Identification of a Novel EIF2AK Variant and Genetics-Assisted Approach to Diagnosis of Pulmonary Capillary Hemangiomatosis.

Pulmonary capillary hemangiomatosis (PCH) is an uncommon type of pulmonary vascular disease characterized by capillary proliferation and very poor prognosis owing to misdiagnosis and lack of effective therapeutic options. Mutations in the eukaryotic translation initiation factor 2α kinase 4 (EIF2AK4) gene have been reported in pulmonary veno-occlusive disease and PCH. In this report, we present a patient whose diagnosis of PCH was delayed by 2 ½ years despite prior surgical lung biopsy and clinical and laboratory findings suggestive of pulmonary hypertension. Genotyping revealed a novel likely pathogenic variant in the EIF2AK4 gene. Review of surgical lung biopsy performed 2 ½ years prior confirmed PCH histology along with constrictive bronchiolitis.

News on infantile haemangioma. Part 1: clinical course and pathomechanism.

Currently, there is no doubt that the first choice of treatment for alarming infantile haemangiomas (IHs) is oral beta-blockers. However, research in this field remains active, as the pathogenesis of IH is still not completely elucidated. Furthermore, there are different approaches to the management of IHs with beta-blockers. In Part 1 of this review we will discuss the state-of-the-art evidence for IH with regard to (i) the definition, epidemiology, course, risk factors and sequelae, and (ii) the pathogenesis, focusing on genetic studies. This review will update the reader on the latest developments in the pathogenesis of IH. Furthermore, we hope this review will give more insight into risk factors and sequelae of IH, thereby contributing to better decisions in the clinical management of patients with IH. The therapy and evaluation of IHs will be discussed in Part 2 of this review.

Pulmonary capillary hemangiomatosis in Chinese patients without EIF2AK4 mutations.

BACKGROUND: Pulmonary capillary hemangiomatosis (PCH) is a very rare and refractory pulmonary vascular disease that causes pulmonary hypertension. Differentiation of PCH from idiopathic pulmonary arterial hypertension (iPAH) is essential because treatment and prognosis can vary greatly between these two diseases. CASE PRESENTATION: A 20-year-old female and a 33-year-old male both presented with progressive exertional dyspnea and cough. High-resolution computed tomography (HRCT) showed bilateral, diffuse, ill-defined centrilobular nodules of ground-glass opacity, without subpleural thickened septal lines or mediastinal lymphadenopathy. Both cases showed clinical and imaging features characteristic of pulmonary veno-occlusive disease (PVOD) or PCH. The entire EIF2AK4 coding sequence was detected with Sanger sequencing, and no pathogenic EIF2AK4 mutations were identified in either case. Video-assisted thoracoscopic surgery (VATS) was safely performed in both cases, and histopathological examinations of biopsies showed that both patients had PCH. CONCLUSION: Two patients presented with clinical and imaging characteristics suspicious for PVOD/PCH. Despite having no pathogenic EIF2AK4 mutations, both were diagnosed with PCH by VATS lung biopsies. The diagnostic distinction of PCH is important to prompt timely evaluations of patients who may need lung transplantations.

Pulmonary capillary haemangiomatosis: a distinct entity?

Pulmonary capillary haemangiomatosis (PCH) is a rare and incompletely understood histopathological finding characterised by abnormal capillary proliferation within the alveolar interstitium, which has long been noted to share many overlapping features with pulmonary veno-occlusive disease (PVOD). But are PCH and PVOD distinct entities that occur in isolation, or are they closely intertwined manifestations along a spectrum of the same disease? The classic clinical features of both PCH and PVOD include signs and symptoms related to pulmonary hypertension, hypoxaemia, markedly impaired diffusion capacity of the lung and abnormal chest imaging with ground glass opacities, septal lines and lymphadenopathy. In recent years, increasing evidence suggests that the clinical presentation, histopathological features, genetic substrate and pathobiological mechanisms of PCH and PVOD are overlapping and usually indistinguishable. The discovery of biallelic mutations in the eukaryotic translation initiation factor 2 α kinase 4 (EIF2AK4) gene in heritable PCH and PVOD greatly advanced our understanding of the overlapping nature of these conditions. Furthermore, recognition of PCH and PVOD-like changes in other pulmonary vascular diseases and in conditions that cause chronic pulmonary venous hyper-perfusion or hypertension suggests that PCH/PVOD may develop as a reactive process to various insults or injuries to the pulmonary vasculature, rather than being primary angiogenic disorders.

Intravitreal Triamcinolone Acetonide in Von Hippel-Lindau Patient with Novel Mutation.

von Hippel-Lindau (VHL) is a multisystemic inherited disease which most commonly affects the retina and central nervous system. The hallmark retinal manifestation of VHL in the eye is retinal capillary hemangioblastoma (RCH). Significant visual morbidity can result from exudative retinal detachments (ERDs) or tractional retinal detachments. Here, we present a 21-year-old male with long-standing poor vision in the right eye. On examination, he was found to have a massive ERD in the right eye with multiple RCH in both eyes. Genetic testing revealed a heterozygous (c.390dupT) mutation in the VHL gene. Intravitreal triamcinolone acetonide injection resulted in subretinal fluid absorption and near total resolution of ERD. Retinal flattening made RCH accessible for laser photocoagulation. Following multiple focal lasers to the RCH, the lesions were regressed with the flat retina and stable vision.

High frequency of GNA14, GNAQ, and GNA11 mutations in cherry hemangioma: a histopathological and molecular study of 85 cases indicating GNA14 as the most commonly mutated gene in vascular neoplasms.

Cherry hemangioma is the most common hemangioma in adult life. Neoplastic and non-neoplastic theories had both been proposed for its pathogenesis, but its nature is still poorly understood. We noted a significant subset of anastomosing hemangiomas and congenital hemangiomas harbored a population of small capillaries surrounded by a perivascular hyaline layer, reminiscent of the vessels seen in cherry hemangioma. Both anastomosing hemangioma and congenital hemangioma harbor recurrent mutations in exon 5 of GNAQ and its paralogues. In this study, we analyzed 68 cherry hemangiomas and 17 cherry hemangioma-like hemangiomas exhibiting additional non-classical features including markedly dilated, cavernous vessels, and/or a deep component extending to the deep dermis. By Sanger sequencing, GNAQ, GNA11, and GNA14 exon 5 mutations were identified in 12, 4, and 32 cherry hemangiomas, respectively, and 5, 3, and 3 cherry hemangioma-like hemangiomas, respectively. MassARRAY analysis detected mutations (including exon 2 GNAQG48V mutations) in additional 8 cherry hemangiomas and 3 cherry hemangioma-like hemangiomas. Overall, the cherry hemangiomas and cherry hemangioma-like hemangiomas had equal GNA mutation rates (82%), and GNA14 and GNAQ mutations were present in approximately half of cherry hemangiomas and cherry hemangioma-like hemangiomas, respectively. All mutations were mutually exclusive. KRASG12V mutation was also detected in one cherry hemangioma-like hemangioma without GNA mutations. In summary, our study demonstrated recurrent GNA14/GNAQ/GNA11 mutations were present in the majority of this very common hemangioma and established its neoplastic nature. Our results also expanded the morphological spectrum of GNA-mutated hemangiomas to include tumors composed of cavernous-like vessels and indicated GNA14 was the most commonly mutated gene in vascular tumors.

Pulmonary Infantile Hemangioma Mimicking a Congenital Cystic Adenomatoid Malformation.

Infantile hemangioma (IH) is the most common benign vascular tumor of infancy, occurring predominantly in the head and neck. It is characterized by specific endothelial expression of glucose transporter-1 (GLUT-1) and involution with time, spontaneous or on beta-blockers treatment. Although some predisposing factors are known, the exact pathogenesis remains unclear. We report a case of pulmonary IH GLUT-1 positive, initially suspected as a cystic pulmonary airway malformation, in a child presenting with both cardiac and renal malformations. The clinical, radiological, pathological, and genetics findings are discussed with a review of the literature. Although pulmonary IH is a rare lesion, it should be suspected when facing a pulmonary cystic mass in a child.

Widening the landscape of heritable pulmonary hypertension mutations in paediatric and adult cases.

BACKGROUND: Heritable forms of pulmonary arterial hypertension (PAH) and pulmonary veno-occlusive disease/pulmonary capillary haemangiomatosis (PVOD/PCH) diverge by lung histopathological lesions, clinical and para-clinical presentation, their responsible genes, and mode of transmission. Since the identification of the BMPR2 gene in families affected by PAH, mutations in several other genes have been discovered for both forms. The mutation landscape in these new genes is not yet well known. METHODS: We set up a next-generation sequencing-based targeted sequencing gene panel allowing known genes for PAH and PVOD/PCH to be analysed simultaneously. Genetic analysis was prospectively performed on 263 PAH and PVOD/PCH patients (adult and paediatric cases). RESULTS: Pathogenic mutations were identified in 19.5% of sporadic PAH patients (n=180), 54.5% of familial PAH patients and 13.3% of PVOD/PCH patients. BMPR2 was the most frequently mutated gene, followed by TBX4 in both paediatric and adult PAH. BMP9 mutations were identified in 1.2% of adult PAH cases. EIF2AK4 biallelic mutations were restricted to PVOD/PCH. A truncating mutation and a predicted loss-of-function variant were also identified in BMP10 in two severely affected sporadic PAH female patients. CONCLUSION: Our results confirm that mutations are found in genes beyond BMPR2 in heritable PAH, emphasise the role of TBX4 and BMP9, and designate BMP10 as a new PAH gene.

Single-nucleotide polymorphisms of VEGF-A and VEGFR-2 genes and risk of infantile hemangioma.

BACKGROUND: Infantile hemangioma (IH) is the most common vascular tumor of childhood and infancy. It is distinguished by rapid proliferation of endothelial cells during the first year of life followed by spontaneous regression thereafter. One of the possible factors responsible for the IH development is vascular endothelial grow factor (VEGF). The purpose of this study was to evaluate the influence of selected polymorphisms in the genes coding for VEGF-A (+405 G/C, rs2010963; +936 C/T, rs3025039) and its receptor VEGFR-2 (+1416 T/A, rs1870377; -271 G/A, rs7667298) on the susceptibility to infantile hemangioma. METHODS: We performed a case-control study of 99 Polish children hospitalized due to IH and compared them with matched healthy control subjects. The polymorphisms were ascertained through genotyping by PCR-RFLP assay, PCR-HRM, or the allelic discrimination method. RESULTS: The study revealed a lower odds of infantile hemangioma in individuals with GG genotype or G allele for +405 G/C VEGF-A polymorphism (ORdis = 0.52, P = 0.023 and ORdis  = 0.63, P = 0.025, respectively). No association was observed for the remaining VEGF and VEGFR-2 polymorphisms and IH risk. CONCLUSIONS: In our study, none of the investigated VEGF-A and VEGFR-2 genes polymorphisms was found to be an independent prognostic marker for infantile hemangioma. However, there is evidence that individuals carrying at least one G allele of +405 G/C VEGF-A polymorphism have significantly lower risk of IH.

NDRG1 and FOXO1 regulate endothelial cell proliferation in infantile haemangioma.

The etiopathogenesis of infantile haemangioma has not been well understood, and it is accepted that angiogenic mediator dysregulation is the main contributor to the abnormal haemangioma capillary formation. The role of NDRG1, a hypoxia-inducible protein; FOXOs, which are tumor suppressor proteins; and the mTOR complex 2 pathway in infantile haemangioma have not been studied yet. The purpose of this study was to investigate NDRG1 and FOXO1 expression in the infantile haemangioma and the correlation of these proteins with proliferation and involution. Primary endothelial cells were obtained, with parental agreement, from 12 infantile haemangioma patients during surgery; 6 patients had proliferating infantile haemangiomas and 6 had involuting IHs. We compared the infantile haemangioma tissues and primary endothelial cells with human vein endothelial cells using microarrays, real-time PCR, Western blotting and immunohistochemical staining. Our data indicated that FOXO1 expression was downregulated in proliferating infantile haemangioma tissue. We found that the expression of NDRG1, a molecule upstream of the FOXO1 pathway, increased during haemangioma proliferation. NDRG1 knockdown decreased haemangioma endothelial cell proliferation and downregulated c-MYC oncoprotein levels. Our findings suggest that NDRG1 positively regulates haemangioma proliferation. FOXO1 dysregulation plays an important role in infantile haemangiomas pathogenesis.

Curcumin induces apoptosis and inhibits proliferation in infantile hemangioma endothelial cells via downregulation of MCL-1 and HIF-1α.

BACKGROUND: Curcumin has been used as an alternative medicine for the treatment of infantile hemangiomas (IHs); however, the mechanism underlying the effectiveness of curcumin in IHs remains largely unclear. METHODS: In this study, we isolated primary human hemangioma endothelial cells (HemECs) from fresh surgical specimens of 3 patients. We treated HemECs by curcumin and investigated the alterations in proliferative and apoptotic signaling pathways with cell counting kit-8, flow cytometry, western blotting, immunofluorescence, and real-time polymerase chain reaction. RESULTS AND CONCLUSION: We found that curcumin potently inhibited proliferation in HemECs, achieving low-micromolar IC50 (the half maximal inhibitory concentration) value. We also observed that treatment with curcumin induced apoptosis in HemECs, as evidenced by positively Annexin-V-FITC staining, caspase-3 activation, and cleavage of poly(adenosine diphosphate-ribose) polymerase (PARP) in the treated cells. Moreover, we showed that curcumin suppressed the expression of antiapoptotic protein myeloid cell leukemia-1 (MCL-1), hypoxia-inducible factor 1α (HIF-1α), and vascular endothelial growth factor (VEGF).Altogether, our study suggests that the effectiveness of curcumin in IHs may be associated with its potent antiproliferative and apoptotic activities in HemECs.

Genetics of pulmonary hypertension in the clinic.

PURPOSE OF REVIEW: Heritable pulmonary arterial hypertension (PAH) is an autosomal dominant disease with incomplete penetrance because of mutations in bone morphogenetic protein receptor-II (BMPR2), activin A receptor type II-like kinase 1, endoglin, caveolin-1, potassium channel subfamily K, member 3, and T-box gene 4 genes. Heritable pulmonary veno-occlusive disease and/or pulmonary capillary hemangiomatosis (PVOD/PCH) is an autosomal recessive disease because of biallelic mutations in the eukaryotic translation initiation factor 2 alpha kinase 4 gene. The 2015 european society of cardiology (ESC) and european respiratory society (ERS) pulmonary hypertension guidelines recommend genetic counselling and testing to adults and children with PAH or PVOD/PCH as well as in adult relatives at risk of carrying a predisposing mutation. RECENT FINDINGS: In France, genetic counseling and testing are offered to all patients displaying sporadic or familial form of PAH or PVOD/PCH and to their relatives at high risk of carrying a predisposing mutation. Patients with a heritable form of PAH are younger at diagnosis with a worse hemodynamic and a dismal prognosis. Patients with a heritable form of PVOD/PCH are younger at diagnosis with a worse response to specific PAH therapies. A program to detect PAH in an early phase was offered to all asymptomatic BMPR2 mutation carriers, according to the 2015 ESC/ERS guidelines. Finally, preimplantation genetic diagnosis has been performed in families with a history of BMPR2 mutations. SUMMARY: Genetic counseling and testing has to be implemented in pulmonary hypertension centers.

An update of 77 cases diagnosed as oral hemangiomas based on GLUT-1 positivity.

OBJECTIVES: To evaluate cases diagnosed as "oral hemangiomas" based on the immunohistochemical expression of human glucose transporter protein (GLUT-1) and on histopathological features, and to investigate whether the classification proposed by the ISSVA was used correctly to classify these lesions. MATERIAL AND METHODS: All cases stored in the archives of an Oral Pathology Service and diagnosed as "oral hemangiomas" were reviewed. Seventy-seven cases were analyzed regarding the expression of GLUT-1. GLUT-1(+) specimens were classified as true infantile hemangioma (IH) and GLUT-1(-) specimens were reclassified based on their histopathological features. The nomenclature of these lesions was evaluated and some cases were reclassified. RESULTS: Only 26 (33.8%) of the specimens were indeed IHs. Among the GLUT-1(-) specimens, 20 (26.0%) were reclassified as pyogenic granulomas (PGs) and 31 (40.2%) as vascular malformations. Considering the previously applied nomenclature, only 47.5% of the cases initially diagnosed as "hemangiomas" were IHs. In the group of "capillary hemangiomas", most cases (56.2%) were PGs. Among the three "cellular hemangiomas", two were PGs and one was IH. Most (88.8%) "cavernous hemangiomas" were vascular malformations. CONCLUSION: Careful and parameterized review of cases of vascular anomalies is necessary using auxiliary tools such as GLUT-1, since the exclusive use of histopathological findings might be insufficient to differentiate some anomalies. CLINICAL RELEVANCE: Accurate clinical examination and the use of biomarkers such as GLUT-1 are essential for the diagnosis.

Congenital capillary proliferation of the kidney: a distinctive renal vascular lesion of childhood.

Renal vascular lesions (RVL) are rare, and their morphological spectrum remains largely unknown, particularly in children. In this study, we characterize the clinicopathological features of RVL in a cohort of 12 children. Seven lesions were classified as previously recognized entities: vascular malformations (4), papillary endothelial hyperplasia (2), and pyogenic granuloma (lobular capillary hemangioma; 1). An eighth lesion showed nonspecific findings, which were interpreted as reactive during our review. The remaining 4 cases presented either prenatally, at birth, or shortly after birth and were morphologically similar. These were characterized by a peculiar pattern of capillary proliferation with entrapment of native renal structures, variable amounts of extramedullary hematopoiesis and reactive lymphocytes, foci of infarction and hemorrhage, and the presence of feeding and draining vessels at their periphery. To our knowledge, this represents a previously undescribed congenital vascular lesion involving the kidney, which we have descriptively and provisionally termed congenital capillary proliferation of the kidney (CCPK). While it is unclear whether CCPK represents a malformation or neoplastic proliferation, it shows overlapping features with congenital hemangioma of the liver (solitary congenital hepatic hemangioma) and congenital nonprogressive hemangioma (CNH) of the skin and soft tissue, suggesting a possible common pathogenesis among these 3 entities.

Clinical phenotypes and outcomes of heritable and sporadic pulmonary veno-occlusive disease: a population-based study.

BACKGROUND: Bi-allelic mutations of the EIF2AK4 gene cause heritable pulmonary veno-occlusive disease and/or pulmonary capillary haemangiomatosis (PVOD/PCH). We aimed to assess the effect of EIF2AK4 mutations on the clinical phenotypes and outcomes of PVOD/PCH. METHODS: We did a population-based study using clinical, functional, and haemodynamic data from the registry of the French Pulmonary Hypertension Network. We reviewed the clinical data and outcomes from all patients referred to the French Referral Centre (Pulmonary Department, Hospital Kremlin-Bicêtre, University Paris-Sud) with either confirmed or highly probable PVOD/PCH with DNA available for mutation screening (excluding patients with other risk factors of pulmonary hypertension, such as chronic respiratory diseases). We sequenced the coding sequence and intronic junctions of the EIF2AK4 gene, and compared clinical characteristics and outcomes between EIF2AK4 mutation carriers and non-carriers. Medical therapies approved for pulmonary arterial hypertension (prostacyclin derivatives, endothelin receptor antagonists and phosphodiesterase type-5 inhibitors) were given to patients according to the clinical judgment and discretion of treating physicians. The primary outcome was the event-free survival (death or transplantation). Secondary outcomes included response to therapies for pulmonary arterial hypertension and survival after lung transplantation. A satisfactory clinical response to specific therapy for pulmonary arterial hypertension was defined by achieving New York Heart Association functional class I or II, a 6-min walk distance of more than 440 m, and a cardiac index greater than 2·5 L/min per m2 at the first reassessment after initiation of specific therapy for pulmonary arterial hypertension. FINDINGS: We obtained data from Jan 1, 2003, to June 1, 2016, and identified 94 patients with sporadic or heritable PVOD/PCH (confirmed or highly probable). 27 (29%) of these patients had bi-allelic EIF2AK4 mutations. PVOD/PCH due to EIF2AK4 mutations occurred from birth to age 50 years, and these patients were younger at presentation than non-carriers (median 26·0 years [range 0-50.3] vs 60·0 years [6·7-81·4] years; p<0·0001). At diagnosis, both mutations carriers and non-carriers had similarly severe precapillary pulmonary hypertension and functional impairment. 22 (81%) of mutations carriers and 63 (94%) of non-carriers received therapy approved for pulmonary arterial hypertension. Drug-induced pulmonary oedema occurred in five (23%) of treated EIF2AK4 mutations carriers and 13 (21%) of treated non-carriers. Follow-up assessment after initiation of treatment showed that only three (4%) patients with PVOD/PCH reached the predefined criteria for satisfactory clinical response. The probabilities of event-free survival (death or transplantation) at 1 and 3 years were 63% and 32% in EIF2AK4 mutations carriers, and 75% and 34% in non-carriers. No significant differences occurred in event-free survival between the 2 groups (p=0·38). Among the 33 patients who had lung transplantation, estimated post-transplantation survival rates at 1, 2, and 5 years were 84%, 81%, and 73%, respectively. INTERPRETATION: Heritable PVOD/PCH due to bi-allelic EIF2AK4 mutations is characterised by a younger age at diagnosis but these patients display similar disease severity compared with mutation non-carriers. Response to therapy approved for pulmonary arterial hypertension in PVOD/PCH is rare. PVOD/PCH is a devastating condition and lung transplantation should be considered for eligible patients. FUNDING: None.

Double Jeopardy: The Rubber Ball Bounces Twice.

Soblet et al. describe cis mutations in TEK/Tie-2 in blue rubber bleb nevus and sporadic vascular malformations. This suggests that the remaining normal allele is required for the phenotype. Second, it suggests therapeutic approaches to treatment signal transduction inhibition.

Rapamycin inhibits the proliferation of endothelial cells in hemangioma by blocking the mTOR-FABP4 pathway.

FABP4 is widely expressed in both normal and pathologic tissues. It promotes cell proliferation, survival and migration of endothelial cells, and therefore, angiogenesis. However, the role of FABP4 in hemangioma or hemangioma endothelial cells (HemECs) has not been explored. In this study, we investigated whether FABP4 directly regulates the proliferation of HemECs. The expression of cell cycle checkpoint genes was analyzed with the microarray data of human dermal microvascular endothelial cells (HDVECs) and infantile hemangioma endothelial cells. Real-time RT-PCR and western blotting were used to examine the expression of FABP4 in HemECs. Next, the FABP4 expression was inhibited in HemECs using siRNA or rapamycin and upregulated using retroviral transduction of HemECs to assess its influence on proliferation of HemECs. The microarray data showed that cell cycle checkpoint genes were upregulated in HemECs. Moreover, HemECs showed significantly higher proliferation rates than HDVECs. The expression of FABP4 and mTOR was increased in the HemECs. While FABP4 knockdown reduced the BrdU incorporation and cell number of HemECs as expected, cell proliferation was accelerated by FABP4 over-expression. Moreover, rapamycin (10nM) inhibited mTOR-FABP4 signaling and HemEC proliferation. Taken together, these results indicated that mTOR signaling pathway-activated FABP4 directly regulates the proliferation of endothelial cells in hemangioma. Rapamycin and inhibitors of FABP4 have therapeutic potential for treating infantile hemangiomas.

Inheritance Patterns of Infantile Hemangioma.

BACKGROUND AND OBJECTIVE: Infantile hemangioma (IH) includes, among its other risk factors, familial clustering, but a definitive understanding of IH's inheritance model and genetic basis is lacking. Our objective was to collect IH pedigrees in Finland, to study the inheritance patterns of IH within these families, and to analyze the characteristics of familial IHs. METHODS: We identified 185 patients with IH who visited our vascular anomaly clinic between 2004 and 2007. Based on hospital records and a questionnaire sent to these patients and their families, IH characteristics and family history of IH were studied. We compared characteristics between patients with positive (familial) and negative (sporadic) IH family history. Families with positive IH family history were further interviewed for extended pedigree data. RESULTS: One-third of our IH cohort's families reported a family history positive for IH, with IH characteristics and perinatal data between the familial and sporadic cases being similar. IH patients with affected first-degree relatives reported higher long-term discomfort rates than the sporadic cases. Of the 40 families interviewed, 11 included ≥4 IH-affected family members; these were most commonly first-degree relatives (63%). Segregation patterns match with autosomal dominant inheritance with an incomplete penetrance or maternal transmission. We also present a case of monozygotic twins that manifest identical IHs. CONCLUSIONS: Based on this large number of IH pedigrees, we suggest at least 2 possible mechanisms of inheritance: autosomal dominant and maternal transmission. This study highlights the need for additional genetic studies to define inheritance of this common disease.