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1.
Ann Neurol ; 2023 Aug 22.
Artículo en Inglés | MEDLINE | ID: mdl-37606373

RESUMEN

OBJECTIVE: Variants in GABRA1 have been associated with a broad epilepsy spectrum, ranging from genetic generalized epilepsies to developmental and epileptic encephalopathies. However, our understanding of what determines the phenotype severity and best treatment options remains inadequate. We therefore aimed to analyze the electroclinical features and the functional effects of GABRA1 variants to establish genotype-phenotype correlations. METHODS: Genetic and electroclinical data of 27 individuals (22 unrelated and 2 families) harboring 20 different GABRA1 variants were collected and accompanied by functional analysis of 19 variants. RESULTS: Individuals in this cohort could be assigned into different clinical subgroups based on the functional effect of their variant and its structural position within the GABRA1 subunit. A homogenous phenotype with mild cognitive impairment and infantile onset epilepsy (focal seizures, fever sensitivity, and electroencephalographic posterior epileptiform discharges) was described for variants in the extracellular domain and the small transmembrane loops. These variants displayed loss-of-function (LoF) effects, and the patients generally had a favorable outcome. A more severe phenotype was associated with variants in the pore-forming transmembrane helices. These variants displayed either gain-of-function (GoF) or LoF effects. GoF variants were associated with severe early onset neurodevelopmental disorders, including early infantile developmental and epileptic encephalopathy. INTERPRETATION: Our data expand the genetic and phenotypic spectrum of GABRA1 epilepsies and permit delineation of specific subphenotypes for LoF and GoF variants, through the heterogeneity of phenotypes and variants. Generally, variants in the transmembrane helices cause more severe phenotypes, in particular GoF variants. These findings establish the basis for a better understanding of the pathomechanism and a precision medicine approach in GABRA1-related disorders. Further studies in larger populations are needed to provide a conclusive genotype-phenotype correlation. ANN NEUROL 2023.

2.
Basic Res Cardiol ; 118(1): 8, 2023 03 02.
Artículo en Inglés | MEDLINE | ID: mdl-36862248

RESUMEN

Whereas cardiomyocytes (CMs) in the fetal heart divide, postnatal CMs fail to undergo karyokinesis and/or cytokinesis and therefore become polyploid or binucleated, a key process in terminal CM differentiation. This switch from a diploid proliferative CM to a terminally differentiated polyploid CM remains an enigma and seems an obstacle for heart regeneration. Here, we set out to identify the transcriptional landscape of CMs around birth using single cell RNA sequencing (scRNA-seq) to predict transcription factors (TFs) involved in CM proliferation and terminal differentiation. To this end, we established an approach combining fluorescence activated cell sorting (FACS) with scRNA-seq of fixed CMs from developing (E16.5, P1, and P5) mouse hearts, and generated high-resolution single-cell transcriptomic maps of in vivo diploid and tetraploid CMs, increasing the CM resolution. We identified TF-networks regulating the G2/M phases of developing CMs around birth. ZEB1 (Zinc Finger E-Box Binding Homeobox 1), a hereto unknown TF in CM cell cycling, was found to regulate the highest number of cell cycle genes in cycling CMs at E16.5 but was downregulated around birth. CM ZEB1-knockdown reduced proliferation of E16.5 CMs, while ZEB1 overexpression at P0 after birth resulted in CM endoreplication. These data thus provide a ploidy stratified transcriptomic map of developing CMs and bring new insight to CM proliferation and endoreplication identifying ZEB1 as a key player in these processes.


Asunto(s)
Miocitos Cardíacos , Transcriptoma , Animales , Ratones , Proliferación Celular , Genes Homeobox , Ploidias , Poliploidía , Homeobox 1 de Unión a la E-Box con Dedos de Zinc , Dedos de Zinc
3.
Genet Med ; 25(9): 100894, 2023 09.
Artículo en Inglés | MEDLINE | ID: mdl-37183800

RESUMEN

PURPOSE: The "NALCN channelosome" is an ion channel complex that consists of multiple proteins, including NALCN, UNC79, UNC80, and FAM155A. Only a small number of individuals with a neurodevelopmental syndrome have been reported with disease causing variants in NALCN and UNC80. However, no pathogenic UNC79 variants have been reported, and in vivo function of UNC79 in humans is largely unknown. METHODS: We used international gene-matching efforts to identify patients harboring ultrarare heterozygous loss-of-function UNC79 variants and no other putative responsible genes. We used genetic manipulations in Drosophila and mice to test potential causal relationships between UNC79 variants and the pathology. RESULTS: We found 6 unrelated and affected patients with UNC79 variants. Five patients presented with overlapping neurodevelopmental features, including mild to moderate intellectual disability and a mild developmental delay, whereas a single patient reportedly had normal cognitive and motor development but was diagnosed with epilepsy and autistic features. All displayed behavioral issues and 4 patients had epilepsy. Drosophila with UNC79 knocked down displayed induced seizure-like phenotype. Mice with a heterozygous loss-of-function variant have a developmental delay in body weight compared with wild type. In addition, they have impaired ability in learning and memory. CONCLUSION: Our results demonstrate that heterozygous loss-of-function UNC79 variants are associated with neurologic pathologies.


Asunto(s)
Epilepsia , Discapacidad Intelectual , Proteínas de la Membrana , Trastornos del Neurodesarrollo , Animales , Humanos , Ratones , Drosophila/genética , Discapacidad Intelectual/genética , Discapacidad Intelectual/patología , Trastornos del Neurodesarrollo/genética , Fenotipo , Proteínas de la Membrana/genética
4.
Mol Genet Metab ; 140(3): 107694, 2023 11.
Artículo en Inglés | MEDLINE | ID: mdl-37708665

RESUMEN

Creatine transporter deficiency (CTD), caused by pathogenic variants in SLC6A8, is the second most common cause of X-linked intellectual disability. Symptoms include intellectual disability, epilepsy, and behavioral disorders and are caused by reduced cerebral creatine levels. Targeted treatment with oral supplementation is available, however the treatment efficacy is still being investigated. There are clinical and theoretical indications that heterozygous females with CTD respond better to supplementation treatment than hemizygous males. Unfortunately, heterozygous females with CTD often have more subtle and uncharacteristic clinical and biochemical phenotypes, rendering diagnosis more difficult. We report a new female case who presented with learning disabilities and seizures. After determining the diagnosis with molecular genetic testing confirmed by proton magnetic resonance spectroscopy (1H-MRS), the patient was treated with supplementation treatment including creatine, arginine, and glycine. After 28 months of treatment, the patient showed prominent clinical improvement and increased creatine levels in the brain. Furthermore, we provide a review of the 32 female cases reported in the current literature including a description of phenotypes, genotypes, diagnostic approaches, and effects of supplementation treatment. Based on this, we find that supplementation treatment should be tested in heterozygous female patients with CTD, and a prospective treatment underlines the importance of diagnosing these patients. The diagnosis should be suspected in a broad clinical spectrum of female patients and can only be made by molecular genetic testing. 1H-MRS of cerebral creatine levels is essential for establishing the diagnosis in females, and especially valuable when assessing variants of unknown significance.


Asunto(s)
Encefalopatías Metabólicas Innatas , Discapacidad Intelectual , Discapacidad Intelectual Ligada al Cromosoma X , Masculino , Humanos , Femenino , Discapacidad Intelectual/genética , Creatina , Encefalopatías Metabólicas Innatas/diagnóstico , Encefalopatías Metabólicas Innatas/genética , Encefalopatías Metabólicas Innatas/tratamiento farmacológico , Discapacidad Intelectual Ligada al Cromosoma X/diagnóstico , Discapacidad Intelectual Ligada al Cromosoma X/genética , Proteínas de Transporte de Neurotransmisores en la Membrana Plasmática/genética , Proteínas del Tejido Nervioso
5.
Epilepsia ; 64(8): e170-e176, 2023 08.
Artículo en Inglés | MEDLINE | ID: mdl-37114479

RESUMEN

IRF2BPL has recently been described as a novel cause of neurodevelopmental disorders with multisystemic regression, epilepsy, cerebellar symptoms, dysphagia, dystonia, and pyramidal signs. We describe a novel IRF2BPL phenotype consistent with progressive myoclonus epilepsy (PME) in three novel subjects and review the features of the 31 subjects with IRF2BPL-related disorders previously reported. Our three probands, aged 28-40 years, harbored de novo nonsense variants in IRF2BPL (c.370C > T, p.[Gln124*] and c.364C > T; p.[Gln122*], respectively). From late childhood/adolescence, they presented with severe myoclonus epilepsy, stimulus-sensitive myoclonus, and progressive cognitive, speech, and cerebellar impairment, consistent with a typical PME syndrome. The skin biopsy revealed massive intracellular glycogen inclusions in one proband, suggesting a similar pathogenic pathway to other storage disorders. Whereas the two older probands were severely affected, the younger proband had a milder PME phenotype, partially overlapping with some of the previously reported IRF2BPL cases, suggesting that some of them might be unrecognized PME. Interestingly, all three patients harbored protein-truncating variants clustered in a proximal, highly conserved gene region around the "coiled-coil" domain. Our data show that PME can be an additional phenotype within the spectrum of IRF2BPL-related disorders and suggest IRF2BPL as a novel causative gene for PME.


Asunto(s)
Epilepsias Mioclónicas , Epilepsia , Epilepsias Mioclónicas Progresivas , Mioclonía , Humanos , Niño , Mutación , Epilepsias Mioclónicas Progresivas/genética , Epilepsias Mioclónicas/patología , Familia , Proteínas Portadoras/genética , Proteínas Nucleares/genética
6.
Brain ; 145(4): 1299-1309, 2022 05 24.
Artículo en Inglés | MEDLINE | ID: mdl-34633442

RESUMEN

A potential link between GABRD encoding the δ subunit of extrasynaptic GABAA receptors and neurodevelopmental disorders has largely been disregarded due to conflicting conclusions from early studies. However, we identified seven heterozygous missense GABRD variants in 10 patients with neurodevelopmental disorders and generalized epilepsy. One variant occurred in two sibs of healthy parents with presumed somatic mosaicism, another segregated with the disease in three affected family members, and the remaining five occurred de novo in sporadic patients. Electrophysiological measurements were used to determine the functional consequence of the seven missense δ subunit variants in receptor combinations of α1ß3δ and α4ß2δ GABAA receptors. This was accompanied by analysis of electroclinical phenotypes of the affected individuals. We determined that five of the seven variants caused altered function of the resulting α1ß3δ and α4ß2δ GABAA receptors. Surprisingly, four of the five variants led to gain-of-function effects, whereas one led to a loss-of-function effect. The stark differences between the gain-of-function and loss-of function effects were mirrored by the clinical phenotypes. Six patients with gain-of-function variants shared common phenotypes: neurodevelopmental disorders with behavioural issues, various degrees of intellectual disability, generalized epilepsy with atypical absences and generalized myoclonic and/or bilateral tonic-clonic seizures. The EEG showed qualitative analogies among the different gain-of-function variant carriers consisting of focal slowing in the occipital regions often preceding irregular generalized epileptiform discharges, with frontal predominance. In contrast, the one patient carrying a loss-of-function variant had normal intelligence and no seizure history, but has a diagnosis of autism spectrum disorder and suffers from elevated internalizing psychiatric symptoms. We hypothesize that increase in tonic GABA-evoked current levels mediated by δ-containing extrasynaptic GABAA receptors lead to abnormal neurotransmission, which represent a novel mechanism for severe neurodevelopmental disorders. In support of this, the electroclinical findings for the gain-of-function GABRD variants resemble the phenotypic spectrum reported in patients with missense SLC6A1 (GABA uptake transporter) variants. This also indicates that the phenomenon of extrasynaptic receptor overactivity is observed in a broader range of patients with neurodevelopmental disorders, because SLC6A1 loss-of-function variants also lead to overactive extrasynaptic δ-containing GABAA receptors. These findings have implications when selecting potential treatment options, as a substantial portion of available antiseizure medication act by enhancing GABAergic function either directly or indirectly, which could exacerbate symptoms in patients with gain-of-function GABRD variants.


Asunto(s)
Trastorno del Espectro Autista , Epilepsia Generalizada , Epilepsia , Proteínas Transportadoras de GABA en la Membrana Plasmática , Trastornos del Neurodesarrollo , Trastorno del Espectro Autista/genética , Epilepsia/genética , Epilepsia Generalizada/genética , Proteínas Transportadoras de GABA en la Membrana Plasmática/genética , Proteínas Transportadoras de GABA en la Membrana Plasmática/metabolismo , Mutación con Ganancia de Función , Humanos , Trastornos del Neurodesarrollo/genética , Receptores de GABA-A/genética , Receptores de GABA-A/metabolismo , Convulsiones/genética , Ácido gamma-Aminobutírico/metabolismo
7.
Brain ; 145(9): 2991-3009, 2022 09 14.
Artículo en Inglés | MEDLINE | ID: mdl-34431999

RESUMEN

We report detailed functional analyses and genotype-phenotype correlations in 392 individuals carrying disease-causing variants in SCN8A, encoding the voltage-gated Na+ channel Nav1.6, with the aim of describing clinical phenotypes related to functional effects. Six different clinical subgroups were identified: Group 1, benign familial infantile epilepsy (n = 15, normal cognition, treatable seizures); Group 2, intermediate epilepsy (n = 33, mild intellectual disability, partially pharmaco-responsive); Group 3, developmental and epileptic encephalopathy (n = 177, severe intellectual disability, majority pharmaco-resistant); Group 4, generalized epilepsy (n = 20, mild to moderate intellectual disability, frequently with absence seizures); Group 5, unclassifiable epilepsy (n = 127); and Group 6, neurodevelopmental disorder without epilepsy (n = 20, mild to moderate intellectual disability). Those in Groups 1-3 presented with focal or multifocal seizures (median age of onset: 4 months) and focal epileptiform discharges, whereas the onset of seizures in patients with generalized epilepsy was later (median: 42 months) with generalized epileptiform discharges. We performed functional studies expressing missense variants in ND7/23 neuroblastoma cells and primary neuronal cultures using recombinant tetrodotoxin-insensitive human Nav1.6 channels and whole-cell patch-clamping. Two variants causing developmental and epileptic encephalopathy showed a strong gain-of-function (hyperpolarizing shift of steady-state activation, strongly increased neuronal firing rate) and one variant causing benign familial infantile epilepsy or intermediate epilepsy showed a mild gain-of-function (defective fast inactivation, less increased firing). In contrast, all three variants causing generalized epilepsy induced a loss-of-function (reduced current amplitudes, depolarizing shift of steady-state activation, reduced neuronal firing). Functional effects were known for 170 individuals. All 136 individuals carrying a functionally tested gain-of-function variant had either focal (n = 97, Groups 1-3) or unclassifiable (n = 39) epilepsy, whereas 34 individuals with a loss-of-function variant had either generalized (n = 14), no (n = 11) or unclassifiable (n = 6) epilepsy; only three had developmental and epileptic encephalopathy. Computational modelling in the gain-of-function group revealed a significant correlation between the severity of the electrophysiological and clinical phenotypes. Gain-of-function variant carriers responded significantly better to sodium channel blockers than to other anti-seizure medications, and the same applied for all individuals in Groups 1-3. In conclusion, our data reveal clear genotype-phenotype correlations between age at seizure onset, type of epilepsy and gain- or loss-of-function effects of SCN8A variants. Generalized epilepsy with absence seizures is the main epilepsy phenotype of loss-of-function variant carriers and the extent of the electrophysiological dysfunction of the gain-of-function variants is a main determinant of the severity of the clinical phenotype in focal epilepsies. Our pharmacological data indicate that sodium channel blockers present a treatment option in SCN8A-related focal epilepsy with onset in the first year of life.


Asunto(s)
Epilepsia Generalizada , Síndromes Epilépticos , Discapacidad Intelectual , Canal de Sodio Activado por Voltaje NAV1.6 , Epilepsia Generalizada/tratamiento farmacológico , Epilepsia Generalizada/genética , Síndromes Epilépticos/tratamiento farmacológico , Síndromes Epilépticos/genética , Estudios de Asociación Genética , Humanos , Lactante , Discapacidad Intelectual/genética , Mutación , Canal de Sodio Activado por Voltaje NAV1.6/genética , Pronóstico , Convulsiones/tratamiento farmacológico , Convulsiones/genética , Bloqueadores de los Canales de Sodio/uso terapéutico
8.
Brain ; 144(12): 3635-3650, 2021 12 31.
Artículo en Inglés | MEDLINE | ID: mdl-34114611

RESUMEN

Variants in KCNT1, encoding a sodium-gated potassium channel (subfamily T member 1), have been associated with a spectrum of epilepsies and neurodevelopmental disorders. These range from familial autosomal dominant or sporadic sleep-related hypermotor epilepsy to epilepsy of infancy with migrating focal seizures (EIMFS) and include developmental and epileptic encephalopathies. This study aims to provide a comprehensive overview of the phenotypic and genotypic spectrum of KCNT1 mutation-related epileptic disorders in 248 individuals, including 66 previously unpublished and 182 published cases, the largest cohort reported so far. Four phenotypic groups emerged from our analysis: (i) EIMFS (152 individuals, 33 previously unpublished); (ii) developmental and epileptic encephalopathies other than EIMFS (non-EIMFS developmental and epileptic encephalopathies) (37 individuals, 17 unpublished); (iii) autosomal dominant or sporadic sleep-related hypermotor epilepsy (53 patients, 14 unpublished); and (iv) other phenotypes (six individuals, two unpublished). In our cohort of 66 new cases, the most common phenotypic features were: (i) in EIMFS, heterogeneity of seizure types, including epileptic spasms, epilepsy improvement over time, no epilepsy-related deaths; (ii) in non-EIMFS developmental and epileptic encephalopathies, possible onset with West syndrome, occurrence of atypical absences, possible evolution to developmental and epileptic encephalopathies with sleep-related hypermotor epilepsy features; one case of sudden unexplained death in epilepsy; (iii) in autosomal dominant or sporadic sleep-related hypermotor epilepsy, we observed a high prevalence of drug-resistance, although seizure frequency improved with age in some individuals, appearance of cognitive regression after seizure onset in all patients, no reported severe psychiatric disorders, although behavioural/psychiatric comorbidities were reported in ∼50% of the patients, sudden unexplained death in epilepsy in one individual; and (iv) other phenotypes in individuals with mutation of KCNT1 included temporal lobe epilepsy, and epilepsy with tonic-clonic seizures and cognitive regression. Genotypic analysis of the whole cohort of 248 individuals showed only missense mutations and one inframe deletion in KCNT1. Although the KCNT1 mutations in affected individuals were seen to be distributed among the different domains of the KCNT1 protein, genotype-phenotype considerations showed many of the autosomal dominant or sporadic sleep-related hypermotor epilepsy-associated mutations to be clustered around the RCK2 domain in the C terminus, distal to the NADP domain. Mutations associated with EIMFS/non-EIMFS developmental and epileptic encephalopathies did not show a particular pattern of distribution in the KCNT1 protein. Recurrent KCNT1 mutations were seen to be associated with both severe and less severe phenotypes. Our study further defines and broadens the phenotypic and genotypic spectrums of KCNT1-related epileptic conditions and emphasizes the increasingly important role of this gene in the pathogenesis of early onset developmental and epileptic encephalopathies as well as of focal epilepsies, namely autosomal dominant or sporadic sleep-related hypermotor epilepsy.


Asunto(s)
Epilepsia/genética , Proteínas del Tejido Nervioso/genética , Canales de potasio activados por Sodio/genética , Adolescente , Niño , Preescolar , Estudios de Cohortes , Femenino , Genotipo , Humanos , Lactante , Masculino , Mutación , Fenotipo , Adulto Joven
9.
Genet Med ; 23(2): 363-373, 2021 02.
Artículo en Inglés | MEDLINE | ID: mdl-33144681

RESUMEN

PURPOSE: Pathogenic variants in the X-linked gene NEXMIF (previously KIAA2022) are associated with intellectual disability (ID), autism spectrum disorder, and epilepsy. We aimed to delineate the female and male phenotypic spectrum of NEXMIF encephalopathy. METHODS: Through an international collaboration, we analyzed the phenotypes and genotypes of 87 patients with NEXMIF encephalopathy. RESULTS: Sixty-three females and 24 males (46 new patients) with NEXMIF encephalopathy were studied, with 30 novel variants. Phenotypic features included developmental delay/ID in 86/87 (99%), seizures in 71/86 (83%) and multiple comorbidities. Generalized seizures predominated including myoclonic seizures and absence seizures (both 46/70, 66%), absence with eyelid myoclonia (17/70, 24%), and atonic seizures (30/70, 43%). Males had more severe developmental impairment; females had epilepsy more frequently, and varied from unaffected to severely affected. All NEXMIF pathogenic variants led to a premature stop codon or were deleterious structural variants. Most arose de novo, although X-linked segregation occurred for both sexes. Somatic mosaicism occurred in two males and a family with suspected parental mosaicism. CONCLUSION: NEXMIF encephalopathy is an X-linked, generalized developmental and epileptic encephalopathy characterized by myoclonic-atonic epilepsy overlapping with eyelid myoclonia with absence. Some patients have developmental encephalopathy without epilepsy. Males have more severe developmental impairment. NEXMIF encephalopathy arises due to loss-of-function variants.


Asunto(s)
Trastorno del Espectro Autista , Encefalopatías , Epilepsia , Trastorno del Espectro Autista/genética , Encefalopatías/genética , Epilepsia/genética , Femenino , Genes Ligados a X/genética , Humanos , Masculino , Proteínas del Tejido Nervioso , Convulsiones/genética
10.
Brain ; 143(4): 1114-1126, 2020 04 01.
Artículo en Inglés | MEDLINE | ID: mdl-32293671

RESUMEN

Congenital disorders of glycosylation are a growing group of rare genetic disorders caused by deficient protein and lipid glycosylation. Here, we report the clinical, biochemical, and molecular features of seven patients from four families with GALNT2-congenital disorder of glycosylation (GALNT2-CDG), an O-linked glycosylation disorder. GALNT2 encodes the Golgi-localized polypeptide N-acetyl-d-galactosamine-transferase 2 isoenzyme. GALNT2 is widely expressed in most cell types and directs initiation of mucin-type protein O-glycosylation. All patients showed loss of O-glycosylation of apolipoprotein C-III, a non-redundant substrate for GALNT2. Patients with GALNT2-CDG generally exhibit a syndrome characterized by global developmental delay, intellectual disability with language deficit, autistic features, behavioural abnormalities, epilepsy, chronic insomnia, white matter changes on brain MRI, dysmorphic features, decreased stature, and decreased high density lipoprotein cholesterol levels. Rodent (mouse and rat) models of GALNT2-CDG recapitulated much of the human phenotype, including poor growth and neurodevelopmental abnormalities. In behavioural studies, GALNT2-CDG mice demonstrated cerebellar motor deficits, decreased sociability, and impaired sensory integration and processing. The multisystem nature of phenotypes in patients and rodent models of GALNT2-CDG suggest that there are multiple non-redundant protein substrates of GALNT2 in various tissues, including brain, which are critical to normal growth and development.


Asunto(s)
Apolipoproteína C-III/sangre , Discapacidades del Desarrollo/genética , N-Acetilgalactosaminiltransferasas/genética , Adolescente , Animales , Apolipoproteína C-III/genética , Niño , Preescolar , Femenino , Glicosilación , Humanos , Mutación con Pérdida de Función , Masculino , Ratones , Linaje , Ratas , Adulto Joven , Polipéptido N-Acetilgalactosaminiltransferasa
11.
Epilepsia ; 61(3): 387-399, 2020 03.
Artículo en Inglés | MEDLINE | ID: mdl-32090326

RESUMEN

OBJECTIVE: Voltage-gated sodium channels (SCNs) share similar amino acid sequence, structure, and function. Genetic variants in the four human brain-expressed SCN genes SCN1A/2A/3A/8A have been associated with heterogeneous epilepsy phenotypes and neurodevelopmental disorders. To better understand the biology of seizure susceptibility in SCN-related epilepsies, our aim was to determine similarities and differences between sodium channel disorders, allowing us to develop a broader perspective on precision treatment than on an individual gene level alone. METHODS: We analyzed genotype-phenotype correlations in large SCN-patient cohorts and applied variant constraint analysis to identify severe sodium channel disease. We examined temporal patterns of human SCN expression and correlated functional data from in vitro studies with clinical phenotypes across different sodium channel disorders. RESULTS: Comparing 865 epilepsy patients (504 SCN1A, 140 SCN2A, 171 SCN8A, four SCN3A, 46 copy number variation [CNV] cases) and analysis of 114 functional studies allowed us to identify common patterns of presentation. All four epilepsy-associated SCN genes demonstrated significant constraint in both protein truncating and missense variation when compared to other SCN genes. We observed that age at seizure onset is related to SCN gene expression over time. Individuals with gain-of-function SCN2A/3A/8A missense variants or CNV duplications share similar characteristics, most frequently present with early onset epilepsy (<3 months), and demonstrate good response to sodium channel blockers (SCBs). Direct comparison of corresponding SCN variants across different SCN subtypes illustrates that the functional effects of variants in corresponding channel locations are similar; however, their clinical manifestation differs, depending on their role in different types of neurons in which they are expressed. SIGNIFICANCE: Variant function and location within one channel can serve as a surrogate for variant effects across related sodium channels. Taking a broader view on precision treatment suggests that in those patients with a suspected underlying genetic epilepsy presenting with neonatal or early onset seizures (<3 months), SCBs should be considered.


Asunto(s)
Síndromes Epilépticos/genética , Canal de Sodio Activado por Voltaje NAV1.1/genética , Canal de Sodio Activado por Voltaje NAV1.2/genética , Canal de Sodio Activado por Voltaje NAV1.3/genética , Canal de Sodio Activado por Voltaje NAV1.6/genética , Canales de Sodio/genética , Edad de Inicio , Trastorno del Espectro Autista/genética , Trastorno del Espectro Autista/fisiopatología , Niño , Preescolar , Codón sin Sentido , Variaciones en el Número de Copia de ADN , Electroencefalografía , Síndromes Epilépticos/tratamiento farmacológico , Síndromes Epilépticos/fisiopatología , Femenino , Mutación con Ganancia de Función , Eliminación de Gen , Duplicación de Gen , Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Genotipo , Humanos , Lactante , Recién Nacido , Mutación con Pérdida de Función , Masculino , Mutación Missense , Canal de Sodio Activado por Voltaje NAV1.1/metabolismo , Canal de Sodio Activado por Voltaje NAV1.2/metabolismo , Canal de Sodio Activado por Voltaje NAV1.3/metabolismo , Canal de Sodio Activado por Voltaje NAV1.6/metabolismo , Trastornos del Neurodesarrollo/genética , Trastornos del Neurodesarrollo/fisiopatología , Fenotipo , Bloqueadores de los Canales de Sodio/uso terapéutico , Canales de Sodio/metabolismo
12.
Brain ; 142(10): 3009-3027, 2019 10 01.
Artículo en Inglés | MEDLINE | ID: mdl-31504254

RESUMEN

N-methyl d-aspartate receptors are ligand-gated ionotropic receptors mediating a slow, calcium-permeable component of excitatory synaptic transmission in the CNS. Variants in genes encoding NMDAR subunits have been associated with a spectrum of neurodevelopmental disorders. Here we report six novel GRIN2D variants and one previously-described disease-associated GRIN2D variant in two patients with developmental and epileptic encephalopathy. GRIN2D encodes for the GluN2D subunit protein; the GluN2D amino acids affected by the variants in this report are located in the pre-M1 helix, transmembrane domain M3, and the intracellular carboxyl terminal domain. Functional analysis in vitro reveals that all six variants decreased receptor surface expression, which may underline some shared clinical symptoms. In addition the GluN2D(Leu670Phe), (Ala675Thr) and (Ala678Asp) substitutions confer significantly enhanced agonist potency, and/or increased channel open probability, while the GluN2D(Ser573Phe), (Ser1271Phe) and (Arg1313Trp) substitutions result in a mild increase of agonist potency, reduced sensitivity to endogenous protons, and decreased channel open probability. The GluN2D(Ser573Phe), (Ala675Thr), and (Ala678Asp) substitutions significantly decrease current amplitude, consistent with reduced surface expression. The GluN2D(Leu670Phe) variant slows current response deactivation time course and increased charge transfer. GluN2D(Ala678Asp) transfection significantly decreased cell viability of rat cultured cortical neurons. In addition, we evaluated a set of FDA-approved NMDAR channel blockers to rescue functional changes of mutant receptors. This work suggests the complexity of the pathological mechanisms of GRIN2D-mediated developmental and epileptic encephalopathy, as well as the potential benefit of precision medicine.


Asunto(s)
Epilepsia Generalizada/genética , Receptores de N-Metil-D-Aspartato/genética , Adulto , Secuencia de Aminoácidos/genética , Animales , Niño , Preescolar , Epilepsia Generalizada/fisiopatología , Femenino , Regulación de la Expresión Génica/genética , Ácido Glutámico/metabolismo , Células HEK293 , Humanos , Masculino , Neuronas/metabolismo , Polimorfismo de Nucleótido Simple/genética , Ratas , Ratas Sprague-Dawley , Receptores de N-Metil-D-Aspartato/metabolismo , Transmisión Sináptica/genética
13.
Neurobiol Dis ; 132: 104581, 2019 12.
Artículo en Inglés | MEDLINE | ID: mdl-31445161

RESUMEN

Mutations in parkin, encoded by the PARK2 gene, causes early-onset familial Parkinson's disease (PD), but dysfunctional parkin has also been implicated in sporadic PD. By combining human isogenic induced pluripotent stem cells (iPSCs) with and without PARK2 knockout (KO) and a novel large-scale mass spectrometry based proteomics and post-translational modification (PTM)-omics approach, we have mapped changes in protein profiles and PTMs caused by parkin deficiency in neurons. Our study identifies changes to several proteins previously shown to be dysregulated in brains of sporadic PD patients. Pathway analysis and subsequent in vitro assays reveal perturbations in migration and neurite outgrowth in the PARK2 KO neurons. We confirm the neurite defects using long-term engraftment of neurons in the striatum of immunosuppressed hemiparkinsonian adult rats. The GTP-binding protein RhoA was identified as a key upstream regulator, and RhoA activity was significantly increased in PARK2 KO neurons. By inhibiting RhoA signalling the migration and neurite outgrowth phenotypes could be rescued. Our study provides new insight into the pathogenesis of PD and demonstrates the broadly applicable potential of proteomics and PTMomics for elucidating the role of disease-causing mutations.


Asunto(s)
Movimiento Celular/fisiología , Neuronas Dopaminérgicas/metabolismo , Neurogénesis/fisiología , Enfermedad de Parkinson/metabolismo , Ubiquitina-Proteína Ligasas/genética , Proteína de Unión al GTP rhoA/metabolismo , Animales , Técnicas de Inactivación de Genes , Humanos , Células Madre Pluripotentes Inducidas , Mutación , Enfermedad de Parkinson/genética , Ratas , Transducción de Señal/fisiología , Ubiquitina-Proteína Ligasas/deficiencia
14.
Brain Behav Immun ; 82: 279-297, 2019 11.
Artículo en Inglés | MEDLINE | ID: mdl-31505254

RESUMEN

BACKGROUND: Although tumor necrosis factor (TNF) inhibitors are used to treat chronic inflammatory diseases, there is little information about how long-term inhibition of TNF affects the homeostatic functions that TNF maintains in the intact CNS. MATERIALS AND METHODS: To assess whether developmental TNF deficiency causes alterations in the naïve CNS, we estimated the number of proliferating cells, microglia, and neurons in the developing neocortex of E13.5, P7 and adult TNF knock out (TNF-/-) mice and wildtype (WT) littermates. We also measured changes in gene and protein expression and monoamine levels in adult WT and TNF-/- mice. To evaluate long-term effects of TNF inhibitors, we treated healthy adult C57BL/6 mice with either saline, the selective soluble TNF inhibitor XPro1595, or the nonselective TNF inhibitor etanercept. We estimated changes in cell number and protein expression after two months of treatment. We assessed the effects of TNF deficiency on cognition by testing adult WT and TNF-/- mice and mice treated with saline, XPro1595, or etanercept with specific behavioral tasks. RESULTS: TNF deficiency decreased the number of proliferating cells and microglia and increased the number of neurons. At the same time, TNF deficiency decreased the expression of WNT signaling-related proteins, specifically Collagen Triple Helix Repeat Containing 1 (CTHRC1) and Frizzled receptor 6 (FZD6). In contrast to XPro1595, long-term inhibition of TNF with etanercept in adult C57BL/6 mice decreased the number of BrdU+ cells in the granule cell layer of the dentate gyrus. Etanercept, but not XPro1595, also impaired spatial learning and memory in the Barnes maze memory test. CONCLUSION: TNF deficiency impacts the organization of neurogenic zones and alters the cell composition in brain. Long-term inhibition of TNF with the nonselective TNF inhibitor etanercept, but not the soluble TNF inhibitor XPro1595, decreases neurogenesis in the adult mouse hippocampus and impairs learning and memory after two months of treatment.


Asunto(s)
Corteza Cerebral/metabolismo , Microglía/metabolismo , Neuronas/metabolismo , Factor de Necrosis Tumoral alfa/deficiencia , Animales , Corteza Cerebral/citología , Corteza Cerebral/efectos de los fármacos , Cognición/efectos de los fármacos , Etanercept/farmacología , Hipocampo/citología , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Masculino , Memoria/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microglía/citología , Microglía/efectos de los fármacos , Neurogénesis/efectos de los fármacos , Neurogénesis/fisiología , Neuronas/citología , Neuronas/efectos de los fármacos , Inhibidores del Factor de Necrosis Tumoral/farmacología , Factor de Necrosis Tumoral alfa/metabolismo , Factor de Necrosis Tumoral alfa/farmacología , Vía de Señalización Wnt
15.
Epilepsia ; 60(5): 830-844, 2019 05.
Artículo en Inglés | MEDLINE | ID: mdl-30968951

RESUMEN

OBJECTIVE: Pathogenic variants in SCN8A have been associated with a wide spectrum of epilepsy phenotypes, ranging from benign familial infantile seizures (BFIS) to epileptic encephalopathies with variable severity. Furthermore, a few patients with intellectual disability (ID) or movement disorders without epilepsy have been reported. The vast majority of the published SCN8A patients suffer from severe developmental and epileptic encephalopathy (DEE). In this study, we aimed to provide further insight on the spectrum of milder SCN8A-related epilepsies. METHODS: A cohort of 1095 patients were screened using a next generation sequencing panel. Further patients were ascertained from a network of epilepsy genetics clinics. Patients with severe DEE and BFIS were excluded from the study. RESULTS: We found 36 probands who presented with an SCN8A-related epilepsy and normal intellect (33%) or mild (61%) to moderate ID (6%). All patients presented with epilepsy between age 1.5 months and 7 years (mean = 13.6 months), and 58% of these became seizure-free, two-thirds on monotherapy. Neurological disturbances included ataxia (28%) and hypotonia (19%) as the most prominent features. Interictal electroencephalogram was normal in 41%. Several recurrent variants were observed, including Ile763Val, Val891Met, Gly1475Arg, Gly1483Lys, Phe1588Leu, Arg1617Gln, Ala1650Val/Thr, Arg1872Gln, and Asn1877Ser. SIGNIFICANCE: With this study, we explore the electroclinical features of an intermediate SCN8A-related epilepsy with mild cognitive impairment, which is for the majority a treatable epilepsy.


Asunto(s)
Epilepsia/genética , Mutación Missense , Canal de Sodio Activado por Voltaje NAV1.6/genética , Anticonvulsivantes/uso terapéutico , Ataxia/genética , Niño , Preescolar , Disfunción Cognitiva/genética , Electroencefalografía , Epilepsia/tratamiento farmacológico , Epilepsia/fisiopatología , Pruebas Genéticas , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Lactante , Discapacidad Intelectual/genética , Trastornos del Desarrollo del Lenguaje/genética , Trastornos del Movimiento/genética , Hipotonía Muscular/genética , Linaje , Índice de Severidad de la Enfermedad
16.
Brain Behav Immun ; 65: 296-311, 2017 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-28587928

RESUMEN

Interleukin-6 (IL-6) is a pleiotropic cytokine with neuroprotective properties. Still, the therapeutic potential of IL-6 after experimental stroke has not yet been investigated in a clinically relevant way. Here, we investigated the therapeutic use of intravenously administered IL-6 and the soluble IL-6 receptor (sIL-6R) alone or in combination, early after permanent middle cerebral artery occlusion (pMCAo) in mice. IL-6 did not affect the infarct volume in C57BL/6 mice, at neither 24 nor 72h after pMCAo but reduced the infarct volume in IL-6 knockout mice at 24h after pMCAo. Assessment of post-stroke behavior showed an improved grip strength after a single IL-6 injection and also improved rotarod endurance after two injections, in C57BL/6 mice at 24h. An improved grip strength and a better preservation of sensory functions was also observed in IL-6 treated IL-6 knockout mice 24h after pMCAo. Co-administration of IL-6 and sIL-6R increased the infarct volume, the number of infiltrating polymorphonuclear leukocytes and impaired the rotarod endurance of C57BL/6 mice 24h after pMCAo. IL-6 administration to naïve C57BL/6 mice lead after 45min to increased plasma-levels of CXCL1 and IL-10, whereas IL-6 administration to C57BL/6 mice lead to a reduction in the ischemia-induced increase in IL-6 and CXCL1 at both mRNA and protein level in brain, and of IL-6 and CXCL1 in serum. We also investigated the expression of IL-6 and IL-6R after pMCAo and found that cortical neurons upregulated IL-6 mRNA and protein, and upregulated IL-6R after pMCAo. In conclusion, the results show a complex but potentially beneficial effect of intravenously administered IL-6 in experimental stroke.


Asunto(s)
Isquemia Encefálica/tratamiento farmacológico , Interleucina-6/farmacología , Animales , Encéfalo/metabolismo , Isquemia Encefálica/metabolismo , Citocinas/metabolismo , Modelos Animales de Enfermedad , Infarto de la Arteria Cerebral Media/metabolismo , Inflamación/metabolismo , Interleucina-6/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neuronas/metabolismo , ARN Mensajero , Receptores de Interleucina-6/metabolismo , Receptores de Interleucina-6/fisiología , Receptores de Interleucina-6/uso terapéutico , Accidente Cerebrovascular/metabolismo , Resultado del Tratamiento
17.
J Immunol ; 194(3): 1141-53, 2015 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-25539816

RESUMEN

The attenuated yellow fever (YF) vaccine (YF-17D) was developed in the 1930s, yet little is known about the protective mechanisms underlying its efficiency. In this study, we analyzed the relative contribution of cell-mediated and humoral immunity to the vaccine-induced protection in a murine model of YF-17D infection. Using different strains of knockout mice, we found that CD4(+) T cells, B cells, and Abs are required for full clinical protection of vaccinated mice, whereas CD8(+) T cells are dispensable for long-term survival after intracerebral challenge. However, by analyzing the immune response inside the infected CNS, we observed an accelerated T cell influx into the brain after intracerebral challenge of vaccinated mice, and this T cell recruitment correlated with improved virus control in the brain. Using mice deficient in B cells we found that, in the absence of Abs, YF vaccination can still induce some antiviral protection, and in vivo depletion of CD8(+) T cells from these animals revealed a pivotal role for CD8(+) T cells in controlling virus replication in the absence of a humoral response. Finally, we demonstrated that effector CD8(+) T cells also contribute to viral control in the presence of circulating YF-specific Abs. To our knowledge, this is the first time that YF-specific CD8(+) T cells have been demonstrated to possess antiviral activity in vivo.


Asunto(s)
Anticuerpos Antivirales/inmunología , Linfocitos T CD8-positivos/inmunología , Fiebre Amarilla/inmunología , Virus de la Fiebre Amarilla/inmunología , Inmunidad Adaptativa , Animales , Anticuerpos Neutralizantes/inmunología , Linfocitos B/inmunología , Encéfalo/inmunología , Encéfalo/metabolismo , Encéfalo/virología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Ligando de CD40/metabolismo , Linfocitos T CD8-positivos/metabolismo , Quimiotaxis de Leucocito/inmunología , Modelos Animales de Enfermedad , Femenino , Inmunización Pasiva , Depleción Linfocítica , Ratones , Ratones Noqueados , Vacunación , Replicación Viral , Fiebre Amarilla/genética , Fiebre Amarilla/mortalidad , Fiebre Amarilla/prevención & control , Vacuna contra la Fiebre Amarilla/inmunología
18.
Glia ; 64(3): 407-24, 2016 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-26496662

RESUMEN

Infiltration of myelin-specific T cells into the central nervous system induces the expression of proinflammatory cytokines in patients with multiple sclerosis (MS). We have previously shown that myelin-specific T cells are recruited into zones of axonal degeneration, where they stimulate lesion-reactive microglia. To gain mechanistic insight, we used RNA microarray analysis to compare the transcript profile in hippocampi from perforant pathway axonal-lesioned mice with and without adoptively transferred myelin-specific T cells 2 days postlesion, when microglia are clearly lesion reactive. Pathway analysis revealed that, among the 1,447 differently expressed transcripts, the interleukin (IL)-1 pathway including all IL-1 receptor ligands was upregulated in the presence of myelin-specific T cells. Quantitative polymerase chain reaction showed increased mRNA levels of IL-1ß, IL-1α, and IL-1 receptor antagonist in the T-cell-infiltrated hippocampi from axonal-lesioned mice. In situ hybridization and immunohistochemistry showed a T-cell-enhanced lesion-specific expression of IL-1ß mRNA and protein, respectively, and induction of the apoptosis-associated speck-like protein, ASC, in CD11b(+) cells. Double in situ hybridization showed colocalization of IL-1ß mRNA in a subset of CD11b mRNA(+) cells, of which many were part of cellular doublets or clusters, characteristic of proliferating, lesion-reactive microglia. Double-immunofluorescence showed a T-cell-enhanced colocalization of IL-1ß to CD11b(+) cells, including lesion-reactive CD11b(+) ramified microglia. These results suggest that myelin-specific T cells stimulate lesion-reactive microglial-like cells to produce IL-1ß. These findings are relevant to understand the consequences of T-cell infiltration in white and gray matter lesions in patients with MS.


Asunto(s)
Axones/metabolismo , Interleucina-1beta/metabolismo , Microglía/patología , Vaina de Mielina/patología , Enfermedades Neurodegenerativas/patología , Linfocitos T/fisiología , Traslado Adoptivo , Análisis de Varianza , Animales , Citocinas/genética , Citocinas/metabolismo , Giro Dentado/patología , Modelos Animales de Enfermedad , Femenino , Fluoresceínas/metabolismo , Interleucina-1beta/genética , Ratones , Análisis por Micromatrices , Infiltración Neutrófila , ARN Mensajero/metabolismo , Transducción de Señal/fisiología , Regulación hacia Arriba/genética
19.
J Virol ; 88(24): 14090-104, 2014 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-25253351

RESUMEN

UNLABELLED: Suppressors of cytokine signaling (SOCS) proteins are intracellular proteins that inhibit cytokine signaling in a variety of cell types. A number of viral infections have been associated with SOCS upregulation; however, not much is known about the mechanisms regulating SOCS expression during viral infection. In this study, we used two pathologically distinct intracerebral (i.c.) infection models to characterize temporal and spatial aspects of SOCS expression in the virus-infected central nervous system (CNS), and by employing various knockout mouse models, we sought to identify regulatory mechanisms that may underlie a virus induced upregulation of SOCS in the CNS. We found that i.c. infection with either lymphocytic choriomeningitis virus (LCMV) or yellow fever virus (YF) results in gradual upregulation of SOCS1/3 mRNA expression peaking at day 7 postinfection (p.i.). In the LCMV model, SOCS mRNA was expressed in brain resident cells, including astrocytes and some neurons, and for SOCS1 in particular this upregulation was almost entirely mediated by gamma interferon (IFN-γ) produced by infiltrating T cells. After infection with YF, we also found SOCS expression to be upregulated in brain resident cells with a peak on day 7 p.i., but in this model, the upregulation was only partially dependent on IFN-γ and T cells, indicating that at least one other mediator was involved in the upregulation of SOCS following YF infection. We conclude that virus-induced inflammation of the CNS is associated with upregulation of SOCS1/3 mRNA expression in brain resident cells and that at least two distinctive pathways can lead to this upregulation. IMPORTANCE: In the present report, we have studied the induction of SOCS1 and SOCS3 expression in the context of virus-induced CNS infection. We found that both a noncytolytic and a cytolytic virus induce marked upregulation of SOCS1 and -3 expression. Notably, the kinetics of the observed upregulation follows that of activity within proinflammatory signaling pathways and, interestingly, type II interferon (IFN), which is also a key inducer of inflammatory mediators, seems to be essential in initiating this counterinflammatory response. Another key observation is that not only cells of the immune system but also CNS resident cells are actively involved in both the pro- and the counterinflammatory immune circuits; thus, for example, astrocytes upregulate both C-X-C-motif chemokine 10 (CXCL10) and SOCS when exposed to type II IFN in vivo.


Asunto(s)
Infecciones por Arenaviridae/patología , Encefalitis Viral/patología , Infecciones por Flavivirus/patología , Coriomeningitis Linfocítica/inmunología , Virus de la Coriomeningitis Linfocítica/inmunología , Proteínas Supresoras de la Señalización de Citocinas/biosíntesis , Animales , Infecciones por Arenaviridae/inmunología , Infecciones por Arenaviridae/virología , Encefalitis Viral/inmunología , Encefalitis Viral/virología , Infecciones por Flavivirus/inmunología , Infecciones por Flavivirus/virología , Perfilación de la Expresión Génica , Interferón gamma/inmunología , Ratones Endogámicos C57BL , Ratones Noqueados , ARN Mensajero/análisis , Linfocitos T/inmunología , Factores de Tiempo , Regulación hacia Arriba
20.
Pediatr Neurol ; 158: 17-25, 2024 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-38936258

RESUMEN

BACKGROUND: Neurodevelopmental disorder with dysmorphic facies and distal limb anomalies (NEDDFL) is associated to BPTF gene haploinsufficiency. Epilepsy was not included in the initial descriptions of NEDDFL, but emerging evidence indicates that epileptic seizures occur in some affected individuals. This study aims to investigate the electroclinical epilepsy features in individuals with NEDDFL. METHODS: We enrolled individuals with BPTF-related seizures or interictal epileptiform discharges (IEDs) on electroencephalography (EEG). Demographic, clinical, genetic, raw EEG, and neuroimaging data as well as response to antiseizure medication were assessed. RESULTS: We studied 11 individuals with a null variant in BPTF, including five previously unpublished ones. Median age at last observation was 9 years (range: 4 to 43 years). Eight individuals had epilepsy, one had a single unprovoked seizure, and two showed IEDs only. Key features included (1) early childhood epilepsy onset (median 4 years, range: 10 months to 7 years), (2) well-organized EEG background (all cases) and brief bursts of spikes and slow waves (50% of individuals), and (3) developmental delay preceding seizure onset. Spectrum of epilepsy severity varied from drug-resistant epilepsy (27%) to isolated IEDs without seizures (18%). Levetiracetam was widely used and reduced seizure frequency in 67% of the cases. CONCLUSIONS: Our study provides the first characterization of BPTF-related epilepsy. Early-childhood-onset epilepsy occurs in 19% of subjects, all presenting with a well-organized EEG background associated with generalized interictal epileptiform abnormalities in half of these cases. Drug resistance is rare.


Asunto(s)
Electroencefalografía , Epilepsia , Fenotipo , Humanos , Niño , Masculino , Femenino , Preescolar , Epilepsia/fisiopatología , Epilepsia/tratamiento farmacológico , Epilepsia/genética , Adolescente , Adulto , Adulto Joven
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