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This work aimed to study the effect of trehalose in protecting cells against Sod1 proteinopathy associated with amyotrophic lateral sclerosis (ALS). Humanized yeast cells in which native Sod1 was replaced by wild-type human Sod1 or an ALS mutant (WT-A4V Sod1 heterodimer) were used as the experimental model. Cells were treated with 10% trehalose (p/v) before or after the appearance of hSod1 proteinopathy induced by oxidative stress. In both conditions, trehalose reduced the number of cells with Sod1 inclusions, increased Sod1 activity, and decreased the levels of intracellular oxidation, demonstrating that trehalose avoids Sod1 misfolding and loss of function in response to oxidative stress. The survival rates of ALS Sod1 cells stressed in the presence of trehalose were 60% higher than in their absence. Treatment with trehalose after the appearance of Sod1 inclusions in cells expressing WT Sod1 doubled longevity; after 5 days, non-treated cells did not survive, but 15% of cells treated with sugar were still alive. Altogether, our results emphasize the potential of trehalose as a novel therapy, which might be applied preventively in ALS patients with a family history of the disease or after diagnosis in ALS patients who discover the disease following the first symptoms.

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Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease usually associated with severe weakness and death within 2-5 years. SOD1 mutations cause hereditary ALS in autosomal dominant and rarely in recessive pattern. We describe a new phenotype of slowly progressive fALS due to homozygous SOD1 mutations (c.358G > C, p.Val120Leu) in a Brazilian family. We reviewed the medical chart and interviewed the index patient and other relatives. A 41-year-old man developed weakness in his legs, leading to frequent falls, followed over the next few months with progressive arm fasciculations and muscle atrophy. The SOD1 enzymatic activity in erythrocytes was slightly decreased. A genetic test panel disclosed homozygous SOD1 mutations (c.358G > C, p.Val120Leu). His asymptomatic parents also carried one mutant allele and 2 brothers and a sister had died with ALS. We reported a new family with homozygous SOD1 mutation and slowly progressive ALS course. Further studies are necessary to confirm whether this mutation can also lead to disease in heterozygosis with incomplete penetrance.

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ß-Glucans (ßG) are polysaccharides widely distributed in nature with chemopreventive properties. The aim of this study was to investigate the effects of ßG and the combined treatment with doxorubicin (Dox) on cell viability and mRNA levels of genes involved in cell cycle, apoptosis and antioxidant response. ßG was not cytotoxic. The mRNA levels of CCNA2of cells exposed to ß-glucan was upregulated and the exposure to Dox decreased the expression, while the combination led to an upregulation. Modulation of mRNA levels of CASP9suggest that ßG could inhibit promotion and progression steps of carcinogenesis, eliminatingneoplastic cells. The upregulation of CCNA2gene in combined treatment could be occurred due to ability of ßG in restoring the cell cycle distribution pattern after treatment with Dox. The upregulation of SOD1suggests that ßG can enhance the intracellular antioxidant defense, reducing the levels of superoxide dismutase induced by Dox. This response could reduce oxidative damage and attenuate tissue damage during chemotherapeutic treatment. Our data suggest that the drug combination may be less effective in killing tumor cells than the treatment with Dox alone. Thus, future studies should carefully consider this effect on indication of ßG during chemotherapy.Keywords:caspase-9; cyclin A2; superoxide dismutase 1; cell cycle; antioxidant.Received on July 2, 2020.Accepted on February 7, 2022.IntroductionGlucans are polysaccharides widely distributed in nature and oftenstudied due to chemopreventive properties. They are constituent of the cell wall of plants (oats and barley), algae, bacteria and fungi. ß-glucans (ßG)have a common structure comprising a main chain of ß-(1,3) and/or ß-(1,4) D-glucopyranosyl unit and they differ in length and branching structures. ßG of Saccharomyces cerevisiaehave 1→6 side branches while those of bacteria have 1→4 side branches (Chan, Chan, & Sze, 2009). ßGcan prevent DNA damage induced by chemical and physical agents (Ghavami,Goliaei, Taghizadeh, & Nikoofar, 2014). Some authors showed its significant efficacy in preventing mutagenic effects caused by doxorubicin, cyclophosphamide and cisplatin (Tohamy, El-Ghor, El-Nahas, & Noshy, 2003), methyl methanesulfonate (Oliveira et al., 2007)and hydrogen peroxide (Slamenová, 2003). Moreover, some studies have related the antioxidant ability of ßGagainst reactive free radicals formed by endogenous metabolic processes or exogenous chemicals (Tsiapali et al., 2001; Slamenová,2003; Sener, Eksioglu-Demiralp, Cetiner, Ercan, & Yegen, 2006; Guerra Dore et al., 2007; Kofuji et al., 2012; Lei et al., 2015). Yeast-derived ßGhave modulating action of humoral and cellular immune responses (Vetvicka et al., 2007).This activity provides protection to the organism against infections and cancer development (Samuelsen, Schrezenmeir, & Knutsen, 2014; Roudbary, Daneshmand, Hajimorad, Roudbarmohammadip, & Hassan, 2015). Despite postulated modes of action by which ß-glucan works are lacking information about the molecular mechanisms involved in the chemopreventive activity of this polysaccharide. In addition, compounds with chemopreventive properties can contribute to reduce side effects and toxicity during the chemotherapeutic treatment. Therefore, the aim of this study was to investigate the effects of ßG and the combined treatment with doxorubicin (Dox) on the expression of genes related with apoptosis (CASP9), cell cycle control (CCNA2)and antioxidant defense (SOD1)in human breast cancer MCF-7 cells. Doxorubicin (Dox) was chosen because it is one of the most used chemotherapeutic agent for cancer treatment. The limitation on the use of Dox in cancer treatment is the lack of selectivity against cancer cells and, consequently, its toxicity to patients.(AU)

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PURPOSE: To identify inherited or acquired mutations in the VSX1, SOD1, TIMP3 and LOX genes from the combined analysis of corneal and blood samples from patients with Keratoconus. METHODS: The casuistry was consisted of samples of peripheral blood and corneal epithelium from 35 unrelated patients with Keratoconus who were submitted to corneal crosslink treatment. Also, blood and corneal epithelium samples from 89 non-keratoconic patients were used to compose the control group. Ophthalmologic evaluations included a clinical examination, topography and tomography. DNA samples were extracted from peripheral blood and from corneal epithelium in both groups and all coding regions of the VSX1, SOD1, TIMP3 and LOX genes were amplified by polymerase chain reaction, denatured and subjected to polyacrylamide gel electrophoresis. Mutational screening was performed by single-strand conformation polymorphism and direct DNA sequencing. RESULTS: No pathogenic variant was found in all coding regions of VSX1, SOD1, TIMP3 and LOX genes, we detected only few SNPs (single-nucleotide polymorphisms). Among the polymorphisms stand out three of them, corresponding to the synonymous exchange of amino acids: exon 3 of VSX1 Ala182Ala and exon 3 of TIMP3 His83His and Ser87Ser; in patients with Keratoconus and also in control subjects. All the polymorphisms were found in samples of corneal epithelium and corresponding blood. CONCLUSION: There is absence of KC pathogenic related to mutations in the VSX1, SOD1, TIMP3 and LOX genes in the studied patients.

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OBJECTIVE: To determine the presence of the 7-bp deletion c.169+50delTAAACAG in intron 2 of Superoxide Dismutase-1 gene in keratoconic patients from the State of São Paulo, Brazil, which promotes splicing variations, resulting in non-functional Superoxide Dismutase-1 antioxidant proteins, which may damage the corneal structure. RESULTS: A group of 35 keratoconic patients, from whom 35 peripheral blood samples and 58 samples of corneal fragments were evaluated, and a control group of 89 individuals, from whom 41 blood samples and 149 samples of corneal fragments were collected. After the amplification of DNA fragments by polymerase chain reaction, mutational screening analysis was performed by enzymatic digestion, followed by direct sequencing. The absence of the 7-bp c.169+50delTAAACAG mutation in intron 2 of Superoxide Dismutase-1 gene was detected in the analyzed subjects of the 2 groups, both in the cornea and peripheral blood samples. Then, according to our results, there is no involvement of c.169+50delTAAACAG deletion in the pathogenesis of keratoconus in this population, once it was not detected. But we emphasize that studies involving this deletion must be continued in an attempt to elucidate this issue.

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Amyotrophic Lateral Sclerosis (ALS) is a progressive, neurodegenerative disease characterized by loss of upper motor neurons (UMN) and lower motor neurons (LMN). Disease affects people all over the world and is more prevalent in men. Patients with ALS develop extensive muscle wasting, paralysis and ultimately death, with a median survival of usually fewer than five years after disease onset. ALS may be sporadic (sALS, 90%) or familial (fALS, 10%). The large majority of fALS cases are associated with genetic alterations, which are mainly related to the genes SOD1, TDP-43, FUS, and C9ORF72. In vitro and in vivo models have helped elucidate ALS etiology and pathogenesis, as well as its molecular, cellular, and physiological mechanisms. Many studies in cell cultures and animal models, such as Caenorhabditis elegans, Drosophila melanogaster, zebrafish, rodents, and non-human primates have been performed to clarify the relationship of these genes to ALS disease. However, there are inherent limitations to consider when using experimental models. In this review, we provide an updated overview of the most used in vitro and in vivo studies that have contributed to a better understanding of the different ALS pathogenic mechanisms.

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Objetivou-se descrever os achados clínicos, histopatológicos e moleculares associados à MDC em um cão da raça Pastor-Suiço. O cão possuía uma paraparesia progressiva em membros pélvicos e foi submetido a avaliações clínicas, pelas quais se obteve, entre outros diferenciais, o diagnóstico presuntivo de MDC. Com a evolução dos sinais, o tutor optou pela eutanásia. Os achados histopatológicos da medula espinhal foram compatíveis com uma degeneração segmentar axonal e mielínica. O diagnóstico molecular foi realizado por meio da extração do DNA obtido por swab oral. Uma PCR foi otimizada utilizando-se primers descritos em literatura para amplificar a região do gene SOD1. A amostra foi, então, submetida a sequenciamento unidirecional, que revelou que o animal em questão era homozigoto para o alelo A para a mutação c.118G>A no éxon 2 do gene SOD1. O diagnóstico clínico presuntivo da MDC no presente caso foi esclarecido por meio dos achados histopatológicos, associados aos achados clínicos, e da sua caracterização molecular. Ressalta-se a contribuição deste relato, que traz aspectos clínicos, histopatológicos e moleculares associados à MDC na raça Pastor-Suíço, para a qual, até o presente momento, na literatura consultada, não há relato dessa enfermidade.(AU)

The objective of this study was to describe the clinical, histopathological and molecular findings associated with MDC in a Swiss Shepherd dog. The dog had a progressive paraparesis in pelvic limbs and was submitted to clinical evaluations where, among other differentials, the presumptive diagnosis of MDC was obtained. With the progression of the nervous deficits tutor opted for euthanasia. The histopathological findings of the spinal cord were compatible with axonal and myelinic segmental degeneration. Molecular diagnosis was performed by extracting the DNA obtained by oral swab. PCR was optimized using primers described in the literature to amplify the SOD1 gene region. The sample was then subjected to one-way sequencing which revealed that the animal in question was homozygous for the A allele for the c.118G>A mutation in exon 2 of the SOD1 gene. The presumptive diagnosis of MDC in the present case was clarified by histopathological findings, as well as by its molecular characterization. The contribution of this report brings clinical, histopathological and molecular aspects associated with canine degenerative myelopathy in the Swiss Shepherd breed, that until this moment, in the literature consulted, there is no report of this disease in the breed mentioned.(AU)

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Objetivou-se descrever os achados clínicos, histopatológicos e moleculares associados à MDC em um cão da raça Pastor-Suiço. O cão possuía uma paraparesia progressiva em membros pélvicos e foi submetido a avaliações clínicas, pelas quais se obteve, entre outros diferenciais, o diagnóstico presuntivo de MDC. Com a evolução dos sinais, o tutor optou pela eutanásia. Os achados histopatológicos da medula espinhal foram compatíveis com uma degeneração segmentar axonal e mielínica. O diagnóstico molecular foi realizado por meio da extração do DNA obtido por swab oral. Uma PCR foi otimizada utilizando-se primers descritos em literatura para amplificar a região do gene SOD1. A amostra foi, então, submetida a sequenciamento unidirecional, que revelou que o animal em questão era homozigoto para o alelo A para a mutação c.118G>A no éxon 2 do gene SOD1. O diagnóstico clínico presuntivo da MDC no presente caso foi esclarecido por meio dos achados histopatológicos, associados aos achados clínicos, e da sua caracterização molecular. Ressalta-se a contribuição deste relato, que traz aspectos clínicos, histopatológicos e moleculares associados à MDC na raça Pastor-Suíço, para a qual, até o presente momento, na literatura consultada, não há relato dessa enfermidade.(AU)

The objective of this study was to describe the clinical, histopathological and molecular findings associated with MDC in a Swiss Shepherd dog. The dog had a progressive paraparesis in pelvic limbs and was submitted to clinical evaluations where, among other differentials, the presumptive diagnosis of MDC was obtained. With the progression of the nervous deficits tutor opted for euthanasia. The histopathological findings of the spinal cord were compatible with axonal and myelinic segmental degeneration. Molecular diagnosis was performed by extracting the DNA obtained by oral swab. PCR was optimized using primers described in the literature to amplify the SOD1 gene region. The sample was then subjected to one-way sequencing which revealed that the animal in question was homozygous for the A allele for the c.118G>A mutation in exon 2 of the SOD1 gene. The presumptive diagnosis of MDC in the present case was clarified by histopathological findings, as well as by its molecular characterization. The contribution of this report brings clinical, histopathological and molecular aspects associated with canine degenerative myelopathy in the Swiss Shepherd breed, that until this moment, in the literature consulted, there is no report of this disease in the breed mentioned.(AU)

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The superoxide dismutase type 1 (SOD1) gene is the first responsible gene mapped in amyotrophic lateral sclerosis type 1 (ALS1), and it codes for the enzyme SOD1, the function of which is to protect against damage mediated by free radicals deriving from oxygen. Its pathophysiological mechanism in ALS1 is related to ischemia. Several molecular studies of the SOD1 gene show that point mutations are the most frequent. The most common mutations in familial cases are p.A4V, p.I113Y, p.G37R, p.D90A and p.E100G, which account for more than 80% of cases, although intronic mutations have also been described as responsible for ALS1. Sporadic cases are explained by mutations in other genes such as SETX and C9orf72. ALS1 is a complex disease with genetic heterogeneity. On the other hand, familial and sporadic cases have a different etiology, which is explained by molecular heterogeneity and multiple pathogenic mechanisms that lead to ALS1; oxidative stress and ischemia are not the only cause. In Mexico, ALS molecular genetics studies are scarce. Clinical studies show an increase in cytokines such as adipsin in cerebrospinal fluid.

El gen SOD1 es el primer gen responsable mapeado en la esclerosis lateral amiotrófica tipo 1 (ELA1) y codifica para la enzima superóxido dismutasa tipo 1 (SOD1), cuya función es proteger del daño mediado de los radicales libres derivados del oxígeno; su mecanismo fisiopatológico en ELA1 se relaciona con isquemia. Diversos estudios moleculares del gen SOD1 muestran que las mutaciones puntuales son las más frecuentes. Las mutaciones más comunes en los casos familiares son p.A4V, p.I113Y, p.G37R, p.D90A y p.E100G, que explican más de 80 % de los casos, aunque también se han descrito mutaciones intrónicas como responsables de esclerosis lateral amiotrófica tipo 1. Los casos esporádicos se explican por mutaciones en otros genes como SETX y C9orf72. ELA1 es una enfermedad compleja con heterogeneidad genética. Por otra parte, los casos familiares y esporádicos tienen etiología distinta, lo cual se explica por la heterogeneidad molecular y múltiples mecanismos patogénicos que conducen a ELA1; el estrés oxidativo y la isquemia no son la única causa. En México son escasos los estudios de genética molecular de esclerosis lateral amiotrófica. Los estudios clínicos muestran incremento de citocinas como la adipsina en el líquido cefalorraquídeo.

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The superoxide dismutase type 1 (SOD1) gene is the first responsible gene mapped in amyotrophic lateral sclerosis type 1 (ALS1), and it codes for the enzyme SOD1, the function of which is to protect against damage mediated by free radicals deriving from oxygen. Its pathophysiological mechanism in ALS1 is related to ischemia. Several molecular studies of the SOD1 gene show that point mutations are the most frequent. The most common mutations in familial cases are p.A4V, p.I113Y, p.G37R, p.D90A and p.E100G, which account for more than 80% of cases, although intronic mutations have also been described as responsible for ALS1. Sporadic cases are explained by mutations in other genes such as SETX and C9orf72. ALS1 is a complex disease with genetic heterogeneity. On the other hand, familial and sporadic cases have a different etiology, which is explained by molecular heterogeneity and multiple pathogenic mechanisms that lead to ALS1; oxidative stress and ischemia are not the only cause. In Mexico, ALS molecular genetics studies are scarce. Clinical studies show an increase in cytokines such as adipsin in cerebrospinal fluid.

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