Selection of plasma donors for the production of anti-SARS-CoV-2 immunoglobulin-based therapies: Strategies for quantitative antibody measurements.

Even after two years of the pandemic, a completely effective treatment against SARS-CoV-2 has not yet been established. Considering this fact and the emergence of successive new viral variants, the development of therapies based on natural polyclonal antibodies recovered from convalescent plasma remains relevant. This study presents a comparison between different methods of screening antibodies in samples of 41 individuals previously diagnosed with COVID-19. We found a significant correlation between Abbot Architect anti-SARS-CoV-2 IgG and Abbott Allinity SARS-CoV-2 IgG II Quantitative assay intensity of reactivity and neutralizing antibody (nAb) titers. Thus, we propose an initial antibody screening with IgG anti-N Abbott Architect test, with an index of, for example, > 3.25 or SARS-CoV-2 IgG II Quantitative Abbott Allinity assay > 137.65 AU/mL as good predictors of Nab ≥ 1:80. For the quantitative method, this threshold demonstrated a 100 % sensitivity and 80 % specificity, with 97.3 % accuracy. An interesting observation was the increase in the neutralizing activity of the anti-SARS-CoV-2 antibodies with the longest interval between the end of the symptoms and the collection, demonstrating that the delay in plasma collection does not affect the achievement of adequate nAbs levels. These results demonstrate the possibility of using faster and more widely available commercial serological tests with a good correlation with viral neutralization tests in culture, allowing for optimized large-scale donor selection, which will be of utmost importance for the development of therapies such as hyperimmune immunoglobulin.

Low SARS-CoV-2 seroprevalence in a cohort of Brazilian sickle cell disease patients: Possible effects of emphasis on social isolation for a population initially considered to be at very high risk.

Despite being initially considered at higher risk for severe COVID-19, sickle cell disease (SCD) patients have mostly presented clinical severity similar to the general population. As their vulnerability to become infected remains uncertain, we assessed the seroreactivity for SARS-CoV-2 to estimate the prevalence of infection and possible phenotypic and socioeconomic determinants for their contagion. Serologic evaluation was performed on 135 patients with an overall prevalence of 11%; positivity was associated with older age and use of public transportation. We speculate that social distancing instructions recommended by our clinic may have contributed to lower levels of infection, but potential protection factors need further investigation.

Inflammatory Dendritic Cells Contribute to Regulate the Immune Response in Sickle Cell Disease.

Sickle cell disease (SCD), one of the most common hemoglobinopathies worldwide, is characterized by a chronic inflammatory component, with systemic release of inflammatory cytokines, due to hemolysis and vaso-occlusive processes. Patients with SCD demonstrate dysfunctional T and B lymphocyte responses, and they are more susceptible to infection. Although dendritic cells (DCs) are the main component responsible for activating and polarizing lymphocytic function, and are able to produce pro-inflammatory cytokines found in the serum of patients with SCD, minimal studies have thus far been devoted to these cells. In the present study, we identified the subpopulations of circulating DCs in patients with SCD, and found that the bloodstream of the patients showed higher numbers and percentages of DCs than that of healthy individuals. Among all the main DCs subsets, inflammatory DCs (CD14+ DCs) were responsible for this rise and correlated with higher reticulocyte count. The patients had more activated monocyte-derived DCs (mo-DCs), which produced MCP-1, IL-6, and IL-8 in culture. We found that a CD14+ mo-DC subset present in culture from some of the patients was the more activated subset and was mainly responsible for cytokine production, and this subset was also responsible for IL-17 production in co-culture with T lymphocytes. Finally, we suggest an involvement of heme oxygenase in the upregulation of CD14 in mo-DCs from the patients, indicating a potential mechanism for inducing inflammatory DC differentiation from circulating monocytes in the patients, which correlated with inflammatory cytokine production, T lymphocyte response skewing, and reticulocyte count.

SIVA, a target of p53, is downregulated in myelodysplastic syndromes

Background: SIVA is a transcriptional target of p53 that plays a potential role in the development and progression of cancer. In this study, we analyzed SIVA1 and SIVA2 expression, and its association with clinical features and TP53 and MDM2 expression in bone marrow cells from healthy donors and myelodysplastic syndrome (MDS) patients. Methods: Fifty-five untreated patients with MDS and 22 healthy donors were included. Gene expression was evaluated by quantitative PCR. For statistical analysis, Mann­Whitney test, Spearman correlation analysis and Log-rank (Mantel-Cox) were used, as appropriate. A p value <0.05 was considered statistically significant. Results: SIVA1 and SIVA2 transcripts were significantly decreased in bone marrow samples from MDS patients compared to healthy donors, and positively correlated with MDM2 and TP53 expression in MDS patients (all p < 0.05). MDM2 expression was also downregulated in bone marrow samples from MDS patients compared to healthy donors (p < 0.05). However, SIVA1, SIVA2, MDM2 and TP53 expressions did not impact on MDS outcomes. Conclusions: SIVA1 and SIVA2 transcripts are downregulated in bone marrow samples from MDS patients (AU)

Erythropoiesis-driven regulation of hepcidin in human red cell disorders is better reflected through concentrations of soluble transferrin receptor rather than growth differentiation factor 15.

Growth differentiation factor 15 (GDF-15) is a bone marrow-derived cytokine whose ability to suppress iron regulator hepcidin in vitro and increased concentrations found in patients with ineffective erythropoiesis (IE)suggest that hepcidin deficiency mediated by GDF-15 may be the pathophysiological explanation for nontransfusional iron overload. We aimed to compare GDF-15 production in anemic states with different types of erythropoietic dysfunction. Complete blood counts, biochemical markers of iron status, plasma hepcidin, GDF-15, and known hepcidin regulators [interleukin-6 and erythropoietin (EPO)] were measured in 87 patients with red cell disorders comprising IE and hemolytic states: thalassemia, sickle cell anemia, and cobalamin deficiency. Healthy volunteers were also evaluated for comparison. Neither overall increased EPO,nor variable GDF-15 concentrations correlated with circulating hepcidin concentrations (P = 0.265 and P = 0.872). Relative hepcidin deficiency was found in disorders presenting with concurrent elevation of GDF-15 and soluble transferrin receptor (sTfR), a biomarker of erythropoiesis, and sTfR had the strongest correlation with hepcidin (r(s) = 0.584, P < 0.0001). Our data show that high concentrations of GDF-15 in vivo are not necessarily associated with pathological hepcidin reduction, and hepcidin deficiency was only found when associated with sTfR overproduction. sTfR elevation may be a necessary common denominator of erythropoiesis-driven mechanisms to favor iron absorption in anemic states and appears a suitable target for investigative approaches to iron disorders.

ARHGAP21 modulates FAK activity and impairs glioblastoma cell migration.

Glioblastoma multiforme is highly aggressive and is the most common glial tumor type. Although there have been advances in treatment, the average survival expectancy is 12-15 months. Several genes have been shown to influence glioblastoma progression. In the present work, we demonstrate that the RhoGTPase Activating Protein 21 (ARHGAP21) is expressed in the nuclear and perinuclear regions of several cell lines. In T98G and U138MG, glioblastoma derived cell lines, ARHGAP21 interacts with the C-terminal region of Focal Adhesion Kinase (FAK). ARHGAP21 depletion by shRNAi in T98G cells alters cellular morphology and increases: FAK phosphorylation states and activation of downstream signaling; the activity state of Cdc42; the production of metalloproteinase 2 (MMP-2) and cell migration rates. These modifications were found to be mainly due to the loss of ARHGAP21 action on FAK and, consequently, the activation of downstream effectors. These results suggest not only that ARHGAP21 might act as a tumor suppressor gene, but also indicate that ARHGAP21 might be a master regulator of migration having a crucial role in controlling the progression of different tumor types.

Genotype frequencies at codon 129 of the prion protein gene in Brazil: Implications in susceptibility to variant Creutzfeldt-Jakob disease compared to European and Asian populations.

A polymorphism at codon 129 of the prion protein gene has been shown to confer genetic susceptibility to prion diseases, and to influence the epidemic course of variant Creutzfeldt-Jakob disease. We employed a PCR-endonuclease digestion-based assay to investigate this genetic trait in Brazil, and then compared our results to previously published data from several European and Asian countries.

Clinical, hematological, and molecular characterization of sickle cell anemia pediatric patients from two different cities in Brazil

O objetivo desse estudo foi avaliar aspectos clínicos, hematológicos e moleculares de pacientes pediátricos portadores de anemia falciforme em duas cidades brasileiras: Salvador e São Paulo. Foram estudados 71 pacientes com idades variando entre 3 a 18 anos, analisando-se os seguintes aspectos: perfis hematológicos, haplótipos dos genes da globina b, presença de talassemia a-23.7kb, número de internações por vaso-oclusão, infecção, presença de acidente vascular cerebral e litíase biliar. O genótipo Ben/CAR predominou nas duas cidades. Talassemia a-23.7kb teve freqüência de 28,2 por cento em Salvador e 22,5 por cento em São Paulo. Os pacientes de São Paulo apresentaram um número maior de internações por vaso-oclusão nos diferentes genótipos. Esses dados sugeriram um fenótipo com menor gravidade clínica nos pacientes de Salvador, possivelmente relacionados a fatores genéticos, ambientais e sócio-econômicos. Estudos adicionais necessitam ser realizados com intuito de elucidar os efeitos moduladores na expressão gênica da doença.

Clinical, hematological, and molecular characterization of sickle cell anemia pediatric patients from two different cities in Brazil.

This study focused on clinical, hematological, and molecular aspects of sickle cell anemia pediatric patients from two different cites in Brazil. Seventy-one patients from São Paulo and Salvador, aged 3 to 18 years, were evaluated. Hematological analyses, betaS globin gene haplotypes, and alpha2 3.7kb-thalassemia were performed. Numbers of hospitalizations due to vaso-occlusive crises, infections, stroke, and cholelithiasis were investigated. São Paulo had more hospitalizations from vaso-occlusion, cholelithiasis, and stroke than Salvador. The Ben/CAR genotype predominated in both cities. Alpha2 3.7kb-thalassemia had a frequency of 28.2% in Salvador, mostly with Ben/CAR genotype (45.0%), while São Paulo had 22.5% with similar frequencies of the Ben/ CAR and CAR/CAR genotypes. Sickle cell anemia patients from São Paulo also had more episodes of stroke, which was observed among CAR/CAR, atypical, and BEN/CAR haplotypes. In Salvador stroke was only observed in the Ben/CAR genotype. Cholelithiasis had similar frequencies in the two cities. These data suggest a milder phenotype among patients in Salvador, possibly due to genetic, environmental, and socioeconomic factors. Further studies are needed to elucidate modulating factors and phenotype association.

Expression of Sara2 human gene in erythroid progenitors.

A human homologue of Sar1, named Sara2, was shown to be preferentially expressed during erythropoiesis in a culture stimulated by EPO. Previous studies, in yeast, have shown that secretion-associated and Ras-related protein (Sar1p) plays an essential role in protein transport from the endoplasmic reticulum to the Golgi apparatus. Here, we report the molecular analysis of Sara2 in erythroid cell culture. A 1250 bp long cDNA, encoding a 198 amino-acid protein very similar to Sar1 proteins from other organisms, was obtained. Furthermore, we also report a functional study of Sara2 with Real-time quantitative PCR analysis, demonstrating that expression of Sara2 mRNA increases during the initial stages of erythroid differentiation with EPO and that a two-fold increase in expression occurs following the addition of hydroxyurea (HU). In K562 cells, Sara2 mRNA was observed to have a constant expression and the addition of HU also up-regulated the expression in these cells. Our results suggest that Sara2 is an important gene in processes involving proliferation and differentiation and could be valuable for understanding the vesicular transport system during erythropoiesis.

Acute myocardial infarction in sickle cell disease: a possible complication of hydroxyurea treatment.

We describe a 28-year-old man treated with hydroxyurea for sickle cell anemia, who was admitted to the University Hospital with an acute myocardial infarction. The patient had evolved high hematocrit values during his long-term hydroxyurea treatment, suggesting a correlation between a possible increment in blood viscosity and the coronary occlusion without previous lesions. Indeed, several studies associate vasocclusive episodes and severe clinical course with high viscosity. Although hydroxyurea is considered an effective therapeutic option for these patients, care should be taken to monitor hematocrit levels and possible complications. Hematocrit and hemoglobin values of above 30% and 10.5 g/dl in SS patients on hydroxyurea therapy should be avoided or closely monitored.

Rapid identification of the association of hemoglobin D Punjab and hemoglobin S (HbD Punjab/HbS) by the polymerase chain reaction

A hemoglobina D-Punjab foi descrita em vários grupos étnicos, tanto em heterozigose simples quanto em associaçäo com Hb S ou ß-talassemia. Neste trabalho descrevemos uma paciente brasileira, negra de 10 anos de idade que apresentava características clínicas de doença falciforme. O padräo eletroforético sugeriu a associaçäo Hb S/Hb D. A mutaçäo da Hb D foi confirmada através da digestäo do fragmento amplificado do gene da globina ß com a enzima EcoRI e sequencialmente direto do fragmento amplificado. A mutaçäo muda uma base no codon 121 e abole o sítio de reconhecimento normal da enzima, possibilitando o reconhecimento do gene anormal em electroforese de gel de agarose. Estes dados representam a segunda descriçäo comprovada da associaçäo Hb S/Hb D no Brasil e indicam que a presença da Hb D deve ser investigada, pelo método aqui descrito, em casos de doença falciforme com padräo eletroforético anômalo

Mielodisplasia: revisäo de 23 casos / Myelodysplasia: report of 23 cases

Foram analisados os dados clínicos hemogramas, mielograma e biópsia de medula óssea de 23 pacientes com diagnóstico de mielodisplasia. A queixa principal decorreu da anemia (21 casos) que usualmente apresentava-se associada ou näo àpancitopenia; macrocitose; hipogranulaçäo dos neutrófilos e pseudo-Pelger. O mielograma demonstrou diseritropoiese em todos os casos, hiperplasia eritrocítica e atipias megacariocitárias freqüentemente. A biópsia de medula óssea foi importante para o diagnóstico dessa síndrome, tendo-se observado celularidade aumentada em 12 casos, diminuida em 8 e normal em dois casos; focos de blastos (8 casos), megacariocitos atípicos (14 casos) e fibras de reticulina (aumentadas em 11 casos)

Sarcoma granulocítico mais mielodisplasia: relato de um caso / Granulocytic sarcoma with myelodysplasia: report of a case

Apresenta-se um caso de Sarcoma Granulocítico de comprometimento exclusivamente linfonodal, tendo, concomitante ao diagnóstico, uma Síndrome Mielodisplásica com excesso de blastos. Näo houve desenvolvimento de manifestaçöes periféricas ou medulares de leucemia desde o diagnóstico até o presente momento. Discutem-se os aspectos diagnósticos, a terapêutica introduzida e compara-se com os dados da literatura, relacionando-se as 2 entidades

Correlaçäo da citologia obtida pela técnica do "imprint" e histologia da medula por biópsia de agulha nos linfomas näo Hodgkin / Correlation of cytology obtained by \"imprint\" technique and histology of bone marrow by needle biopsy in non Hodgkin's lymphoma

Apresentam estudo de 46 casos de Linfomas näo-Hodgkin em adultos, sendo 27 homens e 19 mulheres, entre 18 e 76 anos (x = 50) com biópsia de medula óssea. Foi realizado histopatologia e citologia pela técnica do "imprint". Correlacionou-se os dados obtidos com os subtipos de linfomas no linfonodo ou tecido diagnóstico, segundo Rappaport (1966), com achados do sangue periférico. Os resultados revelaram grande predomínio de linfomas difusos (93,5%) sobre os nodulares (6,5%). A histopatologia de medula óssea mostrou positividade de 57,7%. O estudo citológico pela técnica do "imprint" revelou positividade de 39,5%. Quando o padräo de positividade da medula óssea foi intersticial ou difuso, a citologia também foi positiva, exceto um caso (14 em 15). Ao contrário, quando a positividade foi focal, apenas 2 em 11 puderam ser diagnosticados à citologia. No sangue periférico, os achados mais importantes foram linfopenia (33%), anemia (28%), plaquetopenia (11% - tidos com medula óssea positiva com padräo difuso ou intersticial) e achados de células linfomatosas (11%). Concluiu-se que a realizaçäo da biópsia de medula óssea é fundamental no estadiamento dos Linfomas näo-Hodgkin e que a técnica do "imprint" pode revelar a positividade nos casos de infiltraçäo intersticial e difusa, com rapidez e segurança. Além disso, foi notória a incidência de acometimento difuso da medula óssea ocorre nos linfomas histiocíticos e pobremente diferenciado, o que tornou os métodos histológicos e citológicos estatisticamente semelhantes. Tal fato näo ocorre nos linfomas mistos onde a biópsia é indispensável pela maior freqüência do acometimento focal