Detecção de glicosilceramida em plasma de pacientes com doença de Gaucher

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Estudo da hidrólise enzimática de biomassa de microalga empregando uma sequência de enzimas

O emprego de microalgas para desenvolvimento de produtos está em pleno desenvolvimento considerando a composição de diversas espécies e a adaptação das mesmas ao meio de cultivo. Neste trabalho realizou-se o estudo da hidrólise enzimática da biomassa da microalga considerando a potencialidade de produção a partir de cepas de Chlorella sp., Scenedesmus sp., e Spirulina (Arthrospira platensis) empregando uma sequência de enzimas composta de duas amilases e dois complexos de celulases. Para tanto, foi realizado inicialmente a caracterização das espécies, seguindo-se o estudo com a Spirulina, que apresentou 40,02 % de carboidratos e mais que a metade do teor em amido (24,95 %) se mostrando promissor o seu uso para a otimização da hidrólise enzimática. Os melhores resultados de conversão foram encontrados com 12 FPU da enzima CTec2 enquanto que foi com 6,9 FPU da enzima HTec2, alcançando próximo a 100 % de rendimento. Desta forma pode-se concluir que a hidrólise empregando uma sequência de enzimas, alfa-amilase, glucoamilase e celulases mostrou-se eficiente para a microalga Spirulina, mostrando a viabilidade de aplicação da biomassa para o desenvolvimento de outros produtos, tornando a produção de matéria prima para etanol mais renovável

Genome of the Avirulent Human-Infective Trypanosome—Trypanosoma rangeli

Background: Trypanosoma rangeli is a hemoflagellate protozoan parasite infecting humans and other wild and domestic mammals across Central and South America. It does not cause human disease, but it can be mistaken for the etiologic agent of Chagas disease, Trypanosoma cruzi. We have sequenced the T. rangeli genome to provide new tools for elucidating the distinct and intriguing biology of this species and the key pathways related to interaction with its arthropod and mammalian hosts.  Methodology/Principal Findings: The T. rangeli haploid genome is ,24 Mb in length, and is the smallest and least repetitive trypanosomatid genome sequenced thus far. This parasite genome has shorter subtelomeric sequences compared to those of T. cruzi and T. brucei; displays intraspecific karyotype variability and lacks minichromosomes. Of the predicted 7,613 protein coding sequences, functional annotations could be determined for 2,415, while 5,043 are hypothetical proteins, some with evidence of protein expression. 7,101 genes (93%) are shared with other trypanosomatids that infect humans. An ortholog of the dcl2 gene involved in the T. brucei RNAi pathway was found in T. rangeli, but the RNAi machinery is non-functional since the other genes in this pathway are pseudogenized. T. rangeli is highly susceptible to oxidative stress, a phenotype that may be explained by a smaller number of anti-oxidant defense enzymes and heatshock proteins.  Conclusions/Significance: Phylogenetic comparison of nuclear and mitochondrial genes indicates that T. rangeli and T. cruzi are equidistant from T. brucei. In addition to revealing new aspects of trypanosome co-evolution within the vertebrate and invertebrate hosts, comparative genomic analysis with pathogenic trypanosomatids provides valuable new information that can be further explored with the aim of developing better diagnostic tools and/or therapeutic targets

The FANCM:p.Arg658* truncating variant is associated with risk of triple-negative breast cancer

Abstract: Breast cancer is a common disease partially caused by genetic risk factors. Germline pathogenic variants in DNA repair genes BRCA1, BRCA2, PALB2, ATM, and CHEK2 are associated with breast cancer risk. FANCM, which encodes for a DNA translocase, has been proposed as a breast cancer predisposition gene, with greater effects for the ER-negative and triple-negative breast cancer (TNBC) subtypes. We tested the three recurrent protein-truncating variants FANCM:p.Arg658*, p.Gln1701*, and p.Arg1931* for association with breast cancer risk in 67,112 cases, 53,766 controls, and 26,662 carriers of pathogenic variants of BRCA1 or BRCA2. These three variants were also studied functionally by measuring survival and chromosome fragility in FANCM−/− patient-derived immortalized fibroblasts treated with diepoxybutane or olaparib. We observed that FANCM:p.Arg658* was associated with increased risk of ER-negative disease and TNBC (OR = 2.44, P = 0.034 and OR = 3.79; P = 0.009, respectively). In a country-restricted analysis, we confirmed the associations detected for FANCM:p.Arg658* and found that also FANCM:p.Arg1931* was associated with ER-negative breast cancer risk (OR = 1.96; P = 0.006). The functional results indicated that all three variants were deleterious affecting cell survival and chromosome stability with FANCM:p.Arg658* causing more severe phenotypes. In conclusion, we confirmed that the two rare FANCM deleterious variants p.Arg658* and p.Arg1931* are risk factors for ER-negative and TNBC subtypes. Overall our data suggest that the effect of truncating variants on breast cancer risk may depend on their position in the gene. Cell sensitivity to olaparib exposure, identifies a possible therapeutic option to treat FANCM-associated tumors

Worldwide trends in hypertension prevalence and progress in treatment and control from 1990 to 2019: a pooled analysis of 1201 population-representative studies with 104 million participants.

BACKGROUND: Hypertension can be detected at the primary health-care level and low-cost treatments can effectively control hypertension. We aimed to measure the prevalence of hypertension and progress in its detection, treatment, and control from 1990 to 2019 for 200 countries and territories. METHODS: We used data from 1990 to 2019 on people aged 30-79 years from population-representative studies with measurement of blood pressure and data on blood pressure treatment. We defined hypertension as having systolic blood pressure 140 mm Hg or greater, diastolic blood pressure 90 mm Hg or greater, or taking medication for hypertension. We applied a Bayesian hierarchical model to estimate the prevalence of hypertension and the proportion of people with hypertension who had a previous diagnosis (detection), who were taking medication for hypertension (treatment), and whose hypertension was controlled to below 140/90 mm Hg (control). The model allowed for trends over time to be non-linear and to vary by age. FINDINGS: The number of people aged 30-79 years with hypertension doubled from 1990 to 2019, from 331 (95% credible interval 306-359) million women and 317 (292-344) million men in 1990 to 626 (584-668) million women and 652 (604-698) million men in 2019, despite stable global age-standardised prevalence. In 2019, age-standardised hypertension prevalence was lowest in Canada and Peru for both men and women; in Taiwan, South Korea, Japan, and some countries in western Europe including Switzerland, Spain, and the UK for women; and in several low-income and middle-income countries such as Eritrea, Bangladesh, Ethiopia, and Solomon Islands for men. Hypertension prevalence surpassed 50% for women in two countries and men in nine countries, in central and eastern Europe, central Asia, Oceania, and Latin America. Globally, 59% (55-62) of women and 49% (46-52) of men with hypertension reported a previous diagnosis of hypertension in 2019, and 47% (43-51) of women and 38% (35-41) of men were treated. Control rates among people with hypertension in 2019 were 23% (20-27) for women and 18% (16-21) for men. In 2019, treatment and control rates were highest in South Korea, Canada, and Iceland (treatment >70%; control >50%), followed by the USA, Costa Rica, Germany, Portugal, and Taiwan. Treatment rates were less than 25% for women and less than 20% for men in Nepal, Indonesia, and some countries in sub-Saharan Africa and Oceania. Control rates were below 10% for women and men in these countries and for men in some countries in north Africa, central and south Asia, and eastern Europe. Treatment and control rates have improved in most countries since 1990, but we found little change in most countries in sub-Saharan Africa and Oceania. Improvements were largest in high-income countries, central Europe, and some upper-middle-income and recently high-income countries including Costa Rica, Taiwan, Kazakhstan, South Africa, Brazil, Chile, Turkey, and Iran. INTERPRETATION: Improvements in the detection, treatment, and control of hypertension have varied substantially across countries, with some middle-income countries now outperforming most high-income nations. The dual approach of reducing hypertension prevalence through primary prevention and enhancing its treatment and control is achievable not only in high-income countries but also in low-income and middle-income settings. FUNDING: WHO

Worldwide trends in hypertension prevalence and progress in treatment and control from 1990 to 2019: a pooled analysis of 1201 population-representative studies with 104 million participants

Background Hypertension can be detected at the primary health-care level and low-cost treatments can effectively control hypertension. We aimed to measure the prevalence of hypertension and progress in its detection, treatment, and control from 1990 to 2019 for 200 countries and territories. Methods We used data from 1990 to 2019 on people aged 30-79 years from population-representative studies with measurement of blood pressure and data on blood pressure treatment. We defined hypertension as having systolic blood pressure 140 mm Hg or greater, diastolic blood pressure 90 mm Hg or greater, or taking medication for hypertension. We applied a Bayesian hierarchical model to estimate the prevalence of hypertension and the proportion of people with hypertension who had a previous diagnosis (detection), who were taking medication for hypertension (treatment), and whose hypertension was controlled to below 140/90 mm Hg (control). The model allowed for trends over time to be non-linear and to vary by age. Findings The number of people aged 30-79 years with hypertension doubled from 1990 to 2019, from 331 (95% credible interval 306-359) million women and 317 (292-344) million men in 1990 to 626 (584-668) million women and 652 (604-698) million men in 2019, despite stable global age-standardised prevalence. In 2019, age-standardised hypertension prevalence was lowest in Canada and Peru for both men and women; in Taiwan, South Korea, Japan, and some countries in western Europe including Switzerland, Spain, and the UK for women; and in several low-income and middle-income countries such as Eritrea, Bangladesh, Ethiopia, and Solomon Islands for men. Hypertension prevalence surpassed 50% for women in two countries and men in nine countries, in central and eastern Europe, central Asia, Oceania, and Latin America. Globally, 59% (55-62) of women and 49% (46-52) of men with hypertension reported a previous diagnosis of hypertension in 2019, and 47% (43-51) of women and 38% (35-41) of men were treated. Control rates among people with hypertension in 2019 were 23% (20-27) for women and 18% (16-21) for men. In 2019, treatment and control rates were highest in South Korea, Canada, and Iceland (treatment >70%; control >50%), followed by the USA, Costa Rica, Germany, Portugal, and Taiwan. Treatment rates were less than 25% for women and less than 20% for men in Nepal, Indonesia, and some countries in sub-Saharan Africa and Oceania. Control rates were below 10% for women and men in these countries and for men in some countries in north Africa, central and south Asia, and eastern Europe. Treatment and control rates have improved in most countries since 1990, but we found little change in most countries in sub-Saharan Africa and Oceania. Improvements were largest in high-income countries, central Europe, and some upper-middle-income and recently high-income countries including Costa Rica, Taiwan, Kazakhstan, South Africa, Brazil, Chile, Turkey, and Iran. Interpretation Improvements in the detection, treatment, and control of hypertension have varied substantially across countries, with some middle-income countries now outperforming most high-income nations. The dual approach of reducing hypertension prevalence through primary prevention and enhancing its treatment and control is achievable not only in high-income countries but also in low-income and middle-income settings. Copyright (C) 2021 World Health Organization; licensee Elsevier

Worldwide trends in hypertension prevalence and progress in treatment and control from 1990 to 2019: a pooled analysis of 1201 population-representative studies with 104 million participants

Background Hypertension can be detected at the primary health-care level and low-cost treatments can effectively control hypertension. We aimed to measure the prevalence of hypertension and progress in its detection, treatment, and control from 1990 to 2019 for 200 countries and territories. Methods We used data from 1990 to 2019 on people aged 30–79 years from population-representative studies with measurement of blood pressure and data on blood pressure treatment. We defined hypertension as having systolic blood pressure 140 mm Hg or greater, diastolic blood pressure 90 mm Hg or greater, or taking medication for hypertension. We applied a Bayesian hierarchical model to estimate the prevalence of hypertension and the proportion of people with hypertension who had a previous diagnosis (detection), who were taking medication for hypertension (treatment), and whose hypertension was controlled to below 140/90 mm Hg (control). The model allowed for trends over time to be non-linear and to vary by age. Findings The number of people aged 30–79 years with hypertension doubled from 1990 to 2019, from 331 (95% credible interval 306–359) million women and 317 (292–344) million men in 1990 to 626 (584–668) million women and 652 (604–698) million men in 2019, despite stable global age-standardised prevalence. In 2019, age-standardised hypertension prevalence was lowest in Canada and Peru for both men and women; in Taiwan, South Korea, Japan, and some countries in western Europe including Switzerland, Spain, and the UK for women; and in several low-income and middle-income countries such as Eritrea, Bangladesh, Ethiopia, and Solomon Islands for men. Hypertension prevalence surpassed 50% for women in two countries and men in nine countries, in central and eastern Europe, central Asia, Oceania, and Latin America. Globally, 59% (55–62) of women and 49% (46–52) of men with hypertension reported a previous diagnosis of hypertension in 2019, and 47% (43–51) of women and 38% (35–41) of men were treated. Control rates among people with hypertension in 2019 were 23% (20–27) for women and 18% (16–21) for men. In 2019, treatment and control rates were highest in South Korea, Canada, and Iceland (treatment >70%; control >50%), followed by the USA, Costa Rica, Germany, Portugal, and Taiwan. Treatment rates were less than 25% for women and less than 20% for men in Nepal, Indonesia, and some countries in sub-Saharan Africa and Oceania. Control rates were below 10% for women and men in these countries and for men in some countries in north Africa, central and south Asia, and eastern Europe. Treatment and control rates have improved in most countries since 1990, but we found little change in most countries in sub-Saharan Africa and Oceania. Improvements were largest in high-income countries, central Europe, and some upper-middle-income and recently high-income countries including Costa Rica, Taiwan, Kazakhstan, South Africa, Brazil, Chile, Turkey, and Iran. Interpretation Improvements in the detection, treatment, and control of hypertension have varied substantially across countries, with some middle-income countries now outperforming most high-income nations. The dual approach of reducing hypertension prevalence through primary prevention and enhancing its treatment and control is achievable not only in high-income countries but also in low-income and middle-income settings

A case-only study to identify genetic modifiers of breast cancer risk for BRCA1/BRCA2 mutation carriers

Abstract: Breast cancer (BC) risk for BRCA1 and BRCA2 mutation carriers varies by genetic and familial factors. About 50 common variants have been shown to modify BC risk for mutation carriers. All but three, were identified in general population studies. Other mutation carrier-specific susceptibility variants may exist but studies of mutation carriers have so far been underpowered. We conduct a novel case-only genome-wide association study comparing genotype frequencies between 60,212 general population BC cases and 13,007 cases with BRCA1 or BRCA2 mutations. We identify robust novel associations for 2 variants with BC for BRCA1 and 3 for BRCA2 mutation carriers, P < 10−8, at 5 loci, which are not associated with risk in the general population. They include rs60882887 at 11p11.2 where MADD, SP11 and EIF1, genes previously implicated in BC biology, are predicted as potential targets. These findings will contribute towards customising BC polygenic risk scores for BRCA1 and BRCA2 mutation carriers

A case-only study to identify genetic modifiers of breast cancer risk for BRCA1/BRCA2 mutation carriers

Breast cancer (BC) risk for BRCA1 and BRCA2 mutation carriers varies by genetic and familial factors. About 50 common variants have been shown to modify BC risk for mutation carriers. All but three, were identified in general population studies. Other mutation carrier-specific susceptibility variants may exist but studies of mutation carriers have so far been underpowered. We conduct a novel case-only genome-wide association study comparing genotype frequencies between 60,212 general population BC cases and 13,007 cases with BRCA1 or BRCA2 mutations. We identify robust novel associations for 2 variants with BC for BRCA1 and 3 for BRCA2 mutation carriers, P < 10−8, at 5 loci, which are not associated with risk in the general population. They include rs60882887 at 11p11.2 where MADD, SP11 and EIF1, genes previously implicated in BC biology, are predicted as potential targets. These findings will contribute towards customising BC polygenic risk scores for BRCA1 and BRCA2 mutation carriers

Quantificação de glicosilceramida em plasma de pacientes com a doença de Gaucher

A Doença de Gaucher (DG) é uma esfingolipidose causada por mutações no gene da -glicosidase, ocasionando depósito lisossomal de glicosilceramida (GliCer), principalmente, nas células do sistema mononuclear fagocitário. O diagnóstico da DG é, normalmente, confirmado pela determinação enzimática e/ou pela caracterização molecular. Neste trabalho nos propusemos a: (1) confirmar o diagnóstico de uma amostra de indivíduos com DG através da medida da atividade da -glicosidase e da quitotriosidase (QT); (2) extrair, purificar e quantificar a GliCer de plasma de pacientes com DG com e sem tratamento por reposição enzimática (TRE) e de indivíduos normais, comparando o conteúdo deste lipídio entre os grupos mencionados e (3) comparar a concentração plasmática da quemoquina CCL18/PARC entre os 3 grupos acima. Os leucócitos e o plasma de cada indivíduo foram separados por centrifugação. A atividade da -glicosidase em leucócitos dos pacientes com DG foi 5% daquela de indivíduos normais e a quitotriosidase estava 500 vezes aumentada nos pacientes com DG. O plasma de cada indivíduo foi tratado, seqüencialmente, com três sistemas de solventes: clorofórmio (C): metanol (M) em três proporções distintas. Os três extratos lipídicos totais de cada amostra foram reunidos (extrato lipídico total) e evaporados a seco (resíduo). Este resíduo de extrato lipídico total foi submetido, seqüencialmente, a uma coluna de ácido silícico, à metanólise alcalina e a uma coluna Sep-Pack. Desta coluna resultaram os eluatos C:M:água (3:48:47) e (60:30:4,5). Após ajustes necessários obtivemos a purificação de glicosilceramida numa única fração. A análise destes eluatos por cromatografia em camada delgada de alta resolução (HPTLC) e com revelação química (CuSO4/H3PO4) mostrou que ambos continham uma banda com velocidade de migração semelhante a do padrão de GliCer. A confirmação da identidade desta banda foi realizada por imunorevelação usando anticorpo primário anti-GliCer e secundário conjugado a peroxidase. Pacientes com DG sem tratamento por TRE apresentaram, aproximadamente, 20 vezes mais GliCer que indivíduos normais e 11 vezes mais que pacientes com tratamento. Com a dosagem plasmática da quemoquina CCL18/PARC, observamos que pacientes com DG possuíam uma concentração da mesma 28 vezes aumentada em relação aos indivíduos normais e 17 vezes mais elevada que a de pacientes com tratamento. Ou seja, podemos detectar uma redução de 90% da concentração de GliCer e de 40% na concentração de CCL18/PARC nos pacientes com DG com TRE em comparação com as respectivas concentrações nos paciente com DG sem tratamento. A metodologia estabelecida neste trabalho permitiu extrair, purificar, separar, detectar e quantificar GliCer no plasma de pacientes com DG com e sem tratamento por TRE, bem como, GliCer de indivíduos normais. Tanto a quantificação da GliCer quanto a dosagem de CCL18/PARC permitiram identificar, através dos respectivos valores obtidos de plasma, os grupos de indivíduos analisados. Portanto, estas duas avaliações podem ser associadas à determinação da atividade da quitotriosidase como biomarcadores da DG.Gaucher’s disease (GD) is a sphingolipidosis caused by mutations in the β-glucosidase gene, causing lysosomal storage of glucosylceramide (GluCer) mainly in the mononuclear phagocyte system cells. As a rule, GD diagnosis is confirmed by enzyme determination and/or molecular characterization. The objectives of the present study were (1) to confirm GD diagnosis in a sample of subjects by measuring β-glucosidase activity and chitotriosidase activity, (2) to extract, purify and quantify GluCer in the plasma of patients with GD with and without enzyme replacement therapy (ERT) and of healthy individuals, comparing the content of the lipid between the groups, and (3) the compare the plasma concentration of chemokine CCL18/PARC between the three groups. Plasma of each individual was centrifuged to obtain leukocytes. β-glucosidase activity in leukocytes of patients with GD represented a 5% share of that of healthy individuals, while chitotriosidase activity was 500 times higher in GD patients. Plasma was treated sequentially with a chloroform:methanol solvent system as three sequential solution ratios. The three total lipid extracts of each sample were pooled (total lipid extract) and evaporated to dryness to obtain a residue. This residue was sequentially treated in a salicylic acid column and a Sep-Pak™ column for alkaline methanolysis, from which two chloroform:methanol:water eluates were obtained (3:48:47 and 60:30:4.5). After the required adjustments, GluCer was purified as a single fraction. High performance thin-layer chromatography (HPTLC) and chemical development using CuSO4 and H3PO4 showed that both eluates had a band whose migration speed was similar to the GluCer standard. Confirmation of this band’s identity was conducted by immune development using anti-GluCer primary antibody and peroxidase-conjugated secondary antibody. GluCer levels in DG patients without ERT treatment were approximately 20 times higher than in healthy individuals and 11 times higher than in patients under treatment. Plasma chemokine CCL18/PARC levels in DG patients without treatment were 28 and 17 times higher as compared to healthy and individuals DG patients under treatment, respectively. This accounts for a 90% decrease in GluCer levels and a 40% decrease in CCL18/PARC levels in GD patients with ERT in comparison to DG patients without treatment. The methodology described in this paper afforded to quantify GluCer in plasma of DG individuals with and without ERT as well as GluCer levels in healthy individuals. Quantification of GluCer and CCL18/PARC levels in plasma afforded to identify the groups of individuals analyzed. Therefore, these two parameters may be associated to the determination of chitotriosidase activity as GD biomarkers