor and interacts with activin-A and BMP-7 via activin type II receptors, ActRII and ActRIIB, regardless of which
type I receptor partner is coexpressed. However, endoglin binds BMP-2 by interacting with the ligand binding type I receptors,
ALK3 and ALK6. The formation of heteromeric signaling complexes was not altered by the presence of endoglin, although it was
coprecipitated with these complexes. Endoglin did not interact with BMP-7 through complexes containing the BMP type II receptor,
demonstrating specificity of its action. Our data suggest that endoglin is an accessory protein of multiple kinase receptor
complexes of the TGF-β superfamily.
Endoglin is a transforming growth factor-beta (TGF-beta) binding protein expressed on the surface of endothelial cells. Loss-of-function mutations in the human endoglin gene ENG cause hereditary hemorrhagic telangiectasia (HHT1), a disease characterized by vascular malformations. Here it is shown that by gestational day 11.5, mice lacking endoglin die from defective vascular development. However, in contrast to mice lacking TGF-beta, vasculogenesis was unaffected. Loss of endoglin caused poor vascular smooth muscle development and arrested endothelial remodeling. These results demonstrate that endoglin is essential for angiogenesis and suggest a pathogenic mechanism for HHT1.
Endoglin (CD105), an accessory protein of the TGF-beta receptor superfamily, is highly expressed on endothelial cells. Hereditary hemorrhagic telangiectasia type 1 (HHT1) is associated with mutations in the Endoglin gene, leading to haploinsufficiency. To generate a disease model and ascertain the role of endoglin in development, we generated mice lacking 1 or both copies of the gene. Endoglin null embryos die at gestational day 10.0-10.5 due to defects in vessel and heart development. Vessel formation appears normal until hemorrhage occurs in yolk sacs and embryos. The primitive vascular plexus of the yolk sac fails to mature into defined vessels, and vascular channels dilate and rupture. Internal bleeding is seen in the peritoneal cavity, implying fragile vessels. Heart development is arrested at day 9.0, and the atrioventricular canal endocardium fails to undergo mesenchymal transformation and cushion-tissue formation. These data suggest that endoglin is critical for both angiogenesis and heart valve formation. Some heterozygotes, either with an inbred 129/Ola or mixed C57BL/6-129/Ola background, show signs of HHT, such as telangiectases or recurrent nosebleeds. In this murine model of HHT, it appears that epigenetic factors and modifier genes, some of which are present in 129/Ola, contribute to disease heterogeneity.
ALK-1 (activin receptor-like kinase-1), a type I receptor of the transforming growth factor (TGF)-beta superfamily, is the gene mutated in hereditary hemorrhagic telangiectasia type 2 (HHT2) while endoglin is mutated in HHT1. Using a novel polyclonal antibody to ALK-1, we measured ALK-1 expression on human umbilical vein endothelial cells (HUVEC) of newborns from HHT families whose affected members had normal endoglin levels. ALK-1 levels were specifically reduced in three HUVEC with ALK-1 missense mutant codons, and normal in two newborns not carrying the missense mutations present in the clinically affected relatives. Levels were also normal in a HUVEC with deletion of S232 in the ATP binding site of ALK-1. Thus HHT2 appears to be associated with a loss of function of the mutant allele due to a reduction in either protein level or activity. We also report three new ALK-1 missense mutations leading to G48E/A49P, C344Y and E407D substitutions. In COS-1 transfected cells, ALK-1 was found in the TGF-beta1 and -beta3 receptor complexes in association with endoglin and TbetaRII, but not in activin receptor complexes containing endoglin. In HUVEC, ALK-1 was not detectable in the TGF-beta1 or -beta3 receptor complexes. However, in the absence of ligand, ALK-1 and endoglin interactions were observed by immunoprecipitation/western blot in HUVEC from normal as well as HHT1 and HHT2 patients. Our data suggest a transient association between these two proteins of the TGF-beta superfamily, both required at a critical level to ensure vessel wall integrity.
Endoglin is an auxiliary component of the transforming growth factor-beta (TGF-beta) receptor system, able to associate with the signaling receptor types I (TbetaRI) and II (TbetaRII) in the presence of ligand and to modulate the cellular responses to TGF-beta1. Endoglin cannot bind ligand on its own but requires the presence of the signaling receptors, supporting a critical role for the interaction between endoglin and TbetaRI or TbetaRII. This study shows that full-length endoglin interacts with both TbetaRI and TbetaRII, independently of their kinase activation state or the presence of exogenous TGF-beta1. Truncated constructs encoding either the extracellular or the cytoplasmic domains of endoglin demonstrated that the association with the signaling receptors occurs through both extracellular and cytoplasmic domains. However, a more specific mapping revealed that the endoglin/TbetaRI interaction was different from that of endoglin/TbetaRII. TbetaRII interacts with the amino acid region 437-558 of the extracellular domain of endoglin, whereas TbetaRI interacts not only with the region 437-558 but also with the protein region located between amino acid 437 and the N terminus. Both TbetaRI and TbetaRII interact with the cytoplasmic domain of endoglin, but TbetaRI only interacts when the kinase domain is inactive, whereas TbetaRII remains associated in its active and inactive forms. Upon association, TbetaRI and TbetaRII phosphorylate the endoglin cytoplasmic domain, and then TbetaRI, but not TbetaRII, kinase dissociates from the complex. Conversely, endoglin expression results in an altered phosphorylation state of TbetaRII, TbetaRI, and downstream Smad proteins as well as a modulation of TGF-beta signaling, as measured by the reporter gene expression. These results suggest that by interacting through its extracellular and cytoplasmic domains with the signaling receptors, endoglin might affect TGF-beta responses.
Because angiogenesis and inflammation are intimately associated and endoglin is required for angiogenesis, we wished to determine whether it also plays a role in inflammation.
Using an immunohistochemical approach, we examined spatial and temporal changes in endoglin expression during inflammation and angiogenesis.
We found low levels of endoglin expression in quiescent endothelium in a range of normal adult human tissues. However, constitutive levels of expression are higher in dermal capillaries surrounding hair follicles and in alveolar capillaries as well as in the high endothelial cells of lymph tissue. During inflammatory disease, endoglin expression is strongly upregulated and is consistently associated with an infiltrate of inflammatory cells. For the first time, we have determined the relative changes in endoglin expression from the normal quiescent state through the inflammatory changes and angiogenesis that occur during dermal wound healing in vivo using a timed human wound healing model. Endoglin expression increases rapidly, reaching a peak of expression co-incident with maximal T cell infiltrate and persisting at an elevated level for at least 28 days in both activated and proliferating endothelial cells.
Enhanced endoglin expression is not limited to angiogenesis, it is also associated with inflammation.
Endoglin is a transforming growth factor–β (TGF-β) binding protein expressed on the surface of endothelial cells. Loss-of-function
mutations in the human endoglin gene ENGcause hereditary hemorrhagic telangiectasia (HHT1), a disease characterized by vascular malformations. Here it is shown that
by gestational day 11.5, mice lacking endoglin die from defective vascular development. However, in contrast to mice lacking
TGF-β, vasculogenesis was unaffected. Loss of endoglin caused poor vascular smooth muscle development and arrested endothelial
remodeling. These results demonstrate that endoglin is essential for angiogenesis and suggest a pathogenic mechanism for HHT1.
ALK-1 (activin receptor-like kinase-1), a type I receptor of the transforming growth factor (TGF)-β superfamily, is the gene mutated in hereditary hemorrhagic telangiectasia type 2 (HHT2) while endoglin is mutated in HHT1. Using a novel polyclonal antibody to ALK-1, we measured ALK-1 expression on human umbilical vein endothelial cells (HUVEC) of newborns from HHT families whose affected members had normal endoglin levels. ALK-1 levels were specifically reduced in three HUVEC with ALK-1 missense mutant codons, and normal in two newborns not carrying the missense mutations present in the clinically affected relatives. Levels were also normal in a HUVEC with deletion of S232 in the ATP binding site of ALK-1. Thus HHT2 appears to be associated with a loss of function of the mutant allele due to a reduction in either protein level or activity. We also report three new ALK-1 missense mutations leading to G48E/A49P, C344Y and E407D substitutions. In COS-1 transfected cells, ALK-1 was found in the TGF-β1 and -β3 receptor complexes in association with endoglin and TβRII, but not in activin receptor complexes containing endoglin. In HUVEC, ALK-1 was not detectable in the TGF-β1 or -β3 receptor complexes. However, in the absence of ligand, ALK-1 and endoglin interactions were observed by immunoprecipitation/western blot in HUVEC from normal as well as HHT1 and HHT2 patients. Our data suggest a transient association between these two proteins of the TGF-β superfamily, both required at a critical level to ensure vessel wall integrity.
Transforming growth factor-beta s (TGF-beta) are potent enhancers of the expression of several connective tissue genes. In this study we examined the effects of TGF-beta 1 and TGF-beta 2 on human elastin mRNA abundance, promoter activity, and mRNA stability in cultured human skin fibroblasts. Treatment of cell cultures with varying concentrations of TGF-beta 1 or TGF-beta 2 for 24 hours resulted in a dose-dependent increase in the elastin mRNA steady-state levels, with a maximum enhancement of approximately 30-fold being noted with 1 ng/ml. Addition of cycloheximide (10 micrograms/ml) failed to block up-regulation of elastin gene expression by TGF-beta, indicating that this effect can occur in the absence of active protein synthesis. Furthermore, TGF-beta elicited enhancement of elastin mRNA levels could be abrogated by tumor necrosis factor-alpha and partially counteracted by interferon-gamma. Transient transfections of human skin fibroblasts with elastin promoter/chloramphenicol acetyl-transferase reporter gene constructs, which contained up to approximately 5 kb of the 5' flanking DNA, revealed no change in the promoter activity in the presence of TGF-beta. However, TGF-beta appeared to stabilize the elastin mRNA transcripts as determined by Northern hybridizations after inhibition of initiation of the transcription. As a result of this stabilization, the elastin mRNA levels were clearly detectable in TGF-beta 1-treated cultures even up to 48 hours after inhibition of transcription while they were undetectable in the control cells after 24 hours of incubation. These results demonstrate that TGF-beta 1 and TGF-beta 2 are potent enhancers of elastin gene expression and that this effect is mediated, at least in part, post-transcriptionally. These results suggest that TGF-beta s are involved in regulation of elastin deposition during fetal development and tissue repair, as well as in pathological conditions.
The integrity of the hepatic portal vasculature was examined, relative to the resistance to Schistosoma mansoni observed in 68% of 129/Ola mice. The passage of microspheres to the lungs, following their injection via the superior mesenteric vein, indicated the presence of shunts in the majority of both naive and infected mice. There was a negative association between shunting of microspheres to the lungs and paucity of liver worms at 28/35 days post-infection. Schistosomula accumulated in the livers of resistant mice at a slower rate than in susceptible animals, and after day 21 relocated to the lungs. Many lung schistosomula injected via the superior mesenteric passed immediately to the lungs; the shunts thus greatly reduce the probability of trapping in the liver. Some parasites migrated back from the lungs, successfully lodged in the liver and began to feed on blood. Latex infusion demonstrated the location of large intrahepatic connections between the portal and hepatic veins. We suggest that as these liver worms grow, migrating upstream into progressively larger vessels, they reach the connections, pass out of the hepatic portal system, and relocate to the lungs. The presence of the natural shunts thus accounts for the resistant status of the mice.
The pulmonary and portal vasculature of naïve mice of the 129/Ola and CBA/Ca strains has been studied by means of the vasculature casting technique. This involve injection of pigmented vinylite resin into the arterial and venous systems, followed by digestion of the tissues with KOH. The peripheral vessels of the arterial and portal systems of CBA/Ca mice were numerous and highly branched. In contrast, casts prepared from 70-80% of naïve 129/Ola mice showed dramatic reductions in the number and extent of the peripheral vessels. In addition, such vessels appeared severely truncated. The remaining 20-30% of naïve 129/Ola mice yielded lung and liver casts that were indistinguishable from the CBA/Ca casts. Casts prepared from 129/Ola mice infected 6 weeks previously with Schistosoma mansoni cercariae showed the same segregation; faecal smears, together with observations of presence or absence of gross pathology in such mice confirmed that the vascular changes correlated with the 'non-permissive trait'. We propose that such alterations facilitate the reportedly abnormal migration of schistosomes from the liver to the lungs in 'non-permissive' 129/Ola mice.
Mast cells (MC) are widely distributed throughout different organs with a relative predilection for potential portals of entry into the host. In tissues, MC are generally concentrated around small blood vessels and lymphatics, as well as nerves and glandular tissue. This close association with vascular structures suggests an important interaction between MC and endothelial cells (EC). Tissue MC are known to generate and release a number of mediators including histamine, prostaglandin D2, and leukotrienes that induce vasodilatation and increased vascular permeability. These MC-mediated effects on vessels may enhance responses to tissue injury or infection by facilitating the deposition of plasma components and inflammatory cells into involved sites. Important interactions between MC, EC, and peripheral nerves also occur. MC-derived histamine and possibly other mediators induce axon reflexes in unmyelinated sensory nerves leading to the release of neurotransmitters such as substance P, calcitonin gene-related peptide, and adenine nucleotides. These neurotransmitters exert direct effects on blood vessels, and in some instances, may act as MC stimulators. In vitro studies indicate that MC also affect EC growth and new blood vessel formation. Mast cell-derived heparin has been implicated as an important cofactor in tumor-induced angiogenesis, whereas histamine has been reported to augment human dermal EC growth in culture. The recent identification of a tumor necrosis factor-like peptide in MC and the reported cytostatic effects of TNF on EC suggest that MC may inhibit the proliferation of these cells under certain conditions. Taken together, these observations indicate that the interactions between MC and EC are important in both physiologic and pathologic events.
Elastin and collagen are the principal scleroproteins of the aortic wall, and they largely determine its physical and mechanical properties. During perinatal development of the aorta, elastin and collagen accumulate rapidly, being present as inverse gradients by the time of birth. Elastin is most prevalent in the thoracic aorta, decreasing distally, while collagen shows the opposite trend. The present studies have determined the relative and absolute rates of collagen and elastin synthesis in the porcine aorta between 60 days of fetal development (mid-gestation) and 110 days after birth. Although there was measurable elastin synthesis in the upper thoracic aorta at the earliest time evaluated, there was a fourfold increase in relative elastin synthesis (from 4 to 16% of total protein synthesis) between 60 fetal days and birth. Elastin synthesis was maximal in successively distal segments between 1 and 3 weeks after birth. Relative collagen synthesis progressively increased in distal aortic regions between 90 fetal days and 60 days postpartum. Greater than twofold increases over thoracic levels were measured. Both elastin and collagen synthesis largely subsided by 110 days of development. When expressed as absolute rates of protein synthesis, these scleroproteins were maximally expressed in the first 3 postnatal weeks. Elastin mRNA levels were determined with a cloned sheep gene fragment by molecular hybridization. Gradients of elastin message were present at 60 fetal days and at 4 and 14 days after birth, elastin mRNA levels being maximal in the upper thoracic aorta at 14 days after birth. The differentiation of the aortic wall thus follows discrete patterns of phenotypic change which may be coupled to the rheologic stresses accompanying development of the circulatory system.
Hereditary hemorrhagic telangiectasia is a rare autosomal dorminant disease that features abnormal and fragile vascular dilations of terminal vessels in skin and mucous membranes, as well as arteriovenous malformations of internal organs, particularly lungs, brain, and liver. Often patients have not been diagnosed with HHT for a long time, and undiagnosed HHT patients unnecessarily develop serious complications such as severe life-threatening hemorrhage, stroke or brain abscess. Therefore, early detection and appropriate screening is very important. Early detection of HHT allows the appropriate screening for the presence of silent disease such as AVMs in the lungs, liver, or brain, and preventive treatment in the patient and their affected family members. Dentists should be familiar with HHT because the telangiectases on skin and oral mucosa are often the most dramatic and most easily identified component of HHT. Recently, we experienced a case of HHT. We present the case with a review of the literature.
Transforming growth factor-beta (TGF-beta) is released after vascular injury and may influence the healing response of the vessel wall. We investigated the effect of this growth factor on proliferation and migration of human venous smooth muscle cells (SMC) and the signal transduction mechanisms through which TGF-beta exerts this effect.
SMC derived from human saphenous vein were used in a 72-hour proliferation assay and a 6-day migration assay. Cells were exposed to TGF-beta alone and in the presence of platelet-derived growth factor (PDGF), basic fibroblast growth factor (b-FGF), epidermal growth factor, and serum. The ability of TGF-beta to activate tyrosine kinases, phosphatases, or protein kinase C was evaluated by use of Western blotting with an antiphosphotyrosine antibody.
In a concentration-dependent manner, TGF-beta inhibited proliferation induced by PDGF, b-FGF, epidermal growth factor, and serum. This inhibitory effect was independent of SMC density. TGF-beta also inhibited migration induced by PDGF and b-FGF. Exposure of cells to TGF-beta did not lead to tyrosine phosphorylation of cellular substrates or activation of protein kinase C.
TGF-beta inhibits both migration and proliferation of human SMC. This inhibitory effect is not mediated through protein kinase C, mitogen-activated protein kinase, or tyrosine kinases.
Hereditary haemorrhagic telangiectasia (HHT) displays significant variation in severity between affected individuals in the same family, ranging from relatively trivial epistaxis and telangiectasia to gastrointestinal, cerebral and pulmonary involvement. Evidence for successful therapy of HHT-related haemorrhage with oestrogens and progesterones, and recent case reports outlining pulmonary complications of pregnancy in HHT, prompted us to review the outcome of 161 pregnancies in 47 affected women. HHT-related maternal complications developed in eleven patients, ten in the subgroup of 23 pregnancies in which pulmonary arteriovenous malformations (PAVMs) were present at the outset, or documented in the two years following pregnancy. We present six cases of intrapulmonary shunt deterioration, two cases of fatal pulmonary haemorrhage and three cerebrovascular accidents related to pregnancy. A predisposition towards PAVMs in females was observed. Following the recent discovery of mutations in the endoglin gene in this disease, our data support a hypothesis of hormonal modification of the HHT phenotype. In addition, a significant excess of affected offspring are present in HHT families. We consider contributary aetiological factors, and discuss implications for patient management.
Hereditary haemorrhagic telangiectasia, or Osler-Rendu-Weber (ORW) syndrome, is an autosomal dominant vascular dysplasia. So far, two loci have been demonstrated for ORW. Linkage studies established an ORW locus at chromosome 9q3; endoglin was subsequently identified as the ORW1 gene. A second locus, designated ORW2, was mapped to chromosome 12. Here we report a new 4 cM interval for ORW2 that does not overlap with any previously defined. A 1.38-Mb YAC contig spans the entire interval. It includes the activin receptor like kinase 1 gene (ACVRLK1 or ALK1), a member of the serine-threonine kinase receptor family expressed in endothelium. We report three mutations in the coding sequence of the ALK1 gene in those families which show linkage of the ORW phenotype to chromosome 12. Our data suggest a critical role for ALK1 in the control of blood vessel development or repair.
Pulmonary arteriovenous malformations (PAVMs) occur in up to 27% of patients with hereditary haemorrhagic telangiectasia (HHT) and are associated with a rate of paradoxical cerebral embolism at presentation of up to 36%. At least two different loci have been shown for HHT. Mutations in endoglin have been found in some families and the locus designated ORW1. In other families this locus has been excluded. In this paper we confirm that in families linked to ORW1 there is a prevalence of PAVMs among affected members of 29.2%, compared to a prevalence of 2.9% in families in which this locus has been excluded (chi 2 = 19.2, p < 0.001). This information can be used to decide how to screen HHT patients for PAVMs.
In this review we have summarized some of the evidence to support the view that mast cells play a critical role in leukocyte recruitment to sites of inflammation. Initially, data using a pharmacological tool, compound 48/80, which directly activates mast cells, is reviewed, demonstrating that this reagent can induce the multi-step recruitment of leukocytes (rolling, adhesion and emigration) to sites of inflammation. The adhesive mechanisms and pro-inflammatory mediators implicated in mast cell-induced leukocyte recruitment are discussed. Additionally, data are presented to implicate mast cells in delayed-type hypersensitivity reactions as they pertain to leukocyte recruitment. There is a growing body of evidence to suggest that mast cells also recruit leukocytes in IgE-independent leukocyte recruitment. Ischemia/reperfusion- and bacterial toxin- (Helicobacter pylori and Clostridium difficile) induced leukocyte recruitment is at least in part mast cell dependent. Future directions including preliminary work highlighting the role of nitric oxide as a modulator of mast cell function and subsequent leukocyte recruitment is also discussed.
Endoglin (CD105) is expressed on the surface of endothelial and haematopoietic cells in mammals and binds TGFbeta isoforms 1 and 3 in combination with the signaling complex of TGFbeta receptors types I and II. Endoglin expression increases during angiogenesis, wound healing, and inflammation, all of which are associated with TGFbeta signaling and alterations in vascular structure. The importance of endoglin for normal vascular architecture is further indicated by the association of mutations in the endoglin gene with the inherited disorder Hereditary Haemorrhagic Telangiectasia Type 1 (HHT1), a disease characterised by bleeding from vascular malformations. In order to study the role of endoglin in vivo in more detail and to work toward developing an animal model of HHT1, we have derived mice that carry a targeted nonsense mutation in the endoglin gene. Studies on these mice have revealed that endoglin is essential for early development. Embryos homozygous for the endoglin mutation fail to progress beyond 10.5 days postcoitum and fail to form mature blood vessels in the yolk sac. This phenotype is remarkably similar to that of the TGFbeta1 and the TGFbeta receptor II knockout mice, indicating that endoglin is needed in vivo for TGFbeta1 signaling during extraembryonic vascular development. In addition, we have observed cardiac defects in homozygous endoglin-deficient embryos, suggesting endoglin also plays a role in cardiogenesis. We anticipate that heterozygous mice will ultimately serve as a useful disease model for HHT1, as some individuals have dilated and fragile blood vessels similar to vascular malformations seen in HHT patients.
Endoglin is predominantly expressed on endothelium and is mutated in hereditary hemorrhagic telangiectasia (HHT) type 1 (HHT1). We report the analysis of endoglin in tissues of a newborn (family 2), who died of a cerebral arteriovenous malformation (CAVM), and in a lung specimen surgically resected from a 78-year-old patient (family 5), with a pulmonary AVM (PAVM). The clinically affected father of the newborn revealed a novel mutation that was absent in his parents and was identified as a duplication of exons 3 to 8, by quantitative multiplex polymerase chain reaction. The corresponding mutant protein (116-kd monomer) and the missense mutant protein (80-kd monomer) present in family 5 were detected only as transient intracellular species and were unreactive by Western blot analysis and immunostaining. Normal endoglin (90-kd monomer) was reduced by 50% on peripheral blood-activated monocytes of the HHT1 patients. When analyzed by immunostaining and densitometry, presumed normal blood vessels of the newborn lung and brain and vessels adjacent to the adult PAVM showed a 50% reduction in the endoglin/PECAM-1 ratio. A similar ratio was observed in the CAVM and PAVM, suggesting that all blood vessels of HHT1 patients express reduced endoglin in situ and that AVMs are not attributed to a focal loss of endoglin.
Pulmonary arteriovenous malformations (PAVMs) occur in over 25% of patients with the autosomal dominant disorder hereditary haemorrhagic telangiectasia (HHT). Mutations in two genes, endoglin and ALK-1, are known to cause HHT. Each encodes a protein expressed on vascular endothelial cells and involved in signalling by members of the transforming growth factor (TGF)-beta superfamily. To date, PAVMs have not been detected in ALK-1 families. There is evidence from a single HHT family without pulmonary involvement that a third HHT gene may exist. To establish the existence of a further HHT gene responsible for PAVMs, linkage analyses were performed on an expanded PAVM-HHT family in which HHT did not result from endoglin mutations.
Family members were assessed clinically to assign HHT disease status and were screened for PAVMs. DNA was extracted from blood obtained from 20 individuals of known disease status. Short tandem repeat polymorphic markers spanning the intervals containing the endoglin and ALK-1 genes were amplified by the polymerase chain reaction using (33)P-labelled oligonucleotide primers, separated by denaturing polyacrylamide gel electrophoresis (PAGE), and the resultant autoradiographs were examined for allele sizes. Linkage analyses were performed using MLINK and GENEHUNTER.
Twelve members spanning four generations were affected with HHT. Two had proven PAVMs, one with a classical appearance, the other exhibiting microscopic PAVMs exacerbated by pregnancy. Two point lod and multipoint lod scores significantly excluded linkage to endoglin and ALK-1 in this pedigree.
This study confirms the existence of a third HHT locus that accounts for disease in some HHT patients with pulmonary involvement.
Endoglin (CD105) is a cell membrane glycoprotein over-expressed on highly proliferating endothelial cells in culture, and on endothelial cells of angiogenetic blood vessels within benign and malignant tissues. CD105 binds several factors of the Transforming Growth Factor (TGF)-beta superfamily, and its over-expression modulates cellular responses to TGF-beta1. The complex of experimental findings accumulated in the last few years strongly indicate that CD105 is a powerful marker of angiogenesis, and that it might play a critical role in the pathogenesis of vascular diseases and in tumor progression. In this paper, we will review the structural, biological and functional features of CD105, as well as its distribution within normal and neoplastic tissues, emphasizing its foreseeable role as a molecular target for new diagnostic and bioimmunotherapeutic approaches in human malignancies.
Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant disorder because of mutations in the genes coding for endoglin (HHT1) or ALK-1 (HHT2). The disease is associated with haploinsufficiency and a murine model was obtained by engineering mice that express a single Endoglin allele. Of a total of 171 mice that were observed for 1 year, 50 developed clinical signs of HHT. Disease prevalence was high in 129/Ola strain (72%), intermediate in the intercrosses (36%), and low in C57BL/6 backcrosses (7%). Most mice first presented with an ear telangiectasia and/or recurrent external hemorrhage. One-third of mice with HHT showed severe vascular abnormalities such as dilated vessels, hemorrhages, liver and lung congestion, and/or brain and heart ischemia. Disease sequelae included stroke, hydrocephalus, fatal hemorrhage, and congestive heart failure. Thus the murine model reproduces the multiorgan manifestations of the human disease. Levels of circulating latent transforming growth factor (TGF)-beta1 were significantly lower in the 129/Ola than in the C57BL/6 strain. Intercrosses and 129/Ola mice expressing reduced endoglin also showed lower plasma TGF-beta1 levels than control. These data suggest that modifier genes involved in the regulation of TGF-beta1 expression act in combination with a single functional copy of endoglin in the development of HHT.
Endoglin is an auxiliary receptor for the transforming growth factor-beta family of cytokines and is required for angiogenesis and heart development. Endoglin expression during mouse embryogenesis was analysed by monitoring beta-galactosidase expression from a lacZ reporter cassette inserted downstream of the endoglin promoter. Expression was first detected at 6.5 days post-coitum (dpc) in the amniotic fold and developing allantois. Between 7.5 and 8.5 dpc, endoglin was expressed in endothelial cells of the yolk sac, dorsal aorta and primitive heart tube, and from 9.5 to 13.5 dpc in endothelial cells throughout the developing vasculature. Interestingly, this pattern of endoglin expression is almost identical to that reported for Alk1.
An electron microscopic study in Osler's disease
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A murine model of hereditary hemorrhagic telangiectasia
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