Association of Three Different Mutations in the CLCN1 Gene Modulating the Phenotype in a Consanguineous Family with Myotonia Congenita.

Myotonia congenita is a genetic disease caused by mutations in the CLCN1 gene, which encodes for the major chloride skeletal channel ClC-1, involved in the normal repolarization of muscle action potentials and consequent relaxation of the muscle after contraction. Two allelic forms are recognized, depending on the phenotype and the inheritance pattern: the autosomal dominant Thomsen disease with milder symptoms and the autosomal recessive Becker disorder with a severe phenotype. Before the recent advances of molecular testing, the diagnosis and genetic counseling of families was a challenge due to the large number of mutations in the CLCN1 gene, found both in homozygous or in heterozygous state. Here, we studied a consanguineous family in which three members presented a variable phenotype of myotonia, associated to a combination of three different mutations in the CLCN1 gene. A pathogenic splicing site mutation which causes the skipping of exon 17 was present in homozygosis in one very severely affected son. This mutation was present in compound heterozygosis in the consanguineous parents, but interestingly it was associated to a different second variant in the other allele: c.1453 A > G in the mother and c.1842 G > C in the father. Both displayed variable, but less severe phenotypes than their homozygous son. These results highlight the importance of analyzing the combination of different variants in the same gene in particular in families with patients displaying different phenotypes. This approach may improve the diagnosis, prognosis, and genetic counseling of the involved families.

The mdx Mutation in the 129/Sv Background Results in a Milder Phenotype: Transcriptome Comparative Analysis Searching for the Protective Factors.

The mdx mouse is a good genetic and molecular murine model for Duchenne Muscular Dystrophy (DMD), a progressive and devastating muscle disease. However, this model is inappropriate for testing new therapies due to its mild phenotype. Here, we transferred the mdx mutation to the 129/Sv strain with the aim to create a more severe model for DMD. Unexpectedly, functional analysis of the first three generations of mdx129 showed a progressive amelioration of the phenotype, associated to less connective tissue replacement, and more regeneration than the original mdxC57BL. Transcriptome comparative analysis was performed to identify what is protecting this new model from the dystrophic characteristics. The mdxC57BL presents three times more differentially expressed genes (DEGs) than the mdx129 (371 and 137 DEGs respectively). However, both models present more overexpressed genes than underexpressed, indicating that the dystrophic and regenerative alterations are associated with the activation rather than repression of genes. As to functional categories, the DEGs of both mdx models showed a predominance of immune system genes. Excluding this category, the mdx129 model showed a decreased participation of the endo/exocytic pathway and homeostasis categories, and an increased participation of the extracellular matrix and enzymatic activity categories. Spp1 gene overexpression was the most significant DEG exclusively expressed in the mdx129 strain. This was confirmed through relative mRNA analysis and osteopontin protein quantification. The amount of the 66 kDa band of the protein, representing the post-translational product of the gene, was about 4,8 times higher on western blotting. Spp1 is a known DMD prognostic biomarker, and our data indicate that its upregulation can benefit phenotype. Modeling the expression of the DEGs involved in the mdx mutation with a benign course should be tested as a possible therapeutic target for the dystrophic process.

Serum dosage of CPK-MB in dogs with ST deviation by chemiluminescence / A dosagem sérica de CPK-MB em cães com desvio ST por quimiluminescência

Although frequently in humans, hypoxic and ischemic heart diseases are poorly documented in dogs, with only few reports of acute myocardial infarction (AMI) in this species. Some electrocardiographic findings might suggest myocardium hypoxia/ischemia, like ST segment elevation or depression, but there are no studies showing whether deviations in ST segment are associated to myocardial injury and serum increase of creatine phosphokinase (CPK-MB). In order to investigate possible myocardial cells injury in poor perfusion conditions, 38 dogs were studied, 20 with normal electrocardiogram and 18 with ST segment elevation or depression, recorded in lead II, at a paper speed of 50 mm/sec and N sensibility (1mV=1cm). Serum measurement of creatine phosphokinase isoenzyme MB (CPK-MB) in normal dogs (group 1) determined control values (in ng/mL), which were compared to those obtained from dogs with deviation (group 2), which allowed confirmation or not of myocardial injury. CPK-MB mean values obtained from dogs in groups 1 and 2 were 0.540ng/ml (SD±0.890)ng/mL and 0.440ng/mL (SD±1.106), respectively. At a significance level of 5%, the relation of CPK-MB with age, mass and total creatine phosphokinase (CPK-T) was not significant in groups 1 and 2. CPK-MB showed no difference, at 5% level, between groups 1 and 2. In conclusion, it is possible to use the human chemiluminescent immunometric assay kit in canine species and that hypoxia/ischemia revealed by ST segment deviation does not mean significant myocardium injury.(AU)

Embora frequente em humanos, as doenças hipóxicas e isquêmicas do coração são pouco relatadas em cães, com poucos relatos de infarto agudo do miocárdio (IAM) nesta espécie. Alguns achados no eletrocardiograma podem sugerir hipóxia/isquemia miocárdica, como a elevação ou depressão do segmento ST, mas não há estudos que mostram se os desvios do segmento ST estão associados a lesões miocárdicas e aumento sérico da creatinafosfoquinase (CPK-MB). A fim de investigar possíveis lesões nas células miocárdicas em condições de má perfusão, 38 cães foram estudados, 20 com eletrocardiograma normal e 18 com elevação ou depressão do segmento ST, registrados em papel, na derivação II, velocidade de 50 mm/s e sensibilidade N (1mV = 1cm). A mensuração da creatinafosfoquinase isoenzima MB (CPK-MB) em cães normais (grupo 1) determinou os valores controle (em ng/ml), que foram comparados com os obtidos a partir de cães com desvio (grupo 2), permitindo a confirmação ou não da lesão miocárdica. Os valores médios de CPK-MB obtidos de cães nos grupos 1 e 2 foram 0,540ng/ml (DP±0,890) e 0,440ng / ml (DP ± 1.106), respectivamente. A um nível de significância de 5%, a relação de CPK-MB com a idade, massa e creatinofosfoquinase total (CPK-T) não foi significativa nos grupos 1 e 2. Não houve diferenças na CPK-MB, ao nível de 5%, entre os grupos 1 e 2. Conclui-se que é possível utilizar o kit de ensaio imunométrico por quimioluminescência humano na espécie canina e que a hipoxia/isquemia revelada pelos desvios do segmento ST, não significa lesão miocárdica.(AU)