RESUMO
Genetic predisposition and risk factors such as hypertension and smoking can instigate the development of thoracic aortic aneurysm (TAA), which can lead to highly lethal aortic wall dissection and/or rupture. Monogenic defects in multiple genes involved in the elastin-contractile unit and the TGFß signaling pathway have been associated with TAA in recent years, along with several genetic modifiers and risk-conferring polymorphisms. Advances in omics technology have also provided significant insights into the processes behind aortic wall degeneration: inflammation, epigenetics, vascular smooth muscle phenotype change and depletion, reactive oxygen species generation, mitochondrial dysfunction, and angiotensin signaling dysregulation. These recent advances and findings might pave the way for a therapy that is capable of stopping and perhaps even reversing aneurysm progression.
Assuntos
Aneurisma da Aorta Torácica , Dissecção Aórtica , Dissecção Aórtica/genética , Dissecção Aórtica/metabolismo , Animais , Aneurisma da Aorta Torácica/genética , Aneurisma da Aorta Torácica/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , FenótipoRESUMO
Importin 8, encoded by IPO8, is a ubiquitously expressed member of the importin-ß protein family that translocates cargo molecules such as proteins, RNAs, and ribonucleoprotein complexes into the nucleus in a RanGTP-dependent manner. Current knowledge of the cargoes of importin 8 is limited, but TGF-ß signaling components such as SMAD1-4 have been suggested to be among them. Here, we report that bi-allelic loss-of-function variants in IPO8 cause a syndromic form of thoracic aortic aneurysm (TAA) with clinical overlap with Loeys-Dietz and Shprintzen-Goldberg syndromes. Seven individuals from six unrelated families showed a consistent phenotype with early-onset TAA, motor developmental delay, connective tissue findings, and craniofacial dysmorphic features. A C57BL/6N Ipo8 knockout mouse model recapitulates TAA development from 8-12 weeks onward in both sexes but most prominently shows ascending aorta dilatation with a propensity for dissection in males. Compliance assays suggest augmented passive stiffness of the ascending aorta in male Ipo8-/- mice throughout life. Immunohistological investigation of mutant aortic walls reveals elastic fiber disorganization and fragmentation along with a signature of increased TGF-ß signaling, as evidenced by nuclear pSmad2 accumulation. RT-qPCR assays of the aortic wall in male Ipo8-/- mice demonstrate decreased Smad6/7 and increased Mmp2 and Ccn2 (Ctgf) expression, reinforcing a role for dysregulation of the TGF-ß signaling pathway in TAA development. Because importin 8 is the most downstream TGF-ß-related effector implicated in TAA pathogenesis so far, it offers opportunities for future mechanistic studies and represents a candidate drug target for TAA.
Assuntos
Aneurisma da Aorta Torácica/etiologia , Mutação com Perda de Função , Perda de Heterozigosidade , Fenótipo , beta Carioferinas/genética , Adulto , Animais , Aneurisma da Aorta Torácica/metabolismo , Aneurisma da Aorta Torácica/patologia , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Linhagem , Transdução de Sinais , Síndrome , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Adulto Jovem , beta Carioferinas/metabolismoRESUMO
Cardiovascular outcome in Marfan syndrome (MFS) patients most prominently depends on aortic aneurysm progression with subsequent aortic dissection. Angiotensin II receptor blockers (ARBs) prevent aneurysm formation in MFS mouse models. In patients, ARBs only slow down aortic dilation. Downstream signalling from the angiotensin II type 1 receptor (AT1R) is mediated by G proteins and ß-arrestin recruitment. AT1R also interacts with the monocyte chemoattractant protein-1 (MCP-1) receptor, resulting in inflammation. In this study, we explore the targeting of ß-arrestin signalling in MFS mice by administering TRV027. Furthermore, because high doses of the ARB losartan, which has been proven beneficial in MFS, cannot be achieved in humans, we investigate a potential additive effect by combining lower concentrations of losartan (25 mg/kg/day and 5 mg/kg/day) with barbadin, a ß-arrestin blocker, and DMX20, a C-C chemokine receptor type 2 (CCR2) blocker. A high dose of losartan (50 mg/kg/day) slowed down aneurysm progression compared to untreated MFS mice (1.73 ± 0.12 vs. 1.96 ± 0.08 mm, p = 0.0033). TRV027, the combination of barbadin with losartan (25 mg/kg/day), and DMX-200 (90 mg/kg/day) with a low dose of losartan (5 mg/kg/day) did not show a significant beneficial effect. Our results confirm that while losartan effectively halts aneurysm formation in Fbn1C1041G/+ MFS mice, neither TRV027 alone nor any of the other compounds combined with lower doses of losartan demonstrate a notable impact on aneurysm advancement. It appears that complete blockade of AT1R function, achieved by administrating a high dosage of losartan, may be necessary for inhibiting aneurysm progression in MFS.
Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II , Aneurisma Aórtico , Losartan , Síndrome de Marfan , Transdução de Sinais , Camundongos , Síndrome de Marfan/tratamento farmacológico , Síndrome de Marfan/patologia , Modelos Animais de Doenças , Aneurisma Aórtico/tratamento farmacológico , Aneurisma Aórtico/prevenção & controle , Oligopeptídeos/administração & dosagem , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/patologia , Pirimidinas/administração & dosagem , Combinação de Medicamentos , Losartan/administração & dosagem , Antagonistas de Receptores de Angiotensina/administração & dosagem , Transdução de Sinais/efeitos dos fármacos , Humanos , Bloqueadores do Receptor Tipo 1 de Angiotensina II/administração & dosagemRESUMO
PRKACA and PRKACB code for two catalytic subunits (Cα and Cß) of cAMP-dependent protein kinase (PKA), a pleiotropic holoenzyme that regulates numerous fundamental biological processes such as metabolism, development, memory, and immune response. We report seven unrelated individuals presenting with a multiple congenital malformation syndrome in whom we identified heterozygous germline or mosaic missense variants in PRKACA or PRKACB. Three affected individuals were found with the same PRKACA variant, and the other four had different PRKACB mutations. In most cases, the mutations arose de novo, and two individuals had offspring with the same condition. Nearly all affected individuals and their affected offspring shared an atrioventricular septal defect or a common atrium along with postaxial polydactyly. Additional features included skeletal abnormalities and ectodermal defects of variable severity in five individuals, cognitive deficit in two individuals, and various unusual tumors in one individual. We investigated the structural and functional consequences of the variants identified in PRKACA and PRKACB through the use of several computational and experimental approaches, and we found that they lead to PKA holoenzymes which are more sensitive to activation by cAMP than are the wild-type proteins. Furthermore, expression of PRKACA or PRKACB variants detected in the affected individuals inhibited hedgehog signaling in NIH 3T3 fibroblasts, thereby providing an underlying mechanism for the developmental defects observed in these cases. Our findings highlight the importance of both Cα and Cß subunits of PKA during human development.
Assuntos
Anormalidades Múltiplas/genética , Disfunção Cognitiva/genética , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico/genética , Dedos/anormalidades , Mutação em Linhagem Germinativa , Defeitos dos Septos Cardíacos/genética , Polidactilia/genética , Dedos do Pé/anormalidades , Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/patologia , Adolescente , Adulto , Animais , Sequência de Bases , Disfunção Cognitiva/diagnóstico , Disfunção Cognitiva/patologia , AMP Cíclico/metabolismo , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico/química , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico/deficiência , Feminino , Dedos/patologia , Regulação da Expressão Gênica no Desenvolvimento , Defeitos dos Septos Cardíacos/diagnóstico , Defeitos dos Septos Cardíacos/patologia , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Holoenzimas/química , Holoenzimas/deficiência , Holoenzimas/genética , Humanos , Recém-Nascido , Masculino , Camundongos , Modelos Moleculares , Mosaicismo , Células NIH 3T3 , Linhagem , Polidactilia/diagnóstico , Polidactilia/patologia , Estrutura Secundária de Proteína , Dedos do Pé/patologiaRESUMO
Different pathogenic variants in the fibrillin-1 gene (FBN1) cause Marfan syndrome and acromelic dysplasias. Whereas the musculoskeletal features of Marfan syndrome involve tall stature, arachnodactyly, joint hypermobility, and muscle hypoplasia, acromelic dysplasia patients present with short stature, brachydactyly, stiff joints, and hypermuscularity. Similarly, pathogenic variants in the fibrillin-2 gene (FBN2) cause either a Marfanoid congenital contractural arachnodactyly or a FBN2-related acromelic dysplasia that most prominently presents with brachydactyly. The phenotypic and molecular resemblances between both the FBN1 and FBN2-related disorders suggest that reciprocal pathomechanistic lessons can be learned. In this review, we provide an updated overview and comparison of the phenotypic and mutational spectra of both the "tall" and "short" fibrillinopathies. The future parallel functional study of both FBN1/2-related disorders will reveal new insights into how pathogenic fibrillin variants differently affect the fibrillin microfibril network and/or growth factor homeostasis in clinically opposite syndromes. This knowledge may eventually be translated into new therapeutic approaches by targeting or modulating the fibrillin microfibril network and/or the signaling pathways under its control.
Assuntos
Fibrilina-1 , Fibrilina-2 , Síndrome de Marfan , Anormalidades Musculoesqueléticas/genética , Braquidactilia , Fibrilina-1/genética , Fibrilina-2/genética , Humanos , Síndrome de Marfan/genética , FenótipoRESUMO
PURPOSE: In a large cohort of 373 pediatric patients with Marfan syndrome (MFS) with a severe cardiovascular phenotype, we explored the proportion of patients with MFS with a pathogenic FBN1 variant and analyzed whether the type/location of FBN1 variants was associated with specific clinical characteristics and response to treatment. Patients were recruited on the basis of the following criteria: aortic root z-score > 3, age 6 months to 25 years, no prior or planned surgery, and aortic root diameter < 5 cm. METHODS: Targeted resequencing and deletion/duplication testing of FBN1 and related genes were performed. RESULTS: We identified (likely) pathogenic FBN1 variants in 91% of patients. Ectopia lentis was more frequent in patients with dominant-negative (DN) variants (61%) than in those with haploinsufficient variants (27%). For DN FBN1 variants, the prevalence of ectopia lentis was highest in the N-terminal region (84%) and lowest in the C-terminal region (17%). The association with a more severe cardiovascular phenotype was not restricted to DN variants in the neonatal FBN1 region (exon 25-33) but was also seen in the variants in exons 26 to 49. No difference in the therapeutic response was detected between genotypes. CONCLUSION: Important novel genotype-phenotype associations involving both cardiovascular and extra-cardiovascular manifestations were identified, and existing ones were confirmed. These findings have implications for prognostic counseling of families with MFS.
Assuntos
Ectopia do Cristalino , Síndrome de Marfan , Variação Biológica da População , Criança , Ectopia do Cristalino/complicações , Ectopia do Cristalino/genética , Fibrilina-1/genética , Fibrilinas/genética , Genótipo , Humanos , Síndrome de Marfan/genética , Mutação , FenótipoRESUMO
BACKGROUND: Although carpal tunnel syndrome (CTS) is the most common form of peripheral entrapment neuropathy, its pathogenesis remains largely unknown. An estimated heritability index of 0.46 and an increased familial occurrence indicate that genetic factors must play a role in the pathogenesis. METHODS AND RESULTS: We report on a family in which CTS occurred in subsequent generations at an unusually young age. Additional clinical features included brachydactyly and short Achilles tendons resulting in toe walking in childhood. Using exome sequencing, we identified a heterozygous variant (c.5009T>G; p.Phe1670Cys) in the fibrillin-2 (FBN2) gene that co-segregated with the phenotype in the family. Functional assays showed that the missense variant impaired integrin-mediated cell adhesion and migration. Moreover, we observed an increased transforming growth factor-ß signalling and fibrosis in the carpal tissues of affected individuals. A variant burden test in a large cohort of patients with CTS revealed a significantly increased frequency of rare (6.7% vs 2.5%-3.4%, p<0.001) and high-impact (6.9% vs 2.7%, p<0.001) FBN2 variants in patient alleles compared with controls. CONCLUSION: The identification of a novel FBN2 variant (p.Phe1670Cys) in a unique family with early onset CTS, together with the observed increased frequency of rare and high-impact FBN2 variants in patients with sporadic CTS, strongly suggest a role of FBN2 in the pathogenesis of CTS.
Assuntos
Síndrome do Túnel Carpal/genética , Fibrilina-2/genética , Tendão do Calcâneo/anormalidades , Estatura/genética , Síndrome do Túnel Carpal/diagnóstico por imagem , Síndrome do Túnel Carpal/etiologia , Humanos , Masculino , Mutação de Sentido Incorreto , LinhagemRESUMO
Marfan syndrome is a rare connective tissue disorder that causes aortic dissection-related sudden death. Current conventional treatments, beta-blockers, and type 1 angiotensin II receptor blockers are prescribed to slow down aortic aneurysm progression and delay (prophylactic) aortic surgery. However, neither of these treatments ceases aortic growth completely. This review focuses on potential alternative therapeutic leads in the field, ranging from widely used medication with beneficial effects on the aorta to experimental inhibitors with the potential to stop aortic growth in Marfan syndrome. Clinical trials are warranted to uncover their full potential.
Assuntos
Síndrome de Marfan , Síndrome de Marfan/tratamento farmacológico , Humanos , Animais , Doenças da Aorta/tratamento farmacológico , Doenças da Aorta/etiologia , Aneurisma AórticoRESUMO
Marfan syndrome (MFS) is a connective tissue disorder with pleiotropic manifestations in the ocular, skeletal and cardiovascular system. Ruptured aortic aneurysms in MFS patients are associated with high mortality rates. MFS is typically caused by pathogenic variants in the fibrillin-1 (FBN1) gene. Here, we report a generated induced pluripotent cell (iPSC) line of a MFS patient with a FBN1 c.5372G > A (p.Cys1791Tyr) variant. For that, skin fibroblasts of a MFS patient carrying a FBN1 c.5372G > A (p.Cys1791Tyr) variant were successfully reprogrammed into iPSCs using the CytoTune™-iPS 2.0 Sendai Kit (Invitrogen). The iPSCs showed a normal karyotype, expressed pluripotency markers, were able to differentiate into three germ layers and carried the original genotype.
Assuntos
Células-Tronco Pluripotentes Induzidas , Síndrome de Marfan , Humanos , Síndrome de Marfan/genética , Síndrome de Marfan/patologia , Fibrilina-1/genética , Células-Tronco Pluripotentes Induzidas/metabolismo , Mutação , GenótipoRESUMO
Marfan syndrome (MFS) is a connective tissue disorder with pleiotropic manifestations in the ocular, skeletal and cardiovascular system; and is typically cause by pathogenic variants in the fibrillin-1 (FBN1) gene. We report a generated induced pluripotent cell (iPSC) line of a MFS patient with an FBN1 c.7754T > C (p.Ile2585Thr) variant. The cell line was generated from peripheral blood mononuclear cells (PBMCs) and after reprogramming the line showed a no relevant copy number alterations, expression of pluripotency markers and was able to differentiate into three germ layers while carrying the original genotype.
Assuntos
Células-Tronco Pluripotentes Induzidas , Síndrome de Marfan , Humanos , Síndrome de Marfan/genética , Fibrilina-1/genética , Células-Tronco Pluripotentes Induzidas/metabolismo , Leucócitos Mononucleares/metabolismo , MutaçãoRESUMO
Spondyloepiphyseal dysplasia congenita (SEDC) is a severe non-lethal type 2 collagenopathy caused by pathogenic variants in the COL2A1 gene, which encodes the alpha-1 chain of type II collagen. SEDC is clinically characterized by severe short stature, degenerative joint disease, hearing impairment, orofacial anomalies and ocular manifestations. To study and therapeutically target the underlying disease mechanisms, human iPSC-chondrocytes are considered highly suitable as they have been shown to exhibit several key features of skeletal dysplasias. Prior to creating iPSC-chondrocytes, peripheral blood mononuclear cells of two male SEDC patients, carrying the p.Gly1107Arg and p.Gly408Asp pathogenic variants, respectively, were successfully reprogrammed into iPSCs using the CytoTune™-iPS 2.0 Sendai Kit (Invitrogen).
Assuntos
Células-Tronco Pluripotentes Induzidas , Osteocondrodisplasias , Humanos , Masculino , Leucócitos Mononucleares , Osteocondrodisplasias/genética , Colágeno Tipo II/genéticaRESUMO
Hemizygous missense variants in the X-linked BGN gene, encoding the extracellular matrix protein biglycan, cause spondyloepimetaphyseal dysplasia (SEMD, biglycan type), which is clinically characterized by short stature, brachydactyly and osteoarthritis. Little is known about the pathomechanisms underlying SEMD, biglycan type. IPSC-derived chondrocyte disease models have been shown to exhibit several key aspects of known disease mechanisms of skeletal dysplasias and are therefore considered highly suitable human disease models to study SEMD, biglycan type. Prior to creating iPSC-chondrocytes, dermal fibroblasts of two male patients with SEMD, biglycan type, carrying the p.Gly259Val variant were successfully reprogrammed into iPSCs using the CytoTuneTM-iPS 2.0 Sendai Kit (Invitrogen).
Assuntos
Células-Tronco Pluripotentes Induzidas , Osteocondrodisplasias , Humanos , Masculino , Biglicano/genética , Osteocondrodisplasias/genética , Mutação de Sentido Incorreto , Proteínas da Matriz Extracelular/genéticaRESUMO
Induced pluripotent stem cell (iPSC) technology allows pathomechanistic and therapeutic investigation of human heritable disorders affecting tissue types whose collection from patients is difficult or even impossible. Among them are cartilage diseases. Over the past decade, iPSC-chondrocyte disease models have been shown to exhibit several key aspects of known disease mechanisms. Concurrently, an increasing number of protocols to differentiate iPSCs into chondrocytes have been published, each with its respective (dis)advantages. In this review we provide a comprehensive overview of the different differentiation approaches, the hitherto described iPSC-chondrocyte disease models and mechanistic and/or therapeutic insights that have been derived from their investigation, and the current model limitations. Key lessons are that the most appropriate differentiation approach is dependent upon the cartilage disease under investigation and that further optimization is still required to recapitulate the in vivo cartilage. © 2022 American Society for Bone and Mineral Research (ASBMR).
Assuntos
Doenças das Cartilagens , Células-Tronco Pluripotentes Induzidas , Cartilagem/metabolismo , Doenças das Cartilagens/metabolismo , Diferenciação Celular , Condrócitos/metabolismo , Descoberta de Drogas , HumanosRESUMO
Although at first glance chondrodysplasia and aneurysmal thoracic aortopathy seem oddly dissimilar, recent lines of evidences indicate that they share molecular similarities. Chondrodysplasias are a group of skeletal disorders characterized by genetic defects in hyaline cartilage. Aneurysmal thoracic aortopathy is the pathological enlargement of the thoracic aorta due to wall weakness, along with its ensuing life-threatening complications (i.e., aortic dissection and/or rupture). Extracellular matrix dysregulation, abnormal TGF-ß signaling, and, to a more limited extent, endoplasmic reticulum stress emerge as common disease processes. In this review we provide a comprehensive overview of the genetic and pathomechanistic overlap as well as of how these commonalities can guide treatment strategies for both disease entities.
Assuntos
Aneurisma da Aorta Torácica/genética , Dissecção Aórtica/genética , Exostose Múltipla Hereditária/genética , Animais , Retículo Endoplasmático/genética , Estresse do Retículo Endoplasmático/genética , Matriz Extracelular/genética , Humanos , Transdução de Sinais/genéticaRESUMO
PURPOSE: In a significant proportion of children born small for gestational age (SGA) with failure of catch-up growth, the etiology of short stature remains unclear after routine diagnostic workup. We wanted to investigate if extensive analysis of the (epi)genome can unravel the cause of growth failure in a significant portion of these children. PATIENTS AND METHODS: Twenty SGA children treated with GH because of short stature were selected from the BELGROW database of the Belgian Society for Pediatric Endocrinology and Diabetology for exome sequencing, single-nucleotide polymorphism (SNP) array and genome-wide methylation analysis to identify the (epi)genetic cause. First-year response to GH was compared with the response of SGA patients in the KIGS database. RESULTS: We identified (likely) pathogenic variants in 4 children (from 3 families) using exome sequencing and found pathogenic copy number variants in 2 probands using SNP array. In a child harboring a NSD1-containing microduplication, we identified a DNA methylation signature that is opposite to the genome-wide DNA methylation signature of Sotos syndrome. Moreover, we observed multilocus imprinting disturbances in 2 children in whom no other genomic alteration could be identified. Five of 6 children with a genetic diagnosis had an "above average" response to GH. CONCLUSIONS: The study indicates that a more advanced approach with deep genotyping can unravel unexpected (epi)genomic alterations in SGA children with persistent growth failure. Most SGA children with a genetic diagnosis had a good response to GH treatment.
Assuntos
Metilação de DNA/genética , Doenças do Recém-Nascido/genética , Recém-Nascido Pequeno para a Idade Gestacional/fisiologia , Epigenoma , Feminino , Genômica , Humanos , Recém-Nascido , MasculinoRESUMO
Sclerosteosis is a rare autosomal recessive bone disorder marked by hyperostosis of the skull and tubular bones. Initially, we and others reported that sclerosteosis was caused by loss-of-function mutations in SOST, encoding sclerostin. More recently, we identified disease-causing mutations in LRP4, a binding partner of sclerostin, in three sclerosteosis patients. Upon binding to sclerostin, LRP4 can inhibit the canonical WNT signaling that is known to be an important pathway in the regulation of bone formation. To further investigate the role of LRP4 in the bone formation process, we generated an Lrp4 mutated sclerosteosis mouse model by introducing the p.Arg1170Gln mutation in the mouse genome. Extensive analysis of the bone phenotype of the Lrp4R1170Q/R1170Q knock-in (KI) mouse showed the presence of increased trabecular and cortical bone mass as a consequence of increased bone formation by the osteoblasts. In addition, three-point bending analysis also showed that the increased bone mass results in increased bone strength. In contrast to the human sclerosteosis phenotype, we could not observe syndactyly in the forelimbs or hindlimbs of the Lrp4 KI animals. Finally, we could not detect any significant changes in the bone formation and resorption markers in the serum of the mutant mice. However, the serum sclerostin levels were strongly increased and the level of sclerostin in the tibia was decreased in Lrp4R1170Q/R1170Q mice, confirming the role of LRP4 as an anchor for sclerostin in bone. In conclusion, the Lrp4R1170Q/R1170Q mouse is a good model for the human sclerosteosis phenotype caused by mutations in LRP4 and can be used in the future for further investigation of the mechanism whereby LRP4 regulates bone formation. © 2017 American Society for Bone and Mineral Research.