Parasite DNA and Markers of Decreased Immune Activation Associate Prospectively with Cardiac Functional Decline over 10 Years among Trypanosoma cruzi Seropositive Individuals in Brazil.

Parasitemia and inflammatory markers are cross-sectionally associated with chronic Chagas cardiomyopathy (CCC) among patients with Trypanosoma cruzi. However, the prospective association of the parasite load and host immune response-related characteristics with CCC (that is, progressors) among T. cruzi seropositive individuals has only been partially defined. In a cohort of T. cruzi seropositive patients in Montes Claros and São Paulo, Brazil who were followed over 10 years, we identified the association of a baseline T. cruzi parasite load and systemic markers of inflammation with a decline in cardiac function and/or the presence of cardiac congestion 10 years later. The progressors (n = 21) were individuals with a significant decline in the left ventricular ejection fraction and/or elevated markers of cardiac congestion after 10 years. The controls (n = 31) had normal markers of cardiac function and congestion at the baseline and at the follow-up. They were matched with the progressors on age, sex, and genetic ancestry. The progressors had higher mean parasite loads at the baseline than the controls (18.3 vs. 0.605 DNA parasite equivalents/20 mL, p < 0.05). Of the 384 inflammation-related proteins analyzed, 47 differed significantly at a false discovery rate- (FDR-) corrected p < 0.05 between the groups. There were 44 of these 47 proteins that were significantly higher in the controls compared to in the progressors, including the immune activation markers CCL21, CXCL12, and HCLS1 and several of the tumor necrosis factor superfamily of proteins. Among the individuals who were seropositive for T. cruzi at the baseline and who were followed over 10 years, those with incident CCC at the 10-year marker had a comparatively higher baseline of T. cruzi parasitemia and lower baseline markers of immune activation and chemotaxis. These findings generate the hypothesis that the early impairment of pathogen-killing immune responses predisposes individuals to CCC, which merits further study.

Unraveling the role of miRNAs as biomarkers in Chagas cardiomyopathy: Insights into molecular pathophysiology.

BACKGROUND: Chagas cardiomyopathy (ChCM) is a severe form of Chagas disease and a major cause of cardiovascular morbidity and mortality. The dysregulation of the immune response leads to cardiac remodeling and functional disruptions, resulting in life-threatening complications. Conventional diagnostic methods have limitations, and therapeutic response evaluation is challenging. MicroRNAs (miRNAs), important regulators of gene expression, show potential as biomarkers for diagnosis and prognosis. AIM: This review aims to summarize experimental findings on miRNA expression in ChCM and explore the potential of these miRNAs as biomarkers of Chagas disease. METHODS: The search was conducted in the US National Library of Medicine MEDLINE/PubMed public database using the terms "Chagas cardiomyopathy" OR "Chagas disease" AND "microRNA" OR "miRNA" OR "miR." Additionally, bioinformatics analysis was performed to investigate miRNA-target interactions and explore enrichment pathways of gene ontology biological processes and molecular functions. RESULTS: The miR-21, miR-146b, miR-146a, and miR-155 consistently exhibited up-regulation, whereas miR-145 was down-regulated in ChCM. These specific miRNAs have been linked to fibrosis, immune response, and inflammatory processes in heart tissue. Moreover, the findings from various studies indicate that these miRNAs have the potential as biomarkers for the disease and could be targeted in therapeutic strategies for ChCM. CONCLUSION: In this review, we point out miR-21, miR-146b, miR-146a, miR-155, and miR-145-5p role in the complex mechanisms of ChCM. These miRNAs have been shown as potential biomarkers for precise diagnosis, reliable prognostic evaluation, and effective treatment strategies in the ChCM.

Anti-RBD IgG antibodies from endemic coronaviruses do not protect against the acquisition of SARS-CoV-2 infection among exposed uninfected individuals.

Background: The Coronaviridae family comprises seven viruses known to infect humans, classified into alphacoronaviruses (HCoV-229E and HCoV-NL63) and betacoronaviruses (HCoV-OC43 and HCoV-HKU1), which are considered endemic. Additionally, it includes SARS-CoV (severe acute respiratory syndrome), MERS-CoV (Middle East respiratory syndrome), and the novel coronavirus SARS-CoV-2, responsible for COVID-19. SARS-CoV-2 induces severe respiratory complications, particularly in the elderly, immunocompromised individuals and those with underlying diseases. An essential question since the onset of the COVID-19 pandemic has been to determine whether prior exposure to seasonal coronaviruses influences immunity or protection against SARS-CoV-2. Methods: In this study, we investigated a cohort of 47 couples (N=94), where one partner tested positive for SARS-CoV-2 infection via real-time PCR while the other remained negative. Plasma samples, collected at least 30 days post-PCR reaction, were assessed using indirect ELISA and competition assays to measure specific antibodies against the receptor-binding domain (RBD) portion of the Spike (S) protein from SARS-CoV-2, HCoV-229E, HCoV-NL63, HCoV-OC43, and HCoV-HKU1. Results: IgG antibody levels against the four endemic coronavirus RBD proteins were similar between the PCR-positive and PCR-negative individuals, suggesting that IgG against endemic coronavirus RBD regions was not associated with protection from infection. Moreover, we found no significant IgG antibody cross-reactivity between endemic coronaviruses and SARS-CoV-2 RBDs. Conclusions: Taken together, results suggest that anti-RBD antibodies induced by a previous infection with endemic HCoVs do not protect against acquisition of COVID-19 among exposed uninfected individuals.

Post-COVID-19 condition: systemic inflammation and low functional exercise capacity.

Introduction: Post-COVID-19 condition (PCC) is characterised by a plethora of symptoms, with fatigue appearing as the most frequently reported. The alterations that drive both the persistent and post-acute disease newly acquired symptoms are not yet fully described. Given the lack of robust knowledge regarding the mechanisms of PCC we have examined the impact of inflammation in PCC, by evaluating serum cytokine profile and its potential involvement in inducing the different symptoms reported. Methods: In this cross-sectional study, we recruited 227 participants who were hospitalised with acute COVID-19 in 2020 and came back for a follow-up assessment 6-12 months after hospital discharge. The participants were enrolled in two symptomatic groups: Self-Reported Symptoms group (SR, n = 96), who did not present major organ lesions, yet reported several debilitating symptoms such as fatigue, muscle weakness, and persistent loss of sense of smell and taste; and the Self-Reported Symptoms and decreased Pulmonary Function group (SRPF, n = 54), composed by individuals with the same symptoms described by SR, plus diagnosed pulmonary lesions. A Control group (n = 77), with participants with minor complaints following acute COVID-19, was also included in the study. Serum cytokine levels, symptom questionnaires, physical performance tests and general clinical data were obtained in the follow-up assessment. Results: SRPF presented lower IL-4 concentration compared with Control (q = 0.0018) and with SR (q = 0.030), and lower IFN-α2 serum content compared with Control (q = 0.007). In addition, SRPF presented higher MIP-1ß serum concentration compared with SR (q = 0.029). SR presented lower CCL11 (q = 0.012 and q = 0.001, respectively) and MCP-1 levels (q = 0.052 for both) compared with Control and SRPF. SRPF presented lower G-CSF compared to Control (q = 0.014). Female participants in SR showed lower handgrip strength in relation to SRPF (q = 0.0082). Male participants in SR and SRPF needed more time to complete the timed up-and-go test, as compared with men in the Control group (q = 0.0302 and q = 0.0078, respectively). Our results indicate that different PCC symptom profiles are accompanied by distinct inflammatory markers in the circulation. Of particular concern are the lower muscle function findings, with likely long-lasting consequences for health and quality of life, found for both PCC phenotypes.

Inflammation and mitochondria in the pathogenesis of chronic Chagas disease cardiomyopathy.

Chagas disease (CD), caused by the protozoan parasite Trypanosoma cruzi, is a neglected disease affecting around 6 million people. About 30% of CD patients develop chronic Chagas disease cardiomyopathy (CCC), an inflammatory cardiomyopathy that occurs decades after the initial infection, while most infected patients (60%) remain asymptomatic in the so-called indeterminate form (IF). Death results from heart failure or arrhythmia in a subset of CCC patients. Myocardial fibrosis, inflammation, and mitochondrial dysfunction are involved in the arrhythmia substrate and triggering events. Survival in CCC is worse than in other cardiomyopathies, which may be linked to a Th1-T cell rich myocarditis with abundant interferon (IFN)-γ and tumor necrosis factor (TNF)-α, selectively lower levels of mitochondrial energy metabolism enzymes in the heart, and reduced levels of high-energy phosphate, indicating poor adenosine triphosphate (ATP) production. IFN-γ and TNF-α signaling, which are constitutively upregulated in CD patients, negatively affect mitochondrial function in cardiomyocytes, recapitulating findings in CCC heart tissue. Genetic studies such as whole-exome sequencing (WES) in nuclear families with multiple CCC/IF cases has disclosed rare heterozygous pathogenic variants in mitochondrial and inflammatory genes segregating in CCC cases. In this minireview, we summarized studies showing how IFN-γ and TNF-α affect cell energy generation, mitochondrial health, and redox homeostasis in cardiomyocytes, in addition to human CD and mitochondria. We hypothesize that cytokine-induced mitochondrial dysfunction in genetically predisposed patients may be the underlying cause of CCC severity and we believe this mechanism may have a bearing on other inflammatory cardiomyopathies.

Galectina-3 Associada a Formas Graves e Mortalidade em Longo Prazo em Pacientes com Doença de Chagas / Galectin-3 Associated with Severe Forms and Long-term Mortality in Patients with Chagas Disease

Resumo Fundamento As características histopatológicas da doença de Chagas (DCC) são: presença de miocardite, destruição das fibras cardíacas e fibrose miocárdica. A Galectina-3 (Gal-3) é um biomarcador envolvido no mecanismo de fibrose e inflamação que pode ser útil para a estratificação de indivíduos com DCC por risco. Objetivos Nosso objetivo foi avaliar se níveis elevados de Gal-3 estão associados a formas graves de cardiomiopatia chagásica (CC) e são preditivos de mortalidade. Métodos Estudamos doadores de sangue (DS) positivos para anti-T. cruzi: não-CC-DS (187 DS sem CC com eletrocardiograma [ECG] e fração de ejeção do ventrículo esquerdo [FEVE] normais); CC-Não-Dis-DS (46 DS com CC e apresentando ECG anormal, mas FEVE normal); e 153 controles negativos correspondentes. Esta amostra foi composta por 97 pacientes com CC grave (CC-Dis). Usamos as correlações de Kruskall-Wallis e Spearman para testar a hipótese de associações, assumindo um p bicaudal <0,05 como significativo. Resultados O nível de Gal-3 foi de 12,3 ng/mL para não-CC-DS, 12,0 ng/mL para CC-Não-Dis-DS, 13,8 ng/mL para controles e 15,4 ng/mL para CC-Dis. FEVE <50 foi associada a níveis mais elevados de Gal-3 (p=0,0001). Em nosso modelo de regressão linear ajustado, encontramos associação entre os níveis de Gal-3 e os parâmetros do ecocardiograma em indivíduos positivos para T. cruzi. Nos pacientes CC-Dis, encontramos uma associação significativa de níveis mais elevados de Gal-3 (≥15,3 ng/mL) e morte ou transplante cardíaco em acompanhamento de cinco anos (Hazard ratio - HR 3,11; IC95% 1,21- 8,04; p=0,019). Conclusões Em pacientes com CC, níveis mais elevados de Gal-3 estiveram significativamente associados a formas graves da doença e maior taxa de mortalidade em longo prazo, o que significa que pode ser um meio efetivo para identificar pacientes de alto risco. (Arq Bras Cardiol. 2021; 116(2):248-256)

Abstract Background The histopathological characteristics of Chagas disease (ChD) are: presence of myocarditis, destruction of heart fibers, and myocardial fibrosis. Galectin-3 (Gal-3) is a biomarker involved in the mechanism of fibrosis and inflammation that may be useful for risk stratification of individuals with ChD. Objectives We sought to evaluate whether high Gal-3 levels are associated with severe forms of Chagas cardiomyopathy (CC) and whether they are predictive of mortality. Methods We studied anti-T. cruzi positive blood donors (BD): Non-CC-BD (187 BD without CC with normal electrocardiogram [ECG] and left ventricular ejection fraction [LVEF]); CC-Non-Dys-BD (46 BD with CC with abnormal ECG but normal LVEF); and 153 matched serum-negative controls. This cohort was composed of 97 patients with severe CC (CC-Dys). We used Kruskall-Wallis and Spearman's correlation to test hypothesis of associations, assuming a two-tailed p<0.05 as significant. Results The Gal-3 level was 12.3 ng/mL for Non-CC-BD, 12.0 ng/mL for CC-Non-Dys-BD, 13.8 ng/mL for controls, and 15.4 ng/mL for CC-Dys. LVEF<50 was associated with higher Gal-3 levels (p=0.0001). In our linear regression adjusted model, we found association between Gal-3 levels and echocardiogram parameters in T. cruzi-seropositive subjects. In CC-Dys patients, we found a significant association of higher Gal-3 levels (≥15.3 ng/mL) and subsequent death or heart transplantation in a 5-year follow-up (Hazard ratio - HR 3.11; 95%CI 1.21-8.04; p=0.019). Conclusions In ChD patients, higher Gal-3 levels were significantly associated with severe forms of the disease and more long-term mortality, which means it may be a useful means to identify high-risk patients. (Arq Bras Cardiol. 2021; 116(2):248-256)

Co-administration of plasmid-encoded granulocyte-macrophage colony-stimulating factor increases human immunodeficiency virus-1 DNA vaccine-induced polyfunctional CD4+ T-cell responses

T-cell based vaccines against human immunodeficiency virus (HIV) generate specific responses that may limit both transmission and disease progression by controlling viral load. Broad, polyfunctional, and cytotoxic CD4+T-cell responses have been associated with control of simian immunodeficiency virus/HIV-1 replication, supporting the inclusion of CD4+ T-cell epitopes in vaccine formulations. Plasmid-encoded granulocyte-macrophage colony-stimulating factor (pGM-CSF) co-administration has been shown to induce potent CD4+ T-cell responses and to promote accelerated priming and increased migration of antigen-specific CD4+ T-cells. However, no study has shown whether co-immunisation with pGM-CSF enhances the number of vaccine-induced polyfunctional CD4+ T-cells. Our group has previously developed a DNA vaccine encoding conserved, multiple human leukocyte antigen (HLA)-DR binding HIV-1 subtype B peptides, which elicited broad, polyfunctional and long-lived CD4+ T-cell responses. Here, we show that pGM-CSF co-immunisation improved both magnitude and quality of vaccine-induced T-cell responses, particularly by increasing proliferating CD4+ T-cells that produce simultaneously interferon-γ, tumour necrosis factor-α and interleukin-2. Thus, we believe that the use of pGM-CSF may be helpful for vaccine strategies focused on the activation of anti-HIV CD4+ T-cell immunity.

Mycobacterium tuberculosis epitope-specific interferon-g production in healthy Brazilians reactive and non-reactive to tuberculin skin test

The interferon (IFN)-γ response to peptides can be a useful diagnostic marker of Mycobacterium tuberculosis (MTB) latent infection. We identified promiscuous and potentially protective CD4+ T-cell epitopes from the most conserved regions of MTB antigenic proteins by scanning the MTB antigenic proteins GroEL2, phosphate-binding protein 1 precursor and 19 kDa antigen with the TEPITOPE algorithm. Seven peptide sequences predicted to bind to multiple human leukocyte antigen (HLA)-DR molecules were synthesised and tested with IFN-γ enzyme-linked immunospot (ELISPOT) assays using peripheral blood mononuclear cells (PBMCs) from 16 Mantoux tuberculin skin test (TST)-positive and 16 TST-negative healthy donors. Eighty-eight percent of TST-positive donors responded to at least one of the peptides, compared to 25% of TST-negative donors. Each individual peptide induced IFN-γ production by PBMCs from at least 31% of the TST-positive donors. The magnitude of the response against all peptides was 182 ± 230 x 106 IFN-γ spot forming cells (SFC) among TST-positive donors and 36 ± 62 x 106 SFC among TST-negative donors (p = 0.007). The response to GroEL2 (463-477) was only observed in the TST-positive group. This combination of novel MTB CD4 T-cell epitopes should be tested in a larger cohort of individuals with latent tuberculosis (TB) to evaluate its potential to diagnose latent TB and it may be included in ELISPOT-based IFN-γ assays to identify individuals with this condition.

The effect of beta-blockade on myocardial remodelling in Chagas' cardiomyopathy

OBJECTIVE: Chagas' disease has spread throughout Latin America because of the high rate of migration among these countries. Approximately 30% of Chagas' patients will develop cardiomyopathy, and 10% of these will develop severe cardiac damage leading to heart failure. Beta-blockade improves symptoms and survival in heart failure patients; however, its efficacy has not been well established in Chagas' disease. We evaluated the role of carvedilol in cardiac remodeling and mortality in a Chagas' cardiomyopathy animal model. METHODS: We studied Trypanosoma cruzi infection in 55 Syrian hamsters that were divided into three groups: control (15), infected (20), and infected + carvedilol (20). Animals underwent echocardiography, electrocardiography, and morphometry for collagen evaluation in ventricles stained with picrosirius red. RESULTS: The left ventricular diastolic diameter did not change between groups, although it was slightly larger in infected groups, as was left ventricular systolic diameter. Fractional shortening also did not change between groups, although it was slightly lower in infected groups. Collagen accumulation in the interstitial myocardial space was significantly higher in infected groups and was not attenuated by carvedilol. The same response was observed in the perivascular space. The survival curve showed significantly better survival in the control group compared with the infected groups; but no benefit of carvedilol was observed during the study. However, in the acute phase (up to 100 days of infection), carvedilol did reduce mortality. CONCLUSION: Carvedilol did not attenuate cardiac remodeling or mortality in this model of Chagas' cardiomyopathy. The treatment did improve survival in the acute phase of the disease.

Aspectos imunológicos na cardiopatia chagásica crônica / Immunological features in Chagas cardiomyopathy

A patogenia da cardiomiopatia chagásica crônica ainda é pouco conhecida, e diversos mecanismos patogênicos foram propostos, como a disautonomia cardíaca, os distúrbios da circulação microvascular, e o dano tecidual imunológico-inflamatório. As observações de que a miocardite é mais frequente e intensa nos estágios mais avançados da doença e de que o prognóstico da cardiopatia chagásica crônica é pior que o de outras cardiomiopatias dilatadas de etiologia não-inflamatória sugerem que o infiltrado inflamatório desempenha papel importante no dano miocárdio. Na última década, tem sido evidenciada a participação de citocinas inflamatórias e de quimiocinas na gênese do infiltrado inflamatório e dano tecidual, assim como de polimorfismos genéticos de genes envolvidos na resposta inflamatória. Cardiopatas chagásicos apresentam produção periférica aumentada de interferon-gama e de fator de necrose tumoral alfa (TNF-alfa), comparativamente a pacientes da forma indeterminada, e linfócitos T Th1, produtores de interferon-gama e TNF-alfa, são frequentes no infiltrado inflamatório da cardiopatia chagásica crônica. Neste trabalho, revisaremos os mecanismos imunológicos e moleculares de dano miocárdico na cardiopatias chagásica crônica humana, com ênfase nos resultados obtidos pelo nosso grupo de pesquisa.

Immunological and non-immunological effects of cytokines and chemokines in the pathogenesis of chronic Chagas disease cardiomyopathy

The pathogenesis of Chagas disease cardiomyopathy (CCC) is not well understood. Since studies show that myocarditis is more frequent during the advanced stages of the disease, and the prognosis of CCC is worse than that of other dilated cardiomyopathies of non-inflammatory aetiology, which suggest that the inflammatory infiltrate plays a major role in myocardial damage. In the last decade, increasing evidence has shown that inflammatory cytokines and chemokines play a role in the generation of the inflammatory infiltrate and tissue damage. CCC patients have an increased peripheral production of the inflammatory Th1 cytokines IFN-³ and TNF-± when compared to patients with the asymptomatic/indeterminate form. Moreover, Th1-T cells are the main producers of IFN-³ and TNF-± and are frequently found in CCC myocardial inflammatory infiltrate. Over the past several years, our group has collected evidence that shows several cytokines and chemokines produced in the CCC myocardium may also have a non-immunological pathogenic effect via modulation of gene and protein expression in cardiomyocytes and other myocardial cell types. Furthermore, genetic polymorphisms of cytokine, chemokine and innate immune response genes have been associated with disease progression. We will review the molecular and immunological mechanisms of myocardial damage in human CCC in light of recent findings.

Chagas disease cardiomyopathy: current concepts of an old disease

Chagas disease continues to be a significant public health problem, as ca. 10 million people are still infected with T. cruzi in Latin America. Decades after primary infection, 30 percent of individuals can develop a form of chronic inflammatory cardiomyopathy known as Chagas disease cardiomyopathy (CCC). Data from both murine models and human studies support the view that an autoimmune response as well as a parasite-driven immune response involving inflammatory cytokines and chemokines may both play a role in generating the heart lesions leading to CCC. This review aims to summarize recent advances in the understanding of the immunopathogenesis of Chagas disease cardiomyopathy.

A doença de Chagas continua sendo importante problema de saúde pública uma vez que cerca de 10 milhões de indivíduos ainda estão infectados pelo T. cruzi. Décadas após a infecção primária, aproximadamente 30 por cento dos indivíduos podem desenvolver uma cardiomiopatia inflamatória crônica, a chamada Cardiomiopatia Chagásica Crônica (CCC). Dados de modelos murinos e de estudos em humanos apóiam a visão de que tanto respostas auto-imunes como as determinadas pelo parasita em conjunto com citocinas e quimiocinas inflamatórias participam da geração das lesões cardíacas típicas da CCC. A presente revisão tem como objetivo sumarizar os recentes avanços no entendimento da imunopatogênese da Cardiomiopatia Chagásica Crônica.

Autoimmunity in Chagas' heart disease

The time scale dissociation between high parasitemia and tissue pathology allied to the absence of parasites in the heart lesions of chronic Chagas' disease cardiopathy, casted doubt on the direct participation of Trypanosoma cruzi in tissue lesions. Moreover, the heart tissue lesions in chronic Chagas' disease cardiopathy are associated to an inflammatory mononuclear cell infiltrate, presumably the ultimate effectors of tissue damage. It has been hypothesized that the inflammatory cell infiltrate could mediate a delayed hypersensitivity process directed to the heart tissue components, an autoimmune response triggered by immunological cross-reactivity in the course of a protective immune response against some T. cruzi antigen homologous to heart proteins. However, little is known about the efector role of the T cells in the infiltrate, or about the nature of the antigen that lead to their accumulation in tissue. In this paper, we will review the published evidence on autoimmunity and immunological cross-reactivity between T cruzi and the mammalian host, along with data generated in our laboratory. The definition of the precise role played by autoimmunity in the pathogenesis of Chagas' disease cardiopathy may have important consequences both for immunoprophylaxis and for the therapeutic approach of chronic Chagas' disease.

Increased plasma levels of tumor necrosis factor-alpha in asymptomatic/"indeterminate" and Chagas disease cardiomyopathy patients

We compared plasma tumor necrosis factor-alpha (TNF-alpha) levels among asymptomatic/"indeterminate" Chagas disease patients (ASY) and patients across the clinical spectrum of chronic Chagas disease cardiomyopathy (CCC). Idiopathic dilated cardiomyopathy (DCM) patients and normal controls (NC) were included as controls. ASY Chagas disease patients had significantly higher plasma TNF-alpha levels than NC. TNF-alpha levels among severe CCC patients with significant left ventricular (LV) dysfunction were similar to those of DCM patients, showing average 2-fold higher levels than CCC patients without LV dysfunction and ASY patients, and 8-fold higher levels than NC. In Chagas disease, chronic TNF-a production prior to heart failure may play a role in CCC progression

Disfunção ventricular esquerda em hamsters com doença de Chagas tratados com etanercept / Left ventricular dysfunction in hamsters with Chagas disease and treated with etanercept

Objetivo: a doença afeta mais de 10 milhões de pessoas na América Latina. Leva a cardiomiopatia dilatada inflamatória em 30% dos pacientes como conseqüência tardia da infecção pelo protozoário Trypanosoma cruzi, com pior prognóstico que as outras cardiomiopatias dilatadas. estudos prévios mostram aumento dos níveis circulantes do fator de necrose tumoral-alfa (TNF-x) em pacientes com cardiomiopatia chagásica crônica. Assim, o objetivo do presente trabalho foi avaliar efeito do bloqueio do TNF-x com Etanercept na função ventricular esquerda em hamsters sírios cronicamente infectados pelo T. cruzi...

Análise proteômica das principais proteínas antigênicas do veneno da vespa agelaia pallipes / Proteomic analysis of mayor antigenic proteins from agelaia pallipes venom

Objetivo: alguns pacientes alérgicos ao veneno de vespas apresentam pesquisa negativa de IgE específica com os extratos disponíveis. Para investigar esta falta de reação cruzada, este estudo pretende caracterizar os antígenos principais do veneno de uma das espécies encontradas no Brasil, a Agelaia pallipes, utilizando a análise proteômica. Método: realizamos eletroforese bidimensional com veneno de Agelaia pallipes. Na primeira dimensão utilizamos tiras de gel de 7 cm com gradiente de pH de 3.0 -10.0, e na segunda SDS-PAGE 15%. Com géis feitos em duplicata, o primeiro foi transferido para nitrocelulose e incubado com o soro de paciente sensibilizado (diluição 1:5). A imunodetecção foi realizada com anti-IgE humana biotinilada e ECL (Enhanced Chemiluminescence). No segundo gel, corado Coomassie, os spots correspondentes às proteínas reconhecidas pela IgE através do immuniblotting foram processados e analisados no espectrometro de massa do tipo MALDI-ToF. A identificação foi obtida por PMF - Peptide Mass Fingerprinting. Resultados: a eletroforese bidimensional com o veneno da Agelaia pallipes evidenciou várias proteínas com peso molecular (PM) abaixo de 20kDa. Com immunoblotting foram detectadas proteínas reconhecidas pela IgE com PM entre 20 e 38 kDa. Pela análise proteômica, estas proteínas foram identificadas principalmente como antígeno 5 e serino-proteases. Conclusão: este é o primeiro trabalho a identificar alérgenos de vespas neotropicais com análise proteômica. Além do antígeno 5, identificamos serino-proteases que apenas recentemente foram citadas neste tipo de amostra biológica, mostrando semelhança parcial entre estas proteínas de venenos de vertebrados (serpentes). Nosso projeto futuro será o sequenciamento das amostras.

Auto-imunidade na doença de Chagas / Autoimmunity in Chagas'disease

A dissociaçäo na escala do tempo que ocorre entre o quadro de infecçäo aguda por Trypanosoma cruzi, que cursa com parasitemia e profusa parasitose tecidual, semanas após o contágio, e a cardiopatia chagásica crônica pobre em formas teciduais do parasita que se manifesta décadas mais tarde, em vigência de parasitemia indectável, levantou dúvidas sobre a participaçäo direta do T. cruzi na cardiopatia chagásica crônica. Surgiu a hipótese de que o dano ao miocárdio seria secundário a um processo de hipersensibilidade retardada dirigido ao próprio tecido cardíaco, mediado pelo infiltrado inflamatório linfomononuclear universalmente associado às lesöes. Segundo essa hipótese, a resposta auto-imune dirigida ao miocárdio seria causada por reaçäo cruzada desencadeada durante a resposta imune dirigida ao miocárdio seria causada por reaçäo cruzada desencadeada durante a resposta imune de defesa contra algum antígeno de T.cruzi homólogo a estruturas cardíacas. Entretanto, pouco se sabe sobre o papel efetor dos linfócitos do infiltrado, ou do antígeno desencadeador de seu acúmulo no tecido cardíaco. Neste trabalho, revisaremos a literatura publicada sobre auto-imunidade ou de reaçäo cruzada imunológica na doença de Chagas, e discutiremos os resultados obtdiso em nosso laboratório. A definiçäo do papel da auto-imunidade na patogênese pode ter importantes repercussöes para a imunoprofilaxia e a abordagem terapêutica da cardiopatia chagásica crônica.