Effect of a Klamath algae product ("AFA-B12") on blood levels of vitamin B12 and homocysteine in vegan subjects: a pilot study

none8Vitamin B12 is a critical nutrient that is often inadequate in a plant-based (vegan) diet, thus the inclusion of a reliable vitamin B12 source in a vegan diet is recommended as essential. Unfortunately, many natural sources of vitamin B12 have been proven to contain biologically inactive vitamin B12 analogues, inadequate for human supplementation. The aim of this non-randomized open trial was to determine whether supplementation with a natural Klamath algae-based product ("AFA-B12", Aphanizomenon flos-aquae algae plus a proprietary mix of enzymes) could favorably affect the vitamin B12 status of a group of 15 vegan subjects. By assessing blood concentration of vitamin B12, folate, and more importantly homocysteine (Hcy, a reliable marker in vegans of their B12 absorption), the vitamin B12 status of the participants at the end of the 3-month intervention period, while receiving the Klamath-algae supplement (T2), was compared with their vitamin B12 status at the end of the 3-month control period (T1), when they were not receiving any supplement, having stopped taking their usual vitamin B12 supplement at the beginning of the study (T0). Compared to the control period, in the intervention period participants improved their vitamin B12 status, significantly reducing Hcy blood concentration (p=0.003). In conclusion, the Klamath algae product AFA-B12 appears to be, in a preliminary study, an adequate and reliable source of vitamin B12 in humans.openL.Baroni; S.Scoglio; S.Benedetti; C.Bonetto; S.Pagliarani; Y.Benedetti; M.Rocchi; F.CanestrariL., Baroni; S., Scoglio; Benedetti, Serena; C., Bonetto; S., Pagliarani; Y., Benedetti; Rocchi, MARCO BRUNO LUIGI; Canestrari, Franc

Translational approach to address therapy in myotonia permanens due to a new SCN4A mutation

Objective: We performed a clinical, functional, and pharmacologic characterization of the novel p.P1158L Nav1.4 mutation identified in a young girl presenting a severe myotonic phenotype. Methods: Wild-type hNav1.4 channel and P1158L mutant were expressed in tsA201 cells for functional and pharmacologic studies using patch-clamp. Results: The patient shows pronounced myotonia, slowness of movements, and generalized muscle hypertrophy. Because of general discomfort with mexiletine, she was given flecainide with satisfactory response. In vitro, mutant channels show a slower current decay and a rightward shift of the voltage dependence of fast inactivation. The voltage dependence of activation and slow inactivation were not altered. Mutant channels were less sensitive to mexiletine, whereas sensitivity to flecainide was not altered. The reduced inhibition of mutant channels by mexiletine was also observed using clinically relevant drug concentrations in a myotonic-like condition. Conclusions: Clinical phenotype and functional alterations of P1158L support the diagnosis of myotonia permanens. Impairment of fast inactivation is consistent with the possible role of the channel domain III S4-S5 loop in the inactivation gate docking site. The reduced sensitivity of P1158L to mexiletine may have contributed to the unsatisfactory response of the patient. The success of flecainide therapy underscores the usefulness of in vitro functional studies to help in the choice of the best drug for each individual

Glycogen storage disease type III: A novel Agl knockout mouse model

AbstractGlycogen storage disease type III is an autosomal recessive disease characterized by a deficiency in the glycogen debranching enzyme, encoded by AGL. Essential features of this disease are hepatomegaly, hypoglycemia, hyperlipidemia, and growth retardation. Progressive skeletal myopathy, neuropathy, and/or cardiomyopathy become prominent in adults. Currently, there is no available cure. We generated an Agl knockout mouse model by deletion of the carboxy terminus of the protein, including the carboxy end of the glucosidase domain and the glycogen-binding domain. Agl knockout mice presented serious hepatomegaly, but we did not observe signs of cirrhosis or adenomas. In affected tissues, glycogen storage was higher than in wild-type mice, even in the central nervous system which has never been tested in GSDIII patients. The biochemical findings were in accordance with histological data, which clearly documented tissue impairment due to glycogen accumulation. Indeed, electron microscopy revealed the disruption of contractile units due to glycogen infiltrations. Furthermore, adult Agl knockout animals appeared less prompt to move, and they exhibited kyphosis. Three-mo-old Agl knockout mice could not run, and adult mice showed exercise intolerance. In addition, older affected animals exhibited an accelerated respiratory rate even at basal conditions. This observation was correlated with severe glycogen accumulation in the diaphragm. Diffuse glycogen deposition was observed in the tongues of affected mice. Our results demonstrate that this Agl knockout mouse is a reliable model for human glycogenosis type III, as it recapitulates the essential phenotypic features of the disease

MYH2 myopathy, a new case expands the clinical and pathological spectrum of the recessive form

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Case report: Clinical and molecular characterization of two siblings affected by Brody myopathy

Exercise-induced muscle stiffness is the hallmark of Brody disease, an autosomal recessive myopathy due to biallelic pathogenic variants in ATP2A1, encoding the sarcoplasmic/endoplasmic reticulum Ca2+ ATPase SERCA1. About 40 patients have been reported so far. Our knowledge about the natural history of this disorder, genotype–phenotype correlations and the effect of symptomatic treatment is partial. This results in incomplete recognition and underdiagnosis of the disease. Here, we report the clinical, instrumental, and molecular features of two siblings presenting childhood-onset exercise-induced muscle stiffness without pain. Both the probands display difficulty in climbing stairs and running, frequent falls, delayed muscle relaxation after exertion. Cold temperatures worsen these symptoms. No myotonic discharges were observed at electromyography. Whole Exome Sequencing analysis in the probands revealed the presence of two ATP2A1 variants: the previously reported frameshift microdeletion c.2464delC and the likely pathogenic novel splice-site variant c.324 + 1G > A, whose detrimental effect was demonstrated in ATP2A1 transcript analysis. The bi-allelic inheritance was verified by Sanger sequencing in the unaffected parents. This study expands the molecular defects associated with Brody myopathy

Case report: A novel ACTA1 variant in a patient with nemaline rods and increased glycogen deposition

BackgroundCongenital myopathies are a group of heterogeneous inherited disorders, mainly characterized by early-onset hypotonia and muscle weakness. The spectrum of clinical phenotype can be highly variable, going from very mild to severe presentations. The course also varies broadly resulting in a fatal outcome in the most severe cases but can either be benign or lead to an amelioration even in severe presentations. Muscle biopsy analysis is crucial for the identification of pathognomonic morphological features, such as core areas, nemaline bodies or rods, nuclear centralizations and congenital type 1 fibers disproportion. However, multiple abnormalities in the same muscle can be observed, making more complex the myopathological scenario.Case presentationHere, we describe an Italian newborn presenting with severe hypotonia, respiratory insufficiency, inability to suck and swallow, requiring mechanical ventilation and gastrostomy feeding. Muscle biopsy analyzed by light microscopy showed the presence of vacuoles filled with glycogen, suggesting a metabolic myopathy, but also fuchsinophilic inclusions. Ultrastructural studies confirmed the presence of normally structured glycogen, and the presence of minirods, directing the diagnostic hypothesis toward a nemaline myopathy. An expanded Next Generation Sequencing analysis targeting congenital myopathies genes revealed the presence of a novel heterozygous c.965 T > A p. (Leu322Gln) variant in the ACTA1 gene, which encodes the skeletal muscle alpha-actin.ConclusionOur case expands the repertoire of molecular and pathological features observed in actinopathies. We highlight the value of ultrastructural examination to investigate the abnormalities detected at the histological level. We also emphasized the use of expanded gene panels in the molecular analysis of neuromuscular patients, especially for those ones presenting multiple bioptic alterations

Translational approach to address therapy in myotonia permanens due to a new SCN4A

Objective: We performed a clinical, functional, and pharmacologic characterization of the novel p.P1158L Nav1.4 mutation identified in a young girl presenting a severe myotonic phenotype. Methods: Wild-type hNav1.4 channel and P1158L mutant were expressed in tsA201 cells for functional and pharmacologic studies using patch-clamp. Results: The patient shows pronounced myotonia, slowness of movements, and generalized muscle hypertrophy. Because of general discomfort with mexiletine, she was given flecainide with satisfactory response. In vitro, mutant channels show a slower current decay and a rightward shift of the voltage dependence of fast inactivation. The voltage dependence of activation and slow inactivation were not altered. Mutant channels were less sensitive to mexiletine, whereas sensitivity to flecainide was not altered. The reduced inhibition of mutant channels by mexiletine was also observed using clinically relevant drug concentrations in a myotonic-like condition. Conclusions: Clinical phenotype and functional alterations of P1158L support the diagnosis of myotonia permanens. Impairment of fast inactivation is consistent with the possible role of the channel domain III S4-S5 loop in the inactivation gate docking site. The reduced sensitivity of P1158L to mexiletine may have contributed to the unsatisfactory response of the patient. The success of flecainide therapy underscores the usefulness of in vitro functional studies to help in the choice of the best drug for each individual

Data_Sheet_1_Case report: Dihydropyridine receptor (CACNA1S) congenital myopathy, a novel phenotype with early onset periodic paralysis.docx

IntroductionCACNA1S related congenital myopathy is an emerging recently described entity. In this report we describe 2 sisters with mutations in the CACNA1S gene and the novel phenotype of congenital myopathy and infantile onset episodic weakness.Clinical descriptionBoth sisters had neonatal onset hypotonia, muscle weakness, and delayed walking. Episodic weakness started in infancy and continued thereafter, provoked mostly by cold exposure. Muscle imaging revealed fat replacement of gluteus maximus muscles. Next generation sequencing found the missense p.Cys944Tyr variant and the novel splicing variant c.3526-2A>G in CACNA1S. Minigene assay revealed the splicing variant caused skipping of exon 28 from the transcript, potentially affecting protein folding and/or voltage dependent activation.ConclusionThis novel phenotype supports the notion that there are age related differences in the clinical expression of CACNA1S gene mutations. This expands our understanding of mutations located in regions of the CACNA1S outside the highly conserved S4 segment, where most mutations thus far have been identified.</p