EXTERNAL MORPHOLOGY OF THE IMMATURES OF POLYBIA PAULISTA (HYMENOPTERA: VESPIDAE)

The immatures of Polybia paulista Ihering were described using light and scanning electron microscopy and the results are compared with previous descriptions within the same or related wasps. This study is based on 2 whole nests collected in the municipality of Rio Claro, Sao Paulo, in Brazil. We have detected the existence of 5 larval instars. The main morphological alterations over development occur in the relative size of structures, yet certain structures appear with subsequent instars and become more evident later in development: increasing density in the number of body spines and papillae; the appearance of body setae in fifth-instar larvae; opening of spiracles upon second-instar larvae; 2 body shapes in fifth-instar larvae; the appearance of a lateral tooth on the mandibles of fourth instar; presence of spines on the maxillae of fifth-instar larvae; altered shape of galea and palps upon third-instar larvae from a cluster of sensilla to a conical elevation; and the appearance of spines on postmentum upon fourth-instar larvae. This way, the present study presents a detailed description of the immatures of P. paulista, and we hope the presented information can be useful to morphological, taxonomic, and phylogenetic studies

Structure–function relationships of the peptide Paulistine: A novel toxin from the venom of the social wasp Polybia paulista

AbstractBackgroundThe peptide Paulistine was isolated from the venom of wasp Polybia paulista. This peptide exists under a natural equilibrium between the forms: oxidised — with an intra-molecular disulphide bridge; and reduced — in which the thiol groups of the cysteine residues do not form the disulphide bridge. The biological activities of both forms of the peptide are unknown up to now.MethodsBoth forms of Paulistine were synthesised and the thiol groups of the reduced form were protected with the acetamidemethyl group [Acm-Paulistine] to prevent re-oxidation. The structure/activity relationships of the two forms were investigated, taking into account the importance of the disulphide bridge.ResultsPaulistine has a more compact structure, while Acm-Paulistine has a more expanded conformation. Bioassays reported that Paulistine caused hyperalgesia by interacting with the receptors of lipid mediators involved in the cyclooxygenase type II pathway, while Acm-Paullistine also caused hyperalgesia, but mediated by receptors involved in the participation of prostanoids in the cyclooxygenase type II pathway.ConclusionThe acetamidemethylation of the thiol groups of cysteine residues caused small structural changes, which in turn may have affected some physicochemical properties of the Paulistine. Thus, the dissociation of the hyperalgesy from the edematogenic effect when the actions of Paulistine and Acm-Paulistine are compared to each other may be resulting from the influence of the introduction of Acm-group in the structure of Paulistine.General significanceThe peptides Paulistine and Acm-Paulistine may be used as interesting tools to investigate the mechanisms of pain and inflammation in future studies

Immunogenicity of personalized dendritic-cell therapy in HIV-1 infected individuals under suppressive antiretroviral treatment:interim analysis from a phase II clinical trial

BACKGROUND: We developed a personalized Monocyte-Derived Dendritic-cell Therapy (MDDCT) for HIV-infected individuals on suppressive antiretroviral treatment and evaluated HIV-specific T-cell responses. METHODS: PBMCs were obtained from 10 HIV(+) individuals enrolled in trial NCT02961829. Monocytes were differentiated into DCs using IFN-α and GM-CSF. After sequencing each patient’s HIV-1 Gag and determining HLA profiles, autologous Gag peptides were selected based on the predicted individual immunogenicity and used to pulse MDDCs. Three doses of the MDDCT were administered every 15 days. To assess immunogenicity, patients’ cells were stimulated in vitro with autologous peptides, and intracellular IL-2, TNF, and interferon-gamma (IFN-γ) production were measured in CD4(+) and CD8(+) T-cells. RESULTS: The protocol of ex-vivo treatment with IFN-α and GM-CSF was able to induce maturation of MDDCs, as well as to preserve their viability for reinfusion. MDDCT administration was associated with increased expression of IL-2 in CD4(+) and CD8(+) T-cells at 15 and/or 30 days after the first MDDCT administration. Moreover, intracellular TNF and IFN-γ expression was significantly increased in CD4(+) T-cells. The number of candidates that increased in vitro the cytokine levels in CD4(+) and CD8(+) T cells upon stimulation with Gag peptides from baseline to day 15 and from baseline to day 30 and day 120 after MDDCT was significant as compared to Gag unstimulated response. This was accompanied by an increasing trend in the frequency of polyfunctional T-cells over time, which was visible when considering both cells expressing two and three out of the three cytokines examined. CONCLUSIONS: MDDC had a mature profile, and this MDDCT promoted in-vitro T-cell immune responses in HIV-infected patients undergoing long-term suppressive antiretroviral treatment. Trial registration NCT02961829: (Multi Interventional Study Exploring HIV-1 Residual Replication: a Step Towards HIV-1 Eradication and Sterilizing Cure, https://www.clinicaltrials.gov/ct2/show/NCT02961829, posted November 11th, 2016) SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12981-021-00426-z

Rationale, study design, and analysis plan of the Alveolar Recruitment for ARDS Trial (ART): Study protocol for a randomized controlled trial

Background: Acute respiratory distress syndrome (ARDS) is associated with high in-hospital mortality. Alveolar recruitment followed by ventilation at optimal titrated PEEP may reduce ventilator-induced lung injury and improve oxygenation in patients with ARDS, but the effects on mortality and other clinical outcomes remain unknown. This article reports the rationale, study design, and analysis plan of the Alveolar Recruitment for ARDS Trial (ART). Methods/Design: ART is a pragmatic, multicenter, randomized (concealed), controlled trial, which aims to determine if maximum stepwise alveolar recruitment associated with PEEP titration is able to increase 28-day survival in patients with ARDS compared to conventional treatment (ARDSNet strategy). We will enroll adult patients with ARDS of less than 72 h duration. The intervention group will receive an alveolar recruitment maneuver, with stepwise increases of PEEP achieving 45 cmH(2)O and peak pressure of 60 cmH2O, followed by ventilation with optimal PEEP titrated according to the static compliance of the respiratory system. In the control group, mechanical ventilation will follow a conventional protocol (ARDSNet). In both groups, we will use controlled volume mode with low tidal volumes (4 to 6 mL/kg of predicted body weight) and targeting plateau pressure <= 30 cmH2O. The primary outcome is 28-day survival, and the secondary outcomes are: length of ICU stay; length of hospital stay; pneumothorax requiring chest tube during first 7 days; barotrauma during first 7 days; mechanical ventilation-free days from days 1 to 28; ICU, in-hospital, and 6-month survival. ART is an event-guided trial planned to last until 520 events (deaths within 28 days) are observed. These events allow detection of a hazard ratio of 0.75, with 90% power and two-tailed type I error of 5%. All analysis will follow the intention-to-treat principle. Discussion: If the ART strategy with maximum recruitment and PEEP titration improves 28-day survival, this will represent a notable advance to the care of ARDS patients. Conversely, if the ART strategy is similar or inferior to the current evidence-based strategy (ARDSNet), this should also change current practice as many institutions routinely employ recruitment maneuvers and set PEEP levels according to some titration method.Hospital do Coracao (HCor) as part of the Program 'Hospitais de Excelencia a Servico do SUS (PROADI-SUS)'Brazilian Ministry of Healt

Análise proteômica do veneno da vespa social Polistes lanio lanio (Hymenoptera; vespidae)

Os venenos dos insetos da ordem Hymenoptera (abelhas, vespas e formigas) são responsáveis por um grande número de acidentes causados por ferroadas. Estes acidentes podem produzir uma série de reações, variando desde uma reação local, até uma reação sistêmica e anafilaxia. Estudos mostraram que 0,8% a 5% da população mundial sofrem de reações sistêmicas generalizadas após ferroadas de insetos pertencentes à ordem Hymenoptera. As espécies pertencentes ao gênero Polistes são tipicamente encontradas no sudeste do Brasil, causando muitos acidentes por ferroadas devido principalmente à proximidade dos ninhos destes insetos das habitações humanas. P.lanio lanio é uma das vespas sociais que mais causam acidentes no Estado de São Paulo e pouco se sabe sobre a composição de seu veneno. As vespas do gênero Polistes são capazes de ferroar múltiplas vezes e causar reações alérgicas severas. Dessa forma, a identificação das proteínas mais abundantes do veneno da vespa social Polistes lanio lanio por uma abordagem proteômica, se faz necessária para uma melhor compreensão dos mecanismos de ação desse veneno. A identificação de várias proteínas do veneno de P. l. lanio, revelou importantes aspectos sobre o processo de envenenamento por vespas do gênero Polistes, os quais podem ajudar no melhor entendimento dos mecanismos de ação destes venenos. A compreensão dos principais alérgenos é uma etapa importante para o desenvolvimento de novos extratos específicos para diagnósticos de alergia e imunoterapia de pacientes sensíveis ao veneno de vespas

O uso de um sistema LC-ESI-IT-TOF/MS e MSn na prospecção de novos componentes peptídicos do veneno da vespa social Polybia paulista

O desenvolvimento dos venenos e aparelhos de ferroar entre os Insecta representa um atributo evolutivo que contribuiu para a adaptação dos insetos em muitos ambientes terrestres diferentes. Os venenos desses insetos são misturas complexas de compostos biologicamente ativos, tais como compostos de baixa massa molecular, peptídeos e proteínas. Estudos recentes têm demonstrado a capacidade de se detectar e identificar diferentes compostos de natureza peptídica, em concentrações bastante reduzidas nos venenos animais, utilizando-se diferentes abordagens de espectrometria de massas. Isso tem permitido a construção de bibliotecas peptídicas de grande interesse aplicado à biotecnologia. O veneno da vespa social Polybia paulista tem sido intensamente investigado, porém somente os peptídeos mais abundantes deste veneno são conhecidos: os mastoparanos Polybia -MPI e -MPII, o peptídeo quimiotáctico Polybia-CP e a Paulistina. Este fato deve-se principalmente à utilização de abordagens clássicas até então, com coletas “off-line” em relação às análises de sequenciamento, fazendo com que somente os peptídeos mais abundantes pudessem ser investigados. Com os avanços na área da espectrometria de massas, incluindo o desenvolvimento da tecnologia de analisadores do tipo “ion-trap” utilizado no presente trabalho, tornou-se possível a investigação de amostras pouco abundantes, com alta velocidade de aquisição de dados e elevada resolução. O presente estudo visou a obtenção de um perfil peptídico detalhado do veneno de P. paulista por uma abordagem analítica moderna e mais sensível, além de padronizar o sequenciamento destes peptídeos por espectrometria de massas sequencial de maneira “on-line”. Os peptídeos foram detectados e sequenciados utilizando-se um sistema LC-ESI-IT-TOF/MS e MSn...The evolution of venoms and their injection apparatuses among the Insecta represents an evolutionary attribute which contributed for thr adaptation of the insects to many different terrestrial environments. The venoms of insects are complex mixtures of biologically active compounds, such as low molecular mass compounds, peptides and proteins. Recent studies have demonstrated the ability to detect and identify different compounds in very low concentrations in animal venoms, using different approaches of mass spectrometry. This has allowed the construction of peptide libraries of great applied interest in biotechnology. The venom of the social wasp Polybia paulista has been intensively investigated, but only the most abundant peptides of this venom are known: the mastoparans Polybia-MPI and -MPII, the chemotactic peptide Polybia-CP and the Paulistine. This fact is mainly due to the use of classical approaches, using offline collections in relation to peptide sequencing, so only the more abundant peptides could be investigated. With the advances in mass spectrometry, including the development of analyzers type ion-trap as the one used in present work, it became possible to investigate low abundance samples, with high speed of data acquisition at high resolution. The objective of this study was to profile and sequence the peptide compounds present in the venom of the social wasp P. paulista using a modern and very sensitive analytical technique, and standardize the sequencing of peptides by high resolution LC-MS and MSn strategy. Peptides were detected and sequenced using a LCESI- IT-TOF/MS and MSn system of high resolution and sensitivity. The analysis of the venom was performed by high performance liquid chromatography on a reverse phase (HPLC), and the analytes (peptides) were characterized ... (Complete abstract click electronic access below)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Estudo peptídico e determinação do perfil de metabólitos de venenos de escorpiões da família buthidae: Tityus serrulatus, Tityus bahiensis e Tityus obscurus

Numerosos estudos produzidos nos últimos 30 anos, vêm elucidando estruturas e atividades biológicas de vários componentes de venenos animais, demonstrando suas complexas composições. Os venenos de escorpiões são misturas complexas de compostos biologicamente ativos, tais como: compostos de baixa massa molar, peptídeos e proteínas, sendo estas últimas as mais amplamente estudadas. Até então, pouco se conhece sobre os pequenos peptídeos e metabolitos dos venenos de escorpiões, sendo necessárias pesquisas para ampliar esse conhecimento. Este estudo visou a obtenção de um perfil peptídico detalhado dos venenos das espécies T. serrulatus, T. bahienis e T. obscurus, com foco principal nos pequenos peptídeos lineares, antes negligenciados em venenos de escorpiões. A extração em acetonitrila 50 % (v/v) foi uma das diferenças estratégicas para esse objetivo, e a separação e sequenciamento destes peptídeos foram realizados utilizando-se um sistema LCESI- IT-TOF. Os componentes peptídicos dos venenos dos escorpiões estudados estão distribuídos na faixa de massa molar de 400-4000 Da, sendo possível detectar a presença de 521 peptídeos distintos no veneno de T. serrulatus, 460 peptídeos no veneno de T. bahiensis e 517 peptídeos no veneno de T. obscurus. Também foi possível estabelecer os perfis peptídicos dos venenos das três espécies estudadas, assim como obter a sequencia de 66 peptídeos, dos quais alguns apresentaram semelhanças com toxinas já conhecidas de escorpiões. Treze dos novos peptídeos detectados, contendo entre 6 e 19 aminoácidos, foram sintetizados e caracterizados com relação às suas atividades biológicas em ensaios de antibiose, hemólise, desgranulação de mastócitos, LDH, ensaio de campo aberto, dor e inflamação em mamíferos, demonstrando algumas atividades principalmente com relação à hemólise, dor e locomoção de mamíferos. Este trabalho também elucidou o perfil de metabólitos nestes venenos em um sistema LCMS-Q-TOF-MS de maneira qualitativa e quantitativa. Para essa finalidade, foi realizada a padronização das condições cromatográficas com utilização de cromatografia de pareamento iônico para a melhor separação e identificação de compostos tais como aminas biogênicas, poliaminas, aminoácidos, derivados de nucleosídeos, etc., detectados nos venenos. A complexidade química demonstrada no presente trabalho corrobora com o fato de que as moléculas pequenas, tanto peptídeos quanto metabólitos, deverão agir em sinergismo com as grandes toxinas nos venenos, potencializando seus efeitos em vítimas de ferroadas por esses animais.Numerous studies produced in the last 30 years, come elucidating structures and biological activities of various components of animal venoms, showing their complex compositions. The scorpion venoms are complex mixtures of biologically active compounds, such as low molecular weight compounds, peptides and proteins, the latter being the most widely studied. To the present time not much is known about the small peptides and metabolites of scorpion venoms, requiring research to expand this knowledge. This study aimed to obtain a detailed peptide profile of the venoms of T. serrulatus, T. bahienis and T. obscurus species, focusing primarily on small linear neglected peptides in scorpion venom. The extraction in 50% acetonitrile was one of the strategic differences for this purpose and the separation and sequencing of these peptides were performed "on-line" using an LC-ESI-ITTOF system. The peptide components of the scorpion venoms are distributed along the molar mass range of 400-4000 Da, and in this study we could detect the presence of 521 different peptides in T. serrulatus venom, 460 peptides in T. bahiensis and 517 in T. obscurus. It was also possible to establish the peptide profiles of the venoms of the three species studied, as well as to obtain the sequence of 66 peptides, some of which had similarities to known already toxins of scorpions. Thirteen of the newly detected peptides, containing between 6 and 19 amino acids, were synthesized and characterized with proposal to obtain their biological activities, such antibiosis, hemolysis, mast cell degranulation, LDH, open field test, pain and inflammation in mammals, showing some particularly activities in hemolysis, pain and mammals locomotion. This work also revealed the metabolites profile in these venoms with a LCMS-Q-TOF-MS system for qualitative and quantitative analysis. For this purpose, we standardized the chromatographic conditions with the use of ion pairing chromatography to better separation and identification of compounds such as biogenic amines, polyamines, amino acids, nucleosides derivatives, etc., detected in the venoms. The chemical complexity demonstrated in this study corroborates with the fact that small peptides and metabolites should act in synergism with large toxins in poisonings, increasing its effects on victims by these animals.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES

External Morphology of the Immatures of Polybia paulista (Hymenoptera: Vespidae)

The immatures of Polybia paulista Ihering were described using light and scanning electron microscopy and the results are compared with previous descriptions within the same or related wasps. This study is based on 2 whole nests collected in the municipality of Rio Claro, São Paulo, in Brazil. We have detected the existence of 5 larval instars. The main morphological alterations over development occur in the relative size of structures, yet certain structures appear with subsequent instars and become more evident later in development: increasing density in the number of body spines and papillae; the appearance of body setae in fifth-instar larvae; opening of spiracles upon second-instar larvae; 2 body shapes in fifth-instar larvae; the appearance of a lateral tooth on the mandibles of fourth instar; presence of spines on the maxillae of fifth-instar larvae; altered shape of galea and palps upon third-instar larvae from a cluster of sensilla to a conical elevation; and the appearance of spines on postmentum upon fourth-instar larvae. This way, the present study presents a detailed description of the immatures of P. paulista, and we hope the presented information can be useful to morphological, taxonomic, and phylogenetic studies.Os imaturos de Polybia paulista Ihering foram descritos e comparados, com o auxílio da microscopia óptica e eletrônica de varredura. Foram analisados os imaturos provenientes de dois ninhos obtidos no município de Rio Claro, SP, Brasil. Verificou-se a existência de cinco instares larvais. Durante o desenvolvimento ocorrem principalmente mudanças no tamanho das estruturas. Além disso, certas estruturas surgem e se tornam evidentes com o desenvolvimento da larva: aumento da densidade de espinhos e papilas no corpo; presença de setas no corpo da larva de último instar; abertura dos espiráculos a partir do segundo instar; dois formatos de corpo na larva de último instar; a partir do quarto instar, a presença de dente lateral na mandíbula; presença de espinhos na maxila do último instar; mudança do formato da galea e palpos a partir do terceiro instar, de um agrupamento de sensilas para um cone; e o surgimento de espinhos na gula a partir do quarto instar. Desta maneira, o presente estudo efetuou uma caracterização detalhada dos imaturos de P. paulista, cujas informações poderão ser talvez úteis para estudos morfológicos, taxonômicos e filogenéticos

Paulistine—The Functional Duality of a Wasp Venom Peptide Toxin

It has been reported that Paulistine in the venom of the wasp Polybia paulista co-exists as two different forms: an oxidized form presenting a compact structure due to the presence of a disulfide bridge, which causes inflammation through an apparent interaction with receptors in the 5-lipoxygenase pathway, and a naturally reduced form (without the disulfide bridge) that exists in a linear conformation and which also causes hyperalgesia and acts in the cyclooxygenase type II pathway. The reduced peptide was acetamidomethylated (Acm-Paulistine) to stabilize this form, and it still maintained its typical inflammatory activity. Oxidized Paulistine docks onto PGHS2 (COX-2) molecules, blocking the access of oxygen to the heme group and inhibiting the inflammatory activity of Acm-Paulistine in the cyclooxygenase type II pathway. Docking simulations revealed that the site of the docking of Paulistine within the PGHS2 molecule is unusual among commercial inhibitors of the enzyme, with an affinity potentially much higher than those observed for traditional anti-inflammatory drugs. Therefore, Paulistine causes inflammatory activity at the level of the 5-lipooxygenase pathway and, in parallel, it competes with its reduced form in relation to the activation of the cyclooxygenase pathway. Thus, while the reduced Paulistine causes inflammation, its oxidized form is a potent inhibitor of this activity