Produção de citocinas e óxido nítrico por macrófagos de camundongos infectados com flavivírus brasileiros

The Flaviviridae family, Flavivirus genus includes viruses that are transmitted to vertebrates by infected mosquitoes or ticks. The genus Flavivirus includes a variety of viruses that cause diseases such as acute febrile illness, encephalitis, and hemorrhagic fever. Flaviviruses primarily infect blood monocytes and tissue macrophages, which have been shown to be permissive, supporting viral replication and serving as virus reservoirs. On the other hand, these cells may have an important antiviral activity related to modulation by cytokine production and by the capacity of these cells to synthesize reactive free radicals such as nitric oxide (NO) which can have a microbicidal effect. The present study was performed in order to determine the production of cytokines interleukin-1beta (IL-1β), tumor necrosis factor -alpha (TNF-α), transforming growth factor- beta (TGF-β) and interferon -alpha (IFN-α) and NO by macrophages infected with one of four Brazilian flaviviruses, Bussuquara virus (BUSV), Yellow Fever virus (YFV), Rocio virus (ROCV) and Encephalitis Saint Louis virus (SLEV), and to verify the possible antiviral effect of NO during macrophage infection with ROCV. Moreover, we asked if the different viruses were able to regulate bacterial lipopolysaccharide (LPS) induced cytokine production. Our results showed that YFV and SLEV reduced the production of IL-1β and TGF-β by LPS-stimulated macrophages, while ROCV only diminished LPS-stimulated TGF-β synthesis. On the other hand, BUSV more likely favored an enhancement of the LPS-induced production of IL-1β by macrophages. Additionally, while most of the viruses stimulated the production of IFN-α, none of them altered the production of TNF-α by murine macrophages. Interestingly, all viruses induced synthesis of NO that was not correlated with antiviral activity for ROCV.A família Flaviviridae, gênero flavivírus inclui vírus que são transmitidos para os vertebrados por mosquitos e carrapatos. O gênero flavivirus inclui uma variedade de vírus que causa doenças como febres, encefalites e febres hemorrágicas. Primeiramente, as flaviviroses infectam monócitos do sangue e macrófagos do tecido, o qual tem mostrado ser permissivo, suportando a replicação viral e servindo como reservatório de vírus. Por outro lado, essas células podem ter uma importante atividade antiviral relacionada à modulação pela produção de citocinas e pela capacidade destas células sintetizar reativos de radicais livres como óxido nítrico (NO) o qual tem efeito microbicida. O presente estudo foi realizado a fim de determinar a produção de citocinas interleucina -1 beta (IL-1β), fator de necrose tumoral-alfa (TNF-α), fator de crescimento transformador-beta (TGF-β), interferon - alfa (IFN-α) e NO pelos macrófagos infectados com os quatros flavivírus como vírus Bussuquara (BUSV), vírus da febre amarela (YFV), vírus Rocio (ROCV) e vírus da Encefalite de Saint Louis (SLEV), e verificar o possível efeito antiviral de NO durante a infecção dos macrófagos com ROCV. Além disso, com os diferentes vírus foram capazes de regular o lipopolissacarídeo bacteriano (LPS) indutor da produção de citocinas. Nossos resultados mostraram que YFV e SLEV reduziram a produção de IL-1β e TGF-β quando macrófagos foram estimulados pelo LPS, enquanto ROCV somente diminuiu a síntese de TGF-β estimulada pelo LPS. Entretanto, BUSV favoreceu uma acentuada produção de IL-1β pelos macrófagos estimulados pelo LPS, enquanto os vírus estimularam a produção de IFN-α, nenhum deles alterou a produção de TNF-α pelos macrófagos murinos. Interessantemente, todos os vírus induziram a síntese de NO que não esteve correlacionada com a atividade antiviral pelo ROCV

Intervenção educacional sobre enteroparasitoses: um estudo quase experimental

Introdução: As enteroparasitoses vêm ocasionando sérios problemas de saúde pública no mundo, sobretudo nos países em desenvolvimento. Mesmo com a população em geral relatando que tem conhecimento sobre as parasitoses intestinais, estudos apontam que não sabem identificar as verminoses. Objetivo: Elaborar, implementar e avaliar um programa educativo sobre parasitoses intestinais em uma escola pública de Ribeirão Preto – SP. Materiais e Métodos: Estudo quase experimental, não randomizado realizado com 56 alunos do 1º a 4º ano do ensino fundamental. Para avaliar o conhecimento dos alunos antes e após intervenção educativa, foi aplicado um questionário semiestruturado (Pré e Pós - teste). Resultados: Houve mudanças nas respostas dos alunos em relação hábitos de higiene, especialmente nas questões sobre como os vermes se alimentam (p=0,008); o que não fazer para não se contaminar com vermes (p=0,05); qual o formato dos vermes quando é ingerido (p=0,001); quais órgãos os vermes atravessam (p=0,001). Assim, ficou evidente que este tipo de atividade educativa foi eficiente para auxiliar na aprendizagem de alunos do ensino fundamental. Discussão: Após a atividade educativa sem notou um crescimento nos acertos para a maioria das questões, contudo ainda houve dificuldades de compreensão sobre os sintomas, o ciclo e quais organismos são vermes. Conclusões: Espera-se que este trabalho incentive profissionais da saúde e da educação a incluir práticas educativas sobre saúde no contexto escolar.Como citar este artigo: Bragagnollo GR, Godoy PCGT, Santos TS, Ribeiro VS, Morero JAP, Ferreira BR. Intervenção educacional sobre enteroparasitoses: um estudo quase experimental. Rev Cuid. 2018; 9(1): 2030-44. http://dx.doi.org/10.15649/cuidarte.v9i1.486

Integral use of immunopeptidomics and immunoinformatics for the characterization of antigen presentation and rational identification of BoLA-DR- presented peptides and epitopes

MHC peptide binding and presentation is the most selective event defining the landscape of T cell epitopes. Consequently, understanding the diversity of MHC alleles in a given population and the parameters that define the set of ligands that can be bound and presented by each of these alleles (the immunopeptidome) has an enormous impact on our capacity to predict and manipulate the potential of protein Ags to elicit functional T cell responses. Liquid chromatography–mass spectrometry analysis of MHC-eluted ligand data has proven to be a powerful technique for identifying such peptidomes, and methods integrating such data for prediction of Ag presentation have reached a high level of accuracy for both MHC class I and class II. In this study, we demonstrate how these techniques and prediction methods can be readily extended to the bovine leukocyte Ag class II DR locus (BoLA-DR). BoLA-DR binding motifs were characterized by eluted ligand data derived from bovine cell lines expressing a range of DRB3 alleles prevalent in Holstein–Friesian populations. The model generated (NetBoLAIIpan, available as a Web server at www.cbs.dtu.dk/services/NetBoLAIIpan) was shown to have unprecedented predictive power to identify known BoLA-DR–restricted CD4 epitopes. In summary, the results demonstrate the power of an integrated approach combining advanced mass spectrometry peptidomics with immunoinformatics for characterization of the BoLA-DR Ag presentation system and provide a prediction tool that can be used to assist in rational evaluation and selection of bovine CD4 T cell epitopes

Molecular cloning and characterization of a highly selective chemokine-binding protein from the tick Rhipicephalus sanguineus.

Abstract Ticks are blood-feeding parasites that secrete a number of immuno-modulatory factors to evade the host immune response. Saliva isolated from different species of ticks has recently been shown to contain chemokine neutralizing activity. To characterize this activity, we constructed a cDNA library from the salivary glands of the common brown dog tick, Rhipicephalus sanguineus. Pools of cDNA clones from the library were transfected into HEK293 cells, and the conditioned media from the transfected cells were tested for chemokine binding activity by chemical cross-linking to radiolabeled CCL3 followed by SDS-PAGE. By de-convolution of a single positive pool of 270 clones, we identified a full-length cDNA encoding a protein of 114 amino acids, which after signal peptide cleavage was predicted to yield a mature protein of 94 amino acids that we called Evasin-1. Recombinant Evasin-1 was produced in HEK293 cells and in insect cells. Using surface plasmon resonance we were able to show that Evasin-1 was exquisitely selective for 3 CC chemokines, CCL3 and CCL4 and the closely related chemokine CCL18, with KD values of 0.16, 0.81, and 3.21 nm, respectively. The affinities for CCL3 and CCL4 were confirmed in competition receptor binding assays. Analysis by size exclusion chromatography demonstrated that Evasin-1 was monomeric and formed a 1:1 complex with CCL3. Thus, unlike the other chemokine-binding proteins identified to date from viruses and from the parasitic worm Schistosoma mansoni, Evasin-1 is highly specific for a subgroup of CC chemokines, which may reflect a specific role for these chemokines in host defense against parasites

Desenvolvimento e validação de tecnologia educacional interativa sobre febre maculosa

Objetivo: desarrollar y validar una tecnología educativa interactiva sobre fiebre maculosa, para ofrecer un método innovador de enseñanza. Método: estudio metodológico que recorrió las siguientes etapas: análisis y diagnóstico; planificación instructiva, diseño didáctico, revisión y validación y producción de la tecnología. Resultados: el análisis y el diagnóstico se obtuvieron a partir de experiencias en actividades de educación y salud para fiebre maculosa. En la planificación instructiva, se definió que la tecnología se presentaría en forma de Laboratorio Interactivo, con estaciones de aprendizaje. La producción del Laboratorio fue realizada por un equipo multidisciplinario constituido por un carpintero, un electricista y un artista plástico, entre otros. El proceso de revisión y validación se subdividió en dos etapas: validación de aspecto y contenido por profesionales de las áreas de biología y educación, y validación semántica por alumnos de las carreras de Enfermería y Pedagogía. Los resultados de la validación de aspecto y contenido mostraron un índice de validez de contenido superior a 0,8 para la gran mayoría de las variables. En la validación semántica, el Laboratorio fue evaluado de manera positiva por los alumnos Conclusión: la trayectoria recorrida para la construcción del Laboratorio Interactivo sobre fiebre maculosa dio sustentación académica y científica al producto construido, ofreciendo un recurso educativo innovador con potencial pedagógico que valora el aprendizaje significativo.Objective: to develop and validate an interactive educational technology on spotted fever, to offer an innovative teaching method. Method: a methodological study that covered the following stages: analysis and diagnosis; instructional planning, didactic design, review, and validation and production of technology. Results: the analysis and diagnosis were obtained from experiences in education and health activities for spotted fever. In the instructional planning, it was defined that the technology would be presented in the form of an Interactive Laboratory, with learning stations. The production of the Laboratory was carried out by a multidisciplinary team made up of a carpenter, an electrician, and a plastic artist, among others. The review and validation process was subdivided into two stages: appearance and content validation by professionals in the fields of biology, and education and semantic validation by students of the Nursing and Pedagogy courses. The results of the appearance and content validation showed a content validity index over 0.8 for the vast majority of the variables. In the semantic validation, the Laboratory was evaluated positively by the students. Conclusion: the trajectory followed for the construction of the Interactive Laboratory on spotted fever gave academic and scientific support to the product, offering an innovative educational resource with pedagogical potential that values significant learning.Objetivo: desenvolver e validar uma tecnologia educacional interativa sobre febre maculosa, para oferecer um método inovador de ensino. Método: estudo metodológico desenvolvido nas seguintes etapas: análise e diagnóstico; planejamento instrucional, desenho didático, revisão e validação e produção da tecnologia. Resultados: a análise e diagnóstico foram obtidos a partir de experiências em atividades de educação e saúde para febre maculosa. No planejamento instrucional, definiu-se que a tecnologia seria apresentada em forma de Laboratório Interativo, com estações de aprendizagem. A produção do Laboratório foi realizada por uma equipe multidisciplinar constituída por marceneiro, eletricista, artista plástico, dentre outros. O processo de revisão e validação foi subdividido em duas etapas: validação de aparência e conteúdo por profissionais das áreas de biologia e educação e validação semântica por alunos do curso de enfermagem e pedagogia. Os resultados da validação de aparência e conteúdo mostraram um índice de validade de conteúdo superior a 0,8 para a grande maioria das variáveis. Na validação semântica, o Laboratório foi avaliado de forma positiva pelos alunos. Conclusão: a trajetória percorrida para a construção do Laboratório Interativo sobre febre maculosa conferiu sustentação acadêmica e científica ao produto construído, oferecendo um recurso educativo inovador com potencial pedagógico que valoriza a aprendizagem significativa

Haplotypes of the bovine IgG2 heavy gamma chain in tick-resistant and tick-susceptible breeds of cattle

Bovines present contrasting, heritable phenotypes of infestations with the cattle tick, Rhipicephalus (Boophilus) microplus. Tick salivary glands produce IgG-binding proteins (IGBPs) as a mechanism for escaping from host antibodies that these ectoparasites ingest during blood meals. Allotypes that occur in the constant region of IgG may differ in their capacity to bind with tick IGBPs; this may be reflected by the distribution of distinct allotypes according to phenotypes of tick infestations. In order to test this hypothesis, we investigated the frequency of haplotypes of bovine IgG2 among tick-resistant and tick-susceptible breeds of bovines. Sequencing of the gene coding for the heavy chain of IgG2 from 114 tick-resistant (Bos taurus indicus, Nelore breed) and tick-susceptible (B. t. taurus, Holstein breed) bovines revealed SNPs that generated 13 different haplotypes, of which 11 were novel and 5 were exclusive of Holstein and 3 of Nelore breeds. Alignment and modeling of coded haplotypes for hinge regions of the bovine IgG2 showed that they differ in the distribution of polar and hydrophobic amino acids and in shape according to the distribution of these amino acids. We also found that there was an association between genotypes of the constant region of the IgG2 heavy chain with phenotypes of tick infestations. These findings open the possibility of investigating if certain IgG allotypes hinder the function of tick IGBPs. If so, they may be markers for breeding for resistance against tick infestations

SARS-CoV-2 susceptibility and COVID-19 disease severity are associated with genetic variants affecting gene expression in a variety of tissues

Variability in SARS-CoV-2 susceptibility and COVID-19 disease severity between individuals is partly due to genetic factors. Here, we identify 4 genomic loci with suggestive associations for SARS-CoV-2 susceptibility and 19 for COVID-19 disease severity. Four of these 23 loci likely have an ethnicity-specific component. Genome-wide association study (GWAS) signals in 11 loci colocalize with expression quantitative trait loci (eQTLs) associated with the expression of 20 genes in 62 tissues/cell types (range: 1:43 tissues/gene), including lung, brain, heart, muscle, and skin as well as the digestive system and immune system. We perform genetic fine mapping to compute 99% credible SNP sets, which identify 10 GWAS loci that have eight or fewer SNPs in the credible set, including three loci with one single likely causal SNP. Our study suggests that the diverse symptoms and disease severity of COVID-19 observed between individuals is associated with variants across the genome, affecting gene expression levels in a wide variety of tissue types