Directional coupling of oligodendrocyte connexin-47 and astrocyte connexin-43 gap junctions.

Intercellular communication via gap junction channels between oligodendrocytes and between astrocytes as well as between these cell types is essential to maintain the integrity of myelin in the central nervous system. Oligodendrocyte gap junction connexin-47 (Cx47) is a key element in this crosstalk and indeed, mutations in human Cx47 cause severe myelin disorders. However, the permeation properties of channels of Cx47 alone and in heterotypic combination with astrocyte Cx43 remain unclear. We show here that Cx47 contains three extra residues at 5' amino-terminus that play a critical role in the channel pore structure and account for relative low ionic conductivity, cationic permselectivity and voltage-gating properties of oligodendrocyte-oligodendrocyte Cx47 channels. Regarding oligodendrocyte-astrocyte coupling, heterotypic channels formed by Cx47 with Cx43 exhibit ionic and chemical rectification, which creates a directional diffusion barrier for the movement of ions and larger negatively charged molecules from cells expressing Cx47 to those with Cx43. The restrictive permeability of Cx47 channels and the diffusion barrier of Cx47-Cx43 channels was abolished by a mutation associated with leukodystrophy, the Cx47P90S, suggesting a novel pathogenic mechanism underlying myelin disorders that involves alterations in the panglial permeation.

Diagnóstico prenatal y diagnóstico preconcepcional. Perspectivas del análisis genético / Prenatal diagnosis and preconception diagnosis. Prospects for genetic analysis

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Miotonía congénita, una miopatía no distrófica / Myotonia congenita, a non-dystrophic myopathy

La miotonía congénita es la forma más común de miotonía no distrófica. Esta miopatía está causada por mutaciones en el gen CLCN1, codificante del principal canal de iones cloruro del músculo esquelético (ClC-1); la alteración de la función de este canal, regulado por voltaje, da lugar al fenómeno de miotonía. La enfermedad se puede heredar con un tipo de herencia dominante (enfermedad de Thomsen) o recesiva (enfermedad de Becker o miotonía congénita generalizada). El fenotipo clínico de ambas formas de la enfermedad es similar aunque la forma recesiva se caracteriza por una mayor gravedad de los síntomas. El diagnóstico clínico de miotonía congénita debe sospecharse cuando encontramos en un paciente episodios de rigidez muscular (miotonía), remisión o alivio de la rigidez con el ejercicio (fenómeno warm-up), miotonía clínica, un patrón electromiográfico característico y/o historia familiar. El diagnóstico molecular de miotonía congénita consiste en el análisis por secuenciación del gen CLCN1 (AU)

Myotonia congenita is the most common form of non-dystrophic myotonia. This myopathy is caused by mutations in the CLCN1 gene, encoding the main skeletal muscle chloride ion channel (ClC-1). Altering the function of this voltage-gated channel, leads to the phenomenon of myotonia. The disease can be inherited with a dominant (Thomsen disease) or recessive type (Becker disease or congenital generalised myotonia). The clinical phenotype of both forms of the disease is similar, although the recessive form is characterised by more severe symptoms. The clinical diagnosis of congenital myotonia should be suspected in a patient who presents with episodes of muscle stiffness (myotonia), remission or relief from stiffness with exercise (warm-up phenomenon), and a characteristic electromyography pattern, and/or family history. Sequencing the CLCN1 gene is the present approach for molecular diagnosis of myotonia congenita (AU)

Fast skeletal myofibers of mdx mouse, model of Duchenne muscular dystrophy, express connexin hemichannels that lead to apoptosis.

Skeletal muscles of patients with Duchenne muscular dystrophy (DMD) show numerous alterations including inflammation, apoptosis, and necrosis of myofibers. However, the molecular mechanism that explains these changes remains largely unknown. Here, the involvement of hemichannels formed by connexins (Cx HCs) was evaluated in skeletal muscle of mdx mouse model of DMD. Fast myofibers of mdx mice were found to express three connexins (39, 43 and 45) and high sarcolemma permeability, which was absent in myofibers of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice (deficient in skeletal muscle Cx43/Cx45 expression). These myofibers did not show elevated basal intracellular free Ca(2+) levels, immunoreactivity to phosphorylated p65 (active NF-κB), eNOS and annexin V/active Caspase 3 (marker of apoptosis) but presented dystrophin immunoreactivity. Moreover, muscles of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice exhibited partial decrease of necrotic features (big cells and high creatine kinase levels). Accordingly, these muscles showed similar macrophage infiltration as control mdx muscles. Nonetheless, the hanging test performance of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice was significantly better than that of control mdx Cx43(fl/fl)Cx45(fl/fl) mice. All three Cxs found in skeletal muscles of mdx mice were also detected in fast myofibers of biopsy specimens from patients with muscular dystrophy. Thus, reduction of Cx expression and/or function of Cx HCs may be potential therapeutic approaches to abrogate myofiber apoptosis in DMD.

Sobre el fraude científico / On scientific fraud

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Yo soy yo y mi microbiota (parafraseando a Ortega y Gasset) / I'm me and my microbiota (paraphrasing Ortega y Gasset)

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Regulation of connexin hemichannel activity by membrane potential and the extracellular calcium in health and disease.

Connexins are thought to solely mediate cell-to-cell communication by forming gap junction channels composed of two membrane-spanning hemichannels positioned end-to-end. However, many if not all connexin isoforms also form functional hemichannels (i.e., the precursors of complete channels) that mediate the rapid exchange of ions, second messengers and metabolites between the cell interior and the interstitial space. Electrical and molecular signaling via connexin hemichannels is now widely recognized to be important in many physiological scenarios and pathological conditions. Indeed, mutations in connexins that alter hemichannel function have been implicated in several diseases. Here, we present a comprehensive overview of how hemichannel activity is tightly regulated by membrane potential and the external calcium concentration. In addition, we discuss the genetic mutations known to alter hemichannel function and their deleterious effects, of which a better understanding is necessary to develop novel therapeutic approaches for diseases caused by hemichannel dysfunction. This article is part of the Special Issue Section entitled 'Current Pharmacology of Gap Junction Channels and Hemichannels'.

Interplay between DMD point mutations and splicing signals in Dystrophinopathy phenotypes.

DMD nonsense and frameshift mutations lead to severe Duchenne muscular dystrophy while in-frame mutations lead to milder Becker muscular dystrophy. Exceptions are found in 10% of cases and the production of alternatively spliced transcripts is considered a key modifier of disease severity. Several exonic mutations have been shown to induce exon-skipping, while splice site mutations result in exon-skipping or activation of cryptic splice sites. However, factors determining the splicing pathway are still unclear. Point mutations provide valuable information regarding the regulation of pre-mRNA splicing and elements defining exon identity in the DMD gene. Here we provide a comprehensive analysis of 98 point mutations related to clinical phenotype and their effect on muscle mRNA and dystrophin expression. Aberrant splicing was found in 27 mutations due to alteration of splice sites or splicing regulatory elements. Bioinformatics analysis was performed to test the ability of the available algorithms to predict consequences on mRNA and to investigate the major factors that determine the splicing pathway in mutations affecting splicing signals. Our findings suggest that the splicing pathway is highly dependent on the interplay between splice site strength and density of regulatory elements.

Screening for mutations in Spanish families with myotonia. Functional analysis of novel mutations in CLCN1 gene.

Myotonia congenita is an inherited muscle disorder caused by mutations in the CLCN1 gene, a voltage-gated chloride channel of skeletal muscle. We have studied 48 families with myotonia, 32 out of them carrying mutations in CLCN1 gene and eight carry mutations in SCN4A gene. We have found 26 different mutations in CLCN1 gene, including 13 not reported previously. Among those 26 mutations, c.180+3A>T in intron 1 is present in nearly one half of the Spanish families in this series, the largest one analyzed in Spain so far. Although scarce data have been published on the frequency of mutation c.180+3A>T in other populations, our data suggest that this mutation is more frequent in Spain than in other European populations. In addition, expression in HEK293 cells of the new missense mutants Tyr137Asp, Gly230Val, Gly233Val, Tyr302His, Gly416Glu, Arg421Cys, Asn567Lys and Gln788Pro, demonstrated that these DNA variants are disease-causing mutations that abrogate chloride currents.

Genetic basis of end-stage hypertrophic cardiomyopathy.

AIMS: Hypertrophic cardiomyopathy (HCM) is characterized by a heterogeneous presentation and clinical course. A minority of HCM patients develop end-stage HCM and require cardiac transplantation. The genetic basis of end-stage HCM is unknown but small series, isolated case reports and animal models have related the most aggressive heart failure course with the presence of multiple mutations. METHODS AND RESULTS: Twenty-six patients (age 40.4 ± 14.5 years; 46% male) transplanted for end-stage HCM underwent genetic screening of 10 HCM-related genes (MYH7, MYBPC3, TNNT2, TNNI3, TPM1, TNNC1, MYL3, MYL2, ACTC, LDB3). Additional genetic screening of LAMP2/PRKAG2 and mitochondrial DNA (mtDNA) was performed in four and three cases, respectively. Findings were correlated with clinical and histological features. Pathogenic mutations were identified in 15 patients (58%). Thirteen patients (50%) had mutations in sarcomeric genes (six in MYH7, three in MYBPC3, two in MYL2, one in TNNI3, and one in MYL3) and two patients had mutations in LAMP2. Only three patients (13%) had double mutations and all in homozygosis. Except for a more frequent family history of HCM, patients with mutations in sarcomeric genes did not show any clinical feature that distinguished them from patients without mutations in these genes. Evaluation of 44 relatives from 12 families identified 13 mutation carriers, 9 of whom had an overt HCM phenotype. CONCLUSION: Heart transplanted HCM has a heterogeneous genetic background where multiple mutations are uncommon. The clinical course of HCM is not primarily dependent on the presence of multiple sarcomeric mutations. Clinical and genetic evaluation of relatives does not support differential clinical management in HCM based on genetics.

Recomendaciones de buenas prácticas para el diagnóstico genético de la distrofia miotónica / Recommendations of good practices for the genetic diagnosis of myotonic dystrophy

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Moroccan consanguineous family with Becker myotonia and review.

Myotonia congenita is a genetic muscle disorder characterized by clinical and electrical myotonia, muscle hypertrophy, and stiffness. It is inherited as either autosomal-dominant or -recessive, known as Thomsen and Becker diseases, respectively. These diseases are distinguished by the severity of their symptoms and their patterns of inheritance. Becker disease usually appears later in childhood than Thomsen disease and causes more severe muscle stiffness and pain. Mutations in the muscular voltage-dependent chloride channel gene (CLCN1), located at 7q35, have been found in both types. We report here the case of a Moroccan consanguineous family with a myotonic autosomal-recessive condition in two children. The molecular studies showed that the patients reported here are homozygous for mutation p.Gly482Arg in the CLCN1 gene. The parents were heterozygote carriers for mutation p.Gly482Arg. This diagnosis allowed us to provide an appropriate management to the patients and to make a genetic counselling to their family.

[High-risk hypertrophic cardiomyopathy associated with a novel mutation in cardiac Myosin-binding protein C]. / Miocardiopatía hipertrófica de alto riesgo asociada con una nueva mutación en la proteína C fijadora de miosina.

Hypertrophic cardiomyopathy is an autosomal dominant inherited disease characterized by ventricular hypertrophy and myofibril disarray. Mutations responsible for hypertrophic cardiomyopathy have been identified in 11 genes that encode for cardiac sarcomere proteins. Traditionally, hypertrophic cardiomyopathy due to mutation of the myosin-binding protein C gene (MYBPC3) has been thought to follow a benign course. We report a family with several members affected by hypertrophic cardiomyopathy in which there was a high incidence of sudden death. Disease was presumably caused by the substitution of cytosine by guanine at nucleotide 269 of MYBPC3 mRNA. This mutation, which has not previously been described, modifies codon 79, which encodes for the incorporation of a tyrosine, and gives rise to a stop codon. The mutation described here appears to confer a higher risk than that previously associated with hypertrophic cardiomyopathy due to MYBPC3 gene mutation.

Miocardiopatía hipertrófica de alto riesgo asociada con una nueva mutación en la proteína C fijadora de miosina / High-risk hypertrophic cardiomyopathy associated with a novel mutation in cardiac Myosin-binding protein C

La miocardiopatía hipertrófica (MCH) es una enfermedad genética de herencia autosómica dominante carac-terizada por incremento en la masa ventricular y desor-ganización miofibrilar. Se han identificado mutaciones causantes de MCH en 11 genes de proteínas del sarcómero cardiaco. La MCH causada por mutaciones en el gen de la proteína C fijadora de miosina (MYBPC3) tradicionalmente se ha considerado de curso benigno. Presentamos a una familia con varios miembros afectados de MCH y alta incidencia de muerte súbita, presumiblemente como consecuencia de la sustitución de citosina por guanina en el nucleótido 269 del ARNm del gen MYBPC3. Esta mutación, no descrita previamente, modifica el codón 79, que codifica la incorporación del aminoácido tirosina y da lugar a un codón de terminación. La mutación descrita parece conferir un riesgo mayor que el atribuido hasta el momento a la MCH secundaria a mutaciones en el gen MYBPC3

Hypertrophic cardiomyopathy is an autosomal dominant inherited disease characterized by ventricular hypertrophy and myofibril disarray. Mutations responsible for hypertrophic cardiomyopathy have been identified in 11 genes that encode for cardiac sarcomere proteins. Traditionally, hypertrophic cardiomyopathy due to mutation of the myosin-binding protein C gene (MYBPC3) has been thought to follow a benign course. We report a family with several members affected by hypertrophic cardiomyopathy in which there was a high incidence of sudden death. Disease was presumably caused by the substitution of cytosine by guanine at nucleotide 269 of MYBPC3 mRNA. This mutation, which has not previously been described, modifies codon 79, which encodes for the incorporation of a tyrosine, and gives rise to a stop codon. The mutation described here appears to confer a higher risk than that previously associated with hypertrophic cardiomyopathy due to MYBPC3 gene mutation

[Hypertrophic cardiomyopathy: infrequent mutation of the cardiac beta-myosin heavy-chain gene]. / Miocardiopatía hipertrófica: baja frecuencia de mutaciones en el gen de la cadena pesada de la betamiosina cardiaca.

The aim of this study was to identify mutations in the cardiac heavy-chain beta-myosin gene (MYH7b) in a group of Spanish patients with hypertrophic cardiomyopathy. The study included 36 families with at least one member who had hypertrophic cardiomyopathy. DNA from exons 3 to 24 of the MYH7b gene was sequenced. Two mutations were identified: Arg858Cys and Met515Val. They occurred in two families, one of which was of Moroccan origin. This corresponds to a MYH7b gene mutation frequency of less than 5%. In contrast to findings in other Caucasian populations, MYH7b gene mutation occurred infrequently in this group of Spanish families with hypertrophic cardiomyopathy.