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1.
Artículo en Inglés | MEDLINE | ID: mdl-34533615

RESUMEN

In the last years, the field of inheritable ventricular arrhythmia disease modelling has changed significantly with a push towards the use of novel cellular cardiomyocyte based models. However, there is a growing need for new in vivo models to study the disease pathology at the tissue and organ level. Zebrafish provide an excellent opportunity for in vivo modelling of inheritable ventricular arrhythmia syndromes due to the remarkable similarity between their cardiac electrophysiology and that of humans. Additionally, many state-of-the-art methods in gene editing and electrophysiological phenotyping are available for zebrafish research. In this review, we give a comprehensive overview of the published zebrafish genetic models for primary electrical disorders and arrhythmogenic cardiomyopathy. We summarise and discuss the strengths and weaknesses of the different technical approaches for the generation of genetically modified zebrafish disease models, as well as the electrophysiological approaches in zebrafish phenotyping. By providing this detailed overview, we aim to draw attention to the potential of the zebrafish model for studying arrhythmia syndromes at the organ level and as a platform for personalised medicine and drug testing.


Asunto(s)
Modelos Genéticos , Pez Cebra , Humanos , Animales , Pez Cebra/genética , Síndrome , Arritmias Cardíacas/genética , Miocitos Cardíacos
2.
Am J Hum Genet ; 108(6): 1115-1125, 2021 06 03.
Artículo en Inglés | MEDLINE | ID: mdl-34010605

RESUMEN

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.


Asunto(s)
Aneurisma de la Aorta Torácica/etiología , Mutación con Pérdida de Función , Pérdida de Heterocigocidad , Fenotipo , beta Carioferinas/genética , Adulto , Animales , Aneurisma de la Aorta Torácica/metabolismo , Aneurisma de la Aorta Torácica/patología , Niño , Preescolar , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Linaje , Transducción de Señal , Síndrome , Factor de Crecimiento Transformador beta/genética , Factor de Crecimiento Transformador beta/metabolismo , Adulto Joven , beta Carioferinas/metabolismo
3.
Europace ; 23(6): 918-927, 2021 06 07.
Artículo en Inglés | MEDLINE | ID: mdl-33221854

RESUMEN

AIMS: We identified the first Belgian SCN5A founder mutation, c.4813 + 3_4813 + 6dupGGGT. To describe the clinical spectrum and disease severity associated with this mutation, clinical data of 101 SCN5A founder mutation carriers and 46 non-mutation carrying family members from 25 Belgian families were collected. METHODS AND RESULTS: The SCN5A founder mutation was confirmed by haplotype analysis. The clinical history and electrocardiographic parameters of the mutation carriers and their family members were gathered and compared. A cardiac electrical abnormality was observed in the majority (82%) of the mutation carriers. Cardiac conduction defects, defined as PR or QRS prolongation on electrocardiogram (ECG), were most frequent, occurring in 65% of the mutation carriers. Brugada syndrome (BrS) was the second most prevalent phenotype identified in 52%, followed by atrial dysrythmia in 11%. Overall, 33% of tested mutation carriers had a normal sodium channel blocker test. Negative tests were more common in family members distantly related to the proband. Overall, 23% of the mutation carriers were symptomatic, with 8% displaying major adverse events. As many as 13% of the patients tested with a sodium blocker developed ventricular arrhythmia. One family member who did not carry the founder mutation was diagnosed with BrS. CONCLUSION: The high prevalence of symptoms and sensitivity to sodium channel blockers in our founder population highlights the adverse effect of the founder mutation on cardiac conduction. The large phenotypical heterogeneity, variable penetrance, and even non-segregation suggest that other genetic (and environmental) factors modify the disease expression, severity, and outcome in these families.


Asunto(s)
Síndrome de Brugada , Canal de Sodio Activado por Voltaje NAV1.5 , Bélgica/epidemiología , Electrocardiografía , Humanos , Mutación , Canal de Sodio Activado por Voltaje NAV1.5/genética , Fenotipo
4.
Int J Mol Sci ; 22(6)2021 Mar 10.
Artículo en Inglés | MEDLINE | ID: mdl-33802229

RESUMEN

Brugada syndrome (BrS) is an inherited cardiac arrhythmia that predisposes to ventricular fibrillation and sudden cardiac death. It originates from oligogenic alterations that affect cardiac ion channels or their accessory proteins. The main hurdle for the study of the functional effects of those variants is the need for a specific model that mimics the complex environment of human cardiomyocytes. Traditionally, animal models or transient heterologous expression systems are applied for electrophysiological investigations, each of these models having their limitations. The ability to create induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), providing a source of human patient-specific cells, offers new opportunities in the field of cardiac disease modelling. Contemporary iPSC-CMs constitute the best possible in vitro model to study complex cardiac arrhythmia syndromes such as BrS. To date, thirteen reports on iPSC-CM models for BrS have been published and with this review we provide an overview of the current findings, with a focus on the electrophysiological parameters. We also discuss the methods that are used for cell derivation and data acquisition. In the end, we critically evaluate the knowledge gained by the use of these iPSC-CM models and discuss challenges and future perspectives for iPSC-CMs in the study of BrS and other arrhythmias.


Asunto(s)
Síndrome de Brugada/metabolismo , Células Madre Pluripotentes Inducidas/metabolismo , Modelos Cardiovasculares , Miocitos Cardíacos/metabolismo , Síndrome de Brugada/patología , Humanos , Células Madre Pluripotentes Inducidas/patología , Miocitos Cardíacos/patología
5.
J Med Genet ; 56(4): 220-227, 2019 04.
Artículo en Inglés | MEDLINE | ID: mdl-29967133

RESUMEN

BACKGROUND: Missense variants in SMAD2, encoding a key transcriptional regulator of transforming growth factor beta signalling, were recently reported to cause arterial aneurysmal disease. OBJECTIVES: The aims of the study were to identify the genetic disease cause in families with aortic/arterial aneurysmal disease and to further define SMAD2 genotype-phenotype correlations. METHODS AND RESULTS: Using gene panel sequencing, we identified a SMAD2 nonsense variant and four SMAD2 missense variants, all affecting highly conserved amino acids in the MH2 domain. The premature stop codon (c.612dup; p.(Asn205*)) was identified in a marfanoid patient with aortic root dilatation and in his affected father. A p.(Asn318Lys) missense variant was found in a Marfan syndrome (MFS)-like case who presented with aortic root aneurysm and in her affected daughter with marfanoid features and mild aortic dilatation. In a man clinically diagnosed with Loeys-Dietz syndrome (LDS) that presents with aortic root dilatation and marked tortuosity of the neck vessels, another missense variant, p.(Ser397Tyr), was identified. This variant was also found in his affected daughter with hypertelorism and arterial tortuosity, as well as his affected mother. The third missense variant, p.(Asn361Thr), was discovered in a man presenting with coronary artery dissection. Variant genotyping in three unaffected family members confirmed its absence. The last missense variant, p.(Ser467Leu), was identified in a man with significant cardiovascular and connective tissue involvement. CONCLUSION: Taken together, our data suggest that heterozygous loss-of-function SMAD2 variants can cause a wide spectrum of autosomal dominant aortic and arterial aneurysmal disease, combined with connective tissue findings reminiscent of MFS and LDS.


Asunto(s)
Aneurisma/etiología , Disección Aórtica/etiología , Disección Aórtica/patología , Arterias/patología , Variación Genética , Proteína Smad2/genética , Adulto , Anciano , Alelos , Sustitución de Aminoácidos , Aneurisma/patología , Niño , Facies , Femenino , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Masculino , Síndrome de Marfan/complicaciones , Síndrome de Marfan/genética , Persona de Mediana Edad , Mutación , Linaje , Fenotipo , Proteína Smad2/metabolismo
6.
Am J Hum Genet ; 99(1): 174-87, 2016 Jul 07.
Artículo en Inglés | MEDLINE | ID: mdl-27392076

RESUMEN

Autosomal-dominant tubulo-interstitial kidney disease (ADTKD) encompasses a group of disorders characterized by renal tubular and interstitial abnormalities, leading to slow progressive loss of kidney function requiring dialysis and kidney transplantation. Mutations in UMOD, MUC1, and REN are responsible for many, but not all, cases of ADTKD. We report on two families with ADTKD and congenital anemia accompanied by either intrauterine growth retardation or neutropenia. Ultrasound and kidney biopsy revealed small dysplastic kidneys with cysts and tubular atrophy with secondary glomerular sclerosis, respectively. Exclusion of known ADTKD genes coupled with linkage analysis, whole-exome sequencing, and targeted re-sequencing identified heterozygous missense variants in SEC61A1-c.553A>G (p.Thr185Ala) and c.200T>G (p.Val67Gly)-both affecting functionally important and conserved residues in SEC61. Both transiently expressed SEC6A1A variants are delocalized to the Golgi, a finding confirmed in a renal biopsy from an affected individual. Suppression or CRISPR-mediated deletions of sec61al2 in zebrafish embryos induced convolution defects of the pronephric tubules but not the pronephric ducts, consistent with the tubular atrophy observed in the affected individuals. Human mRNA encoding either of the two pathogenic alleles failed to rescue this phenotype as opposed to a complete rescue by human wild-type mRNA. Taken together, these findings provide a mechanism by which mutations in SEC61A1 lead to an autosomal-dominant syndromic form of progressive chronic kidney disease. We highlight protein translocation defects across the endoplasmic reticulum membrane, the principal role of the SEC61 complex, as a contributory pathogenic mechanism for ADTKD.


Asunto(s)
Anemia/genética , Heterocigoto , Enfermedades Renales/genética , Mutación , Canales de Translocación SEC/genética , Adulto , Anciano , Alelos , Secuencia de Aminoácidos , Animales , Biopsia , Niño , Enfermedad Crónica , Progresión de la Enfermedad , Retículo Endoplásmico/metabolismo , Exoma/genética , Femenino , Retardo del Crecimiento Fetal/genética , Genes Dominantes , Aparato de Golgi/metabolismo , Humanos , Recién Nacido , Enfermedades Renales/patología , Masculino , Persona de Mediana Edad , Modelos Moleculares , Mutación Missense/genética , Neutropenia/genética , Linaje , Fenotipo , ARN Mensajero/análisis , ARN Mensajero/genética , Canales de Translocación SEC/química , Síndrome , Adulto Joven , Pez Cebra/embriología , Pez Cebra/genética
7.
Hum Mutat ; 39(5): 621-634, 2018 05.
Artículo en Inglés | MEDLINE | ID: mdl-29392890

RESUMEN

The Loeys-Dietz syndrome (LDS) is a connective tissue disorder affecting the cardiovascular, skeletal, and ocular system. Most typically, LDS patients present with aortic aneurysms and arterial tortuosity, hypertelorism, and bifid/broad uvula or cleft palate. Initially, mutations in transforming growth factor-ß (TGF-ß) receptors (TGFBR1 and TGFBR2) were described to cause LDS, hereby leading to impaired TGF-ß signaling. More recently, TGF-ß ligands, TGFB2 and TGFB3, as well as intracellular downstream effectors of the TGF-ß pathway, SMAD2 and SMAD3, were shown to be involved in LDS. This emphasizes the role of disturbed TGF-ß signaling in LDS pathogenesis. Since most literature so far has focused on TGFBR1/2, we provide a comprehensive review on the known and some novel TGFB2/3 and SMAD2/3 mutations. For TGFB2 and SMAD3, the clinical manifestations, both of the patients previously described in the literature and our newly reported patients, are summarized in detail. This clearly indicates that LDS concerns a disorder with a broad phenotypical spectrum that is still emerging as more patients will be identified. All mutations described here are present in the corresponding Leiden Open Variant Database.


Asunto(s)
Estudios de Asociación Genética , Síndrome de Loeys-Dietz/genética , Mutación/genética , Proteína Smad2/genética , Proteína smad3/genética , Factor de Crecimiento Transformador beta2/genética , Factor de Crecimiento Transformador beta3/genética , Animales , Modelos Animales de Enfermedad , Humanos , Síndrome de Loeys-Dietz/diagnóstico , Ratones , Transducción de Señal/genética
8.
BMC Med Genet ; 19(1): 140, 2018 08 08.
Artículo en Inglés | MEDLINE | ID: mdl-30089473

RESUMEN

BACKGROUND: Mutations in the X-linked gene filamin A (FLNA), encoding the actin-binding protein FLNA, cause a wide spectrum of connective tissue, skeletal, cardiovascular and/or gastrointestinal manifestations. Males are typically more severely affected than females with common pre- or perinatal death. CASE PRESENTATION: We provide a genotype- and phenotype-oriented literature overview of FLNA hemizygous mutations and report on two live-born male FLNA mutation carriers. Firstly, we identified a de novo, missense mutation (c.238C > G, p.(Leu80Val)) in a five-year old Indian boy who presented with periventricular nodular heterotopia, increased skin laxity, joint hypermobility, mitral valve prolapse with regurgitation and marked facial features (e.g. a flat face, orbital fullness, upslanting palpebral fissures and low-set ears). Secondly, we identified two cis-located FLNA mutations (c.7921C > G, p.(Pro2641Ala); c.7923delC, p.(Tyr2642Thrfs*63)) in a Bosnian patient with Ehlers-Danlos syndrome-like features such as skin translucency and joint hypermobility. This patient also presented with brain anomalies, pectus excavatum, mitral valve prolapse, pulmonary hypertension and dilatation of the pulmonary arteries. He died from heart failure in his second year of life. CONCLUSIONS: These two new cases expand the list of live-born FLNA mutation-positive males with connective tissue disease from eight to ten, contributing to a better knowledge of the genetic and phenotypic spectrum of FLNA-related disease.


Asunto(s)
Enfermedades del Tejido Conjuntivo/genética , Filaminas/metabolismo , Mutación/genética , Adolescente , Adulto , Niño , Preescolar , Tejido Conectivo/metabolismo , Síndrome de Ehlers-Danlos/genética , Genes Ligados a X/genética , Genotipo , Humanos , Recién Nacido , Masculino , Persona de Mediana Edad , Linaje , Fenotipo , Adulto Joven
9.
Ann Neurol ; 75(3): 382-94, 2014 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-24318194

RESUMEN

OBJECTIVE: Mutations in KCNQ2 and KCNQ3, encoding the voltage-gated potassium channels KV 7.2 and KV 7.3, are known to cause benign familial neonatal seizures mainly by haploinsufficiency. Here, we set out to determine the disease mechanism of 7 de novo missense KCNQ2 mutations that were recently described in patients with a severe epileptic encephalopathy including pharmacoresistant seizures and pronounced intellectual disability. METHODS: Mutations were inserted into the KCNQ2 cDNA. Potassium currents were recorded using 2-microelectrode voltage clamping, and surface expression was analyzed by a biotinylation assay in cRNA-injected Xenopus laevis oocytes. RESULTS: We observed a clear loss of function for all mutations. Strikingly, 5 of 7 mutations exhibited a drastic dominant-negative effect on wild-type KV 7.2 or KV 7.3 subunits, either by globally reducing current amplitudes (3 pore mutations) or by a depolarizing shift of the activation curve (2 voltage sensor mutations) decreasing potassium currents at the subthreshold level at which these channels are known to critically influence neuronal firing. One mutation significantly reduced surface expression. Application of retigabine, a recently marketed KV 7 channel opener, partially reversed these effects for the majority of analyzed mutations. INTERPRETATION: The development of severe epilepsy and cognitive decline in children carrying 5 of the 7 studied KCNQ2 mutations can be related to a dominant-negative reduction of the resulting potassium current at subthreshold membrane potentials. Other factors such as genetic modifiers have to be postulated for the remaining 2 mutations. Retigabine or similar drugs may be used as a personalized therapy for this severe disease.


Asunto(s)
Epilepsia Benigna Neonatal/genética , Predisposición Genética a la Enfermedad/genética , Canal de Potasio KCNQ2/genética , Canales de Potasio con Entrada de Voltaje/genética , Animales , Carbamatos/farmacología , Epilepsia Benigna Neonatal/fisiopatología , Humanos , Canal de Potasio KCNQ2/efectos de los fármacos , Canal de Potasio KCNQ2/fisiología , Potenciales de la Membrana/genética , Mutación Missense , Oocitos , Fenilendiaminas/farmacología , Canales de Potasio con Entrada de Voltaje/efectos de los fármacos , Canales de Potasio con Entrada de Voltaje/fisiología , Xenopus
10.
Sci Rep ; 13(1): 1491, 2023 01 27.
Artículo en Inglés | MEDLINE | ID: mdl-36707549

RESUMEN

Despite numerous prior attempts to improve knock-in (KI) efficiency, the introduction of precise base pair substitutions by the CRISPR-Cas9 technique in zebrafish remains challenging. In our efforts to generate KI zebrafish models of human CACNA1C mutations, we have tested the effect of several CRISPR determinants on KI efficiency across two sites in a single gene and developed a novel method for early selection to ameliorate KI efficiency. We identified optimal KI conditions for Cas9 protein and non-target asymmetric PAM-distal single stranded deoxynucleotide repair templates at both cacna1c sites. An effect of distance to the cut site on the KI efficiency was only observed for a single repair template conformation at one of the two sites. By combining minimally invasive early genotyping with the zebrafish embryo genotyper (ZEG) device and next-generation sequencing, we were able to obtain an almost 17-fold increase in somatic editing efficiency. The added benefit of the early selection procedure was particularly evident for alleles with lower somatic editing efficiencies. We further explored the potential of the ZEG selection procedure for the improvement of germline transmission by demonstrating germline transmission events in three groups of pre-selected embryos.


Asunto(s)
Sistemas CRISPR-Cas , Edición Génica , Animales , Humanos , Sistemas CRISPR-Cas/genética , Edición Génica/métodos , Pez Cebra/genética , Genotipo , Secuenciación de Nucleótidos de Alto Rendimiento
11.
Orphanet J Rare Dis ; 18(1): 23, 2023 01 31.
Artículo en Inglés | MEDLINE | ID: mdl-36721196

RESUMEN

BACKGROUND: The c.1124_1127delTTCA p.(Ile375Argfs*43) pathogenic variant is the most frequently identified molecular defect in the KCNQ1 gene in the cardiogenetics clinic of the Antwerp University Hospital. This variant was observed in nine families presenting with either Jervell-Lange-Nielsen syndrome or long QT syndrome (LQTS). Here, we report on the molecular, clinical and functional characterization of the KCNQ1 c.1124_1127delTTCA variant. RESULTS: Forty-one heterozygous variant harboring individuals demonstrated a predominantly mild clinical and electrophysiological phenotype, compared to individuals harboring other KCNQ1 pathogenic variants (5% symptomatic before 40 years of age, compared to 24% and 29% in p.(Tyr111Cys) and p.(Ala341Val) variant carriers, respectively, 33% with QTc ≤ 440 ms compared to 10% in p.(Tyr111Cys) and p.(Ala341Val) variant carriers). The LQTS phenotype was most comparable to that observed for the Swedish p.(Arg518*) founder mutation (7% symptomatic at any age, compared to 17% in p.(Arg518*) variant carriers, 33% with QTc ≤ 440 ms compared to 16% in p.(Arg518*) variant carriers). Surprisingly, short tandem repeat analysis did not reveal a common haplotype for all families. One KCNQ1 c.1124_1127delTTCA harboring patient was diagnosed with Brugada syndrome (BrS). The hypothesis of a LQTS/BrS overlap syndrome was supported by electrophysiological evidence for both loss-of-function and gain-of-function (acceleration of channel kinetics) in a heterologous expression system. However, BrS phenotypes were not identified in other affected individuals and allelic KCNQ1 expression testing in patient-specific induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) showed nonsense mediated decay of the c.1124_1127delTTCA allele. CONCLUSIONS: The c.1124_1127delTTCA frameshift variant shows a high prevalence in our region, despite not being confirmed as a founder mutation. This variant leads to a mild LQTS phenotype in the heterozygous state. Despite initial evidence for a gain-of-function effect based on in vitro electrophysiological assessment in CHO cells and expression of the KCNQ1 c.1124_1127delTTCA allele in patient blood cells, additional testing in iPSC-CMs showed lack of expression of the mutant allele. This suggests haploinsufficiency as the pathogenic mechanism. Nonetheless, as inter-individual differences in allele expression in (iPSC-) cardiomyocytes have not been assessed, a modifying effect on the BrS phenotype through potassium current modulation cannot be excluded.


Asunto(s)
Canal de Potasio KCNQ1 , Síndrome de QT Prolongado , Animales , Cricetinae , Alelos , Bélgica , Cricetulus , Canal de Potasio KCNQ1/genética , Humanos , Síndrome de Jervell-Lange Nielsen/genética , Síndrome de QT Prolongado/genética
12.
Biol Open ; 11(2)2022 02 15.
Artículo en Inglés | MEDLINE | ID: mdl-35195246

RESUMEN

Cardiomyocytes derived from induced pluripotent stem cells (iPSC-CMs) offer an attractive platform for cardiovascular research. Patient-specific iPSC-CMs are very useful for studying disease development, and bear potential for disease diagnostics, prognosis evaluation and development of personalized treatment. Several monolayer-based serum-free protocols have been described for the differentiation of iPSCs into cardiomyocytes, but data on their performance are scarce. In this study, we evaluated two protocols that are based on temporal modulation of the Wnt/ß-catenin pathway for iPSC-CM differentiation from four iPSC lines, including two control individuals and two patients carrying an SCN5A mutation. The SCN5A gene encodes the cardiac voltage-gated sodium channel (Nav1.5) and loss-of-function mutations can cause the cardiac arrhythmia Brugada syndrome. We performed molecular characterization of the obtained iPSC-CMs by immunostaining for cardiac specific markers and by expression analysis of selected cardiac structural and ionic channel protein-encoding genes with qPCR. We also investigated cell growth morphology, contractility and survival of the iPSC-CMs after dissociation. Finally, we performed electrophysiological characterization of the cells, focusing on the action potential (AP) and calcium transient (CT) characteristics using patch-clamping and optical imaging, respectively. Based on our comprehensive morpho-functional analysis, we concluded that both tested protocols result in a high percentage of contracting CMs. Moreover, they showed acceptable survival and cell quality after dissociation (>50% of cells with a smooth cell membrane, possible to seal during patch-clamping). Both protocols generated cells presenting with typical iPSC-CM AP and CT characteristics, although one protocol (that involves sequential addition of CHIR99021 and Wnt-C59) rendered iPSC-CMs, which were more accessible for patch-clamp and calcium transient experiments and showed an expression pattern of cardiac-specific markers more similar to this observed in human heart left ventricle samples.


Asunto(s)
Células Madre Pluripotentes Inducidas , Potenciales de Acción , Diferenciación Celular , Fenómenos Electrofisiológicos , Humanos , Miocitos Cardíacos
13.
Eur J Med Genet ; 64(11): 104322, 2021 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-34438094

RESUMEN

Sudden cardiac death (SCD) is a common cause of death in young adults. In up to 80% of cases a genetic cause is suspected. Next-generation sequencing of candidate genes can reveal the cause of SCD, provide prognostic management, and facilitate pre-symptomatic testing and prevention in relatives. Here we present a proband who experienced SCD in his sleep for which molecular autopsy was performed. We performed a post-mortem genetic analysis of a 49-year-old male who died during sleep after competitive kayaking, using a Cardiomyopathy and Primary Arrhythmia next-generation sequencing panel, each containing 51 candidate genes. Autopsy was not performed. Genetic testing of the proband resulted in missense variants in KCNQ1 (c.1449C > A; p.(Asn483Lys)) and DSG2 (c.2979G > T; p.(Gln993His)), both absent from the gnomAD database. Familial segregation analysis showed de novo occurrence of the DSG2 variant and presence of the KCNQ1 variant in the proband's mother and daughter. KCNQ1 p.(Asn483Lys) was predicted to be pathogenic by MutationTaster. However, none of the KCNQ1 variant carrying family members showed long QTc on ECG or Holter. We further functionally analysed this variant using patch-clamp in a heterologous expression system (Chinese Hamster Ovary (CHO) cells) expressing the KCNQ1 mutant in combination with KCNE1 wild type protein and showed no significant changes in electrophysiological function of Kv7.1. Based on the above evidence, we concluded that the DSG2 p.(Gln993His) variant is the most likely cause of SCD in the presented case, and that there is insufficient evidence that the identified KCNQ1 p.(Asn483Lys) variant would confer risk for SCD in his mother and daughter. Fortunately, the DSG2 variant was not inherited by the proband's two children. This case report indicates the added value of molecular autopsy and the importance of subsequent functional study of variants to inform patients and family members about the risk of variants they might carry.


Asunto(s)
Arritmias Cardíacas/genética , Muerte Súbita Cardíaca/etiología , Desmogleína 2/genética , Mutación Missense , Animales , Arritmias Cardíacas/patología , Células CHO , Cricetinae , Cricetulus , Desmogleína 2/metabolismo , Frecuencia Cardíaca , Humanos , Canal de Potasio KCNQ1/genética , Canal de Potasio KCNQ1/metabolismo , Masculino , Persona de Mediana Edad
14.
Front Cardiovasc Med ; 7: 117, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32850980

RESUMEN

Aims: Brugada syndrome (BrS) is an inherited cardiac arrhythmia with an increased risk for sudden cardiac death (SCD). About 20% of BrS cases are explained by mutations in the SCN5A gene, encoding the main cardiac sodium Nav1.5 channel. Here we present a severe case of cardiac sodium channelopathy with BrS caused by SCN5A compound heterozygous mutations. We performed a genetic analysis of SCN5A in a male proband who collapsed during cycling at the age of 2 years. Because of atrial standstill, he received a pacemaker, and at the age of 3 years, he experienced a collapse anew with left-sided brain stroke. A later ECG taken during a fever unmasked a characteristic BrS type-1 pattern. The functional effect of the detected genetic variants was investigated. Methods and Results: Next-generation sequencing allowed the detection of two SCN5A variants in trans: c.4813+3_4813+6dupGGGT-a Belgian founder mutation-and c.4711 T>C, p.Phe1571Leu. A familial segregation analysis showed the presence of the founder mutation in the proband's affected father and paternal aunt and the de novo occurrence of the p.Phe1571Leu. The functional effect of the founder mutation was previously described as a loss-of-function. We performed a functional analysis of the p.Phe571Leu variant in HEK293 cells alone or co-expressed with the ß1-subunit. Compared to the SCN5A wild type, p.Phe1571Leu displayed a hyperpolarizing shift in the voltage dependence of inactivation (loss-of-function), while the activation parameters were unaffected. Using the peptide toxin nemertide α-1, the variant's loss-of-function effect could be restored due to a toxin-dependent reduction of channel inactivation. Conclusion: This is the first report providing support for the pathogenicity of the p.Phe1571Leu SCN5A variant which, together with the c.4813+3_4813+6dupGGGT founder mutation, explains the severity of the phenotype of cardiac sodium channelopathy with BrS in the presented case.

15.
Front Med (Lausanne) ; 6: 198, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31555651

RESUMEN

Cardiogeneticsbank@UZA is an academic hospital integrated biobank that collects aortic tissue, blood, cell lines (fibroblasts, vascular smooth muscle cells, peripheral blood mononuclear cells, and induced pluripotent stem cells), and DNA from patients with cardiogenetic disorders, for both diagnostic and research purposes. We adhere to a quality management system and have established standard protocols for the sampling and processing of all cardiogenetic patient related materials. Cardiogeneticsbank@UZA is embedded in the Biobanking and Biomolecular Resources Research Infrastructure Belgium (BBMRI.be) and samples from this biobank are available for commercial and academic researchers, through an established access procedure. Currently, the extremely valuable cardiogenetics collection consists of more than 8,700 DNA samples, 380 tissue samples, and 500 cell lines of 7,578 patients, and is linked with extensive clinical data. Some interesting potential research applications are discussed.

16.
Mol Syndromol ; 9(4): 190-196, 2018 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-30140196

RESUMEN

EFEMP2 mutations are known to be responsible for autosomal recessive cutis laxa type 1B (ARCL1B), a rare multisystem disease affecting skin, skeleton, and vascular structures. We report 2 additional related cases of ARCL1B of particular severity leading to termination of pregnancy. Cardinal signs of this connective tissue disease were already seen during the second trimester of pregnancy, then confirmed and clarified at autopsy. Anomalies included cutis laxa, arachnodactyly, clubfoot, wormian bones, moderate bowing of long bones with slender bone trabeculae, rib fractures, undermuscularized diaphragm, hiatal hernia, and arterial tortuosity with thick vascular walls and disorganized elastic fibers. Sequencing of the EFEMP2 gene revealed a novel homozygous nonsense mutation: c.639C>A (p.Cys213*). We performed a thorough histological analysis and discuss differential diagnoses, genotype-phenotype correlations, and the challenge of prenatal diagnosis of this disease.

17.
Ann Cardiothorac Surg ; 6(6): 582-594, 2017 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-29270370

RESUMEN

Many different heritable connective tissue disorders (HCTD) have been described over the past decades. These syndromes often affect the connective tissue of various organ systems, including heart, blood vessels, skin, joints, bone, eyes, and lungs. The discovery of these HCTD was followed by the identification of mutations in a wide range of genes encoding structural proteins, modifying enzymes, or components of the TGFß-signaling pathway. Three typical examples of HCTD are Marfan syndrome (MFS), Ehlers-Danlos syndrome (EDS), and Loeys-Dietz syndrome (LDS). These syndromes show some degree of phenotypical overlap of cardiovascular, skeletal, and cutaneous features. MFS is typically characterized by cardiovascular, ocular, and skeletal manifestations and is caused by heterozygous mutations in FBN1, coding for the extracellular matrix (ECM) protein fibrillin-1. The most common cardiovascular phenotype involves aortic aneurysm and dissection at the sinuses of Valsalva. LDS is caused by mutations in TGBR1/2, SMAD2/3, or TGFB2/3, all coding for components of the TGFß-signaling pathway. LDS can be distinguished from MFS by the unique presence of hypertelorism, bifid uvula or cleft palate, and widespread aortic and arterial aneurysm and tortuosity. Compared to MFS, LDS cardiovascular manifestations tend to be more severe. In contrast, no association is reported between LDS and the presence of ectopia lentis, a key distinguishing feature of MFS. Overlapping features between MFS and LDS include scoliosis, pes planus, anterior chest deformity, spontaneous pneumothorax, and dural ectasia. EDS refers to a group of clinically and genetically heterogeneous connective tissue disorders and all subtypes are characterized by variable abnormalities of skin, ligaments and joints, blood vessels, and internal organs. Typical presenting features include joint hypermobility, skin hyperextensibility, and tissue fragility. Up to one quarter of the EDS patients show aortic aneurysmal disease. The latest EDS nosology distinguishes 13 subtypes. Many phenotypic features show overlap between the different subtypes, which makes the clinical diagnosis rather difficult and highlights the importance of molecular diagnostic confirmation.

18.
Eur J Hum Genet ; 23(2): 224-8, 2015 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-24736733

RESUMEN

Shprintzen-Goldberg syndrome (SGS) is a rare, systemic connective tissue disorder characterized by craniofacial, skeletal, and cardiovascular manifestations that show a significant overlap with the features observed in the Marfan (MFS) and Loeys-Dietz syndrome (LDS). A distinguishing observation in SGS patients is the presence of intellectual disability, although not all patients in this series present this finding. Recently, SGS was shown to be due to mutations in the SKI gene, encoding the oncoprotein SKI, a repressor of TGFß activity. Here, we report eight recurrent and three novel SKI mutations in eleven SGS patients. All were heterozygous missense mutations located in the R-SMAD binding domain, except for one novel in-frame deletion affecting the DHD domain. Adding our new findings to the existing data clearly reveals a mutational hotspot, with 73% (24 out of 33) of the hitherto described unrelated patients having mutations in a stretch of five SKI residues (from p.(Ser31) to p.(Pro35)). This implicates that the initial molecular testing could be focused on mutation analysis of the first half of exon 1 of SKI. As the majority of the known mutations are located in the R-SMAD binding domain of SKI, our study further emphasizes the importance of TGFß signaling in the pathogenesis of SGS.


Asunto(s)
Aracnodactilia/genética , Craneosinostosis/genética , Proteínas de Unión al ADN/genética , Síndrome de Marfan/genética , Mutación Missense , Proteínas Proto-Oncogénicas/genética , Adolescente , Adulto , Aracnodactilia/diagnóstico , Sitios de Unión , Niño , Preescolar , Craneosinostosis/diagnóstico , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/metabolismo , Exones , Femenino , Humanos , Masculino , Síndrome de Marfan/diagnóstico , Persona de Mediana Edad , Unión Proteica , Proteínas Proto-Oncogénicas/química , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Smad/metabolismo
19.
Nat Genet ; 44(11): 1249-54, 2012 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-23023332

RESUMEN

Elevated transforming growth factor (TGF)-ß signaling has been implicated in the pathogenesis of syndromic presentations of aortic aneurysm, including Marfan syndrome (MFS) and Loeys-Dietz syndrome (LDS). However, the location and character of many of the causal mutations in LDS intuitively imply diminished TGF-ß signaling. Taken together, these data have engendered controversy regarding the specific role of TGF-ß in disease pathogenesis. Shprintzen-Goldberg syndrome (SGS) has considerable phenotypic overlap with MFS and LDS, including aortic aneurysm. We identified causative variation in ten individuals with SGS in the proto-oncogene SKI, a known repressor of TGF-ß activity. Cultured dermal fibroblasts from affected individuals showed enhanced activation of TGF-ß signaling cascades and higher expression of TGF-ß-responsive genes relative to control cells. Morpholino-induced silencing of SKI paralogs in zebrafish recapitulated abnormalities seen in humans with SGS. These data support the conclusions that increased TGF-ß signaling is the mechanism underlying SGS and that high signaling contributes to multiple syndromic presentations of aortic aneurysm.


Asunto(s)
Aneurisma de la Aorta/genética , Aracnodactilia/genética , Craneosinostosis/genética , Proteínas de Unión al ADN , Síndrome de Marfan/genética , Proteínas Proto-Oncogénicas , Factor de Crecimiento Transformador beta , Animales , Aracnodactilia/metabolismo , Células Cultivadas , Craneosinostosis/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Fibroblastos , Humanos , Síndrome de Loeys-Dietz/genética , Síndrome de Marfan/metabolismo , Ratones , Mutación , Fenotipo , Fosforilación , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/metabolismo , Transducción de Señal , Factor de Crecimiento Transformador beta/antagonistas & inhibidores , Factor de Crecimiento Transformador beta/genética , Pez Cebra
20.
Nat Genet ; 44(8): 922-7, 2012 Jul 08.
Artículo en Inglés | MEDLINE | ID: mdl-22772368

RESUMEN

Loeys-Dietz syndrome (LDS) associates with a tissue signature for high transforming growth factor (TGF)-ß signaling but is often caused by heterozygous mutations in genes encoding positive effectors of TGF-ß signaling, including either subunit of the TGF-ß receptor or SMAD3, thereby engendering controversy regarding the mechanism of disease. Here, we report heterozygous mutations or deletions in the gene encoding the TGF-ß2 ligand for a phenotype within the LDS spectrum and show upregulation of TGF-ß signaling in aortic tissue from affected individuals. Furthermore, haploinsufficient Tgfb2(+/-) mice have aortic root aneurysm and biochemical evidence of increased canonical and noncanonical TGF-ß signaling. Mice that harbor both a mutant Marfan syndrome (MFS) allele (Fbn1(C1039G/+)) and Tgfb2 haploinsufficiency show increased TGF-ß signaling and phenotypic worsening in association with normalization of TGF-ß2 expression and high expression of TGF-ß1. Taken together, these data support the hypothesis that compensatory autocrine and/or paracrine events contribute to the pathogenesis of TGF-ß-mediated vasculopathies.


Asunto(s)
Aneurisma de la Aorta Torácica/genética , Mutación , Factor de Crecimiento Transformador beta2/genética , Animales , Aneurisma de la Aorta Torácica/patología , Modelos Animales de Enfermedad , Femenino , Fibrilina-1 , Fibrilinas , Haploinsuficiencia , Humanos , Síndrome de Loeys-Dietz/genética , Síndrome de Loeys-Dietz/patología , Masculino , Síndrome de Marfan/genética , Síndrome de Marfan/patología , Ratones , Ratones Noqueados , Ratones Mutantes , Proteínas de Microfilamentos/genética , Linaje , Fenotipo , Transducción de Señal , Síndrome , Factor de Crecimiento Transformador beta2/deficiencia
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