Resposta imune a doenças infecciosas

The interaction between pathogens and immune system occurs in a dynamic way with sophisticate mechanisms of evasion and immune control of infection, respectively. Therefore, it is a sine qua non condition to understand the complexity of this relationship in order to developed new strategies for infection control. Although the immune system has specialized mechanisms to control infection, different conditions involved in the interaction between immune system and pathogens can or cannot determine the development of disease. Interestingly, a protective immune response against one kind of parasite may not be protective against another. So, each pathogen presents a specific way of interaction with the immune system. The development of research in this area has contributed with a better comprehension of the immune system and pathogen relationship and opened perspective in improving the treatment with the development of new drugs and/or vaccines.A interação do sistema imune com os agentes infecciosos ocorre de uma maneira dinâmica, com mecanismos de controle da infecção e de escape sofisticados. A compreensão dessa complexidade é condição sine qua non para que se estabeleça uma ação total no controle dessas infecções. Embora a resposta imune desenvolvida para controle das diferentes infecções apresente certas particularidades, em geral, apresentam também mecanismos comuns. A priori os mecanismos podem ser redundantes, no entanto existe uma gama de sutilezas entre a interação hospedeiro-parasita que define o estabelecimento ou não de doença. Por outro lado, não se pode deixar de alertar que melhores condições de saneamento básico diminuiriam a incidência de inúmeras doenças. A classificação de uma resposta imune protetora tem que ser avaliada sempre em relação ao tipo de agente agressor, pois um mecanismo protetor conta um vírus pode não ser essencial contra uma bactéria extracelular. De qualquer forma, o avanço na pesquisa com diferentes patógenos tem contribuído para uma melhor compreensão da resposta imune decorrente da interação entre o hospedeiro e parasita o que pode resultar no desenvolvimento de novas drogas e vacinas

Endocytosis of DNA-Hsp65 Alters the pH of the Late Endosome/Lysosome and Interferes with Antigen Presentation

BACKGROUND: Experimental models using DNA vaccine has shown that this vaccine is efficient in generating humoral and cellular immune responses to a wide variety of DNA-derived antigens. Despite the progress in DNA vaccine development, the intracellular transport and fate of naked plasmid DNA in eukaryotic cells is poorly understood, and need to be clarified in order to facilitate the development of novel vectors and vaccine strategies. METHODOLOGY AND PRINCIPAL FINDINGS: Using confocal microscopy, we have demonstrated for the first time that after plasmid DNA uptake an inhibition of the acidification of the lysosomal compartment occurs. This lack of acidification impaired antigen presentation to CD4 T cells, but did not alter the recruitment of MyD88. The recruitment of Rab 5 and Lamp I were also altered since we were not able to co-localize plasmid DNA with Rab 5 and Lamp I in early endosomes and late endosomes/lysosomes, respectively. Furthermore, we observed that the DNA capture process in macrophages was by clathrin-mediated endocytosis. In addition, we observed that plasmid DNA remains in vesicles until it is in a juxtanuclear location, suggesting that the plasmid does not escape into the cytoplasmic compartment. CONCLUSIONS AND SIGNIFICANCE: Taken together our data suggests a novel mechanism involved in the intracellular trafficking of plasmid DNA, and opens new possibilities for the use of lower doses of plasmid DNA to regulate the immune response

Functional Liposomal Configuration And Method For Obtaining Said Functional Liposomal Configuration

CONFIGURAÇÃO LIPOSSOMAL FUNCIONAL E PROCESSO DE OBTENÇÃO DE CONFIGURAÇÃO LIPOSSOMAL FUNCIONAL. A invenção apresenta um produto para terapia e vacinação, constituído de uma configuração lipossomal funcional contendo polinucleotídeos complexados preferencialmente na superfície externa de lipossomas do tipo DRVs, e o seu processo de produção. Essa configuração lipossomal funcional contendo polinucleotídeos possui propriedades de mucoadesão que permite a administração através de vias não invasivas como a via nasal, e é capaz de carrear eficientemente os polinucleotídeos até o interior das células, liberando-os no citoplasma. Essas propriedades potencializam maior eficiência de ação in vivo dos polinucleotídeos carreados, redução da concentração e frequência de doses, e, no caso da vacinação produz proteção contra a doença específica.BRPI0705630 (A2); BRPI0705630 (E2)A61K9/127C12N15/64BR2007PI05630A61K9/127C12N15/6

Detrimental effect of fungal 60-kDa heat shock protein on experimental Paracoccidioides brasiliensis Infection

The genus Paracoccidioides comprises species of dimorphic fungi that cause paracocci-dioidomycosis (PCM), a systemic disease prevalent in Latin America. Here, we investigated whether administration of native 60-kDa heat shock protein of P. brasiliensis (nPbHsp60) or its recombinant counterpart (rPbHsp60) affected the course of experimental PCM. Mice were subcutaneously injected with nPbHsp60 or rPbHsp60 emulsified in complete's Freund Adjuvant (CFA) at three weeks after intravenous injection of P. brasiliensis yeasts. Infected control mice were injected with CFA or isotonic saline solution alone. Thirty days after the nPbHsp60 or rPbHsp60 administration, mice showed remarkably increased fungal load, tissue inflammation, and granulomas in the lungs, liver, and spleen compared with control mice. Further, rPbHsp60 treatment (i) decreased the known protective effect of CFA against PCM and (ii) increased the concentrations of IL-17, TNF-alpha, IL-12, IFN-gamma, IL-4, IL-10, and TGF-beta in the lungs. Together, our results indicated that PbHsp60 induced a harmful immune response, exacerbated inflammation, and promoted fungal dissemination. Therefore, we propose that PbHsp60 contributes to the fungal pathogenesis119FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP2009/14777-1; 2013/12278-3; 2009/03235-3; 2012/08552-

The synergy between structural stability and DNA-binding controls the antibody production in EPC/DOTAP/DOPE liposomes and DOTAP/DOPE lipoplexes

We present a comparative study of the physico-chemical properties, in vitro cytotoxicity and in vivo antibody production of surface-complexed DNA in EPC/DOTAP/DOPE (50/25/25% molar) liposomes and DOTAP/DOPE (50/50% molar) lipoplexes. The study aims to correlate the biological behavior and structural properties of the lipid carriers. We used DNA-hsp65, whose naked action as a gene vaccine against tuberculosis has already been demonstrated. Additionally, surface-complexed DNA-hsp65 in EPC/DOTAP/DOPE (50/25/25% molar) liposomes was effective as a single-dose tuberculosis vaccine. The results obtained showed that the EPC inclusion stabilized the DOTAP/DOPE structure, producing higher melting temperature and lower zeta potential despite a close mean hydrodynamic diameter. Resemblances in morphologies were identified in both structures, although a higher fraction of loaded DNA was not electrostatically bound in EPC/DOTAP/DOPE. EPC also induced a striking reduction in cytotoxicity, similar to naked DNA-hsp65. The proper immune response lead to a polarized antibody production of the IgG2a isotype, even for the cytotoxic DOTAP/DOPE. However, the antibody production was detected at 15 and 30 days for DOTAP/DOPE and EPC/DOTAP/DOPE, respectively. Therefore, the in vivo antibody production neither correlates with the in vitro cytotoxicity, nor with the structural stability alone. The synergistic effect of the structural stability and DNA electrostatic binding upon the surface of structures account for the immunological effects. By adjusting the composition to generate proper packing and cationic lipid/DNA interaction, we allow for the optimization of liposome formulations for required immunization or gene therapy. In a specific manner, our results contribute to studies on the tuberculosis therapy and vaccination732175184FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO – FAPESPsem informaçã

Functional Liposomal Configuration And Method For Obtaining Said Functional Liposomal Configuration

This invention provides a product for therapy and vaccination, constituted by a functional liposomal configuration containing complexes polynucleotides preferably on the outer surface of dehydrated-rehydrated liposomes (DRV), and the production process thereof. This functional liposomal configuration containing polynucleotides has mucoadhesion properties which allow administration by non-invasive routes, such as the nasal route, and is able to efficiently carry the polynucleotides to the interior of cells, releasing them in the cytoplasm. These properties allows for higher in vivo action efficiency of the carried polynucleotides, reduced concentration and dose frequency, and, in the case of vaccination, give protection against the specific disease. The present technology also develops a liposomal configuration carrying polynucleotides, which encode the protein HSP65, for prevention and treatment of tuberculosis.WO2009073941 (A2); WO2009073941 (A3)A61K9/127A61K9/50A61K9/51A61K31/7088WO2008BR00387A61K9/127A61K9/50A61K9/51A61K31/7088US6110745 (A)US2004043954 (A1)WO2006086330 (A2

The synergy between structural stability and DNA-binding controls the antibody production in EPC/DOTAP/DOPE liposomes and DOTAP/DOPE lipoplexes

We present a comparative study of the physico-chemical properties, in vitro cytotoxicity and in vivo antibody production of surface-complexed DNA in EPC/DOTAP/DOPE (50/25/25% molar) liposomes and DOTAP/DOPE (50/50% molar) lipoplexes. The study aims to correlate the biological behavior and structural properties of the lipid carriers. We used DNA-hsp65, whose naked action as a gene vaccine against tuberculosis has already been demonstrated. Additionally, surface-complexed DNA-hsp65 in EPC/DOTAP/DOPE (50/25/25% molar) liposomes was effective as a single-dose tuberculosis vaccine. The results obtained showed that the EPC inclusion stabilized the DOTAP/DOPE structure, producing higher melting temperature and lower zeta potential despite a close mean hydrodynamic diameter. Resemblances in morphologies were identified in both structures, although a higher fraction of loaded DNA was not electrostatically bound in EPC/DOTAP/DOPE. EPC also induced a striking reduction in cytotoxicity, similar to naked DNA-hsp65. The proper immune response lead to a polarized antibody production of the IgG2a isotype, even for the cytotoxic DOTAP/DOPE. However, the antibody production was detected at 15 and 30 days for DOTAP/DOPE and EPC/DOTAP/DOPE, respectively. Therefore, the in vivo antibody production neither correlates with the in vitro cytotoxicity, nor with the structural stability alone. The synergistic effect of the structural stability and DNA electrostatic binding upon the surface of structures account for the immunological effects. By adjusting the composition to generate proper packing and cationic lipid/DNA interaction, we allow for the optimization of liposome formulations for required immunization or gene therapy. In a specific manner, our results contribute to studies on the tuberculosis therapy and vaccination. (C) 2009 Elsevier B.V. All rights reserved.Fundacao de Auxilio a Pesquisa de Sao Paulo (FAPESP

HSP65 DNA as therapeutic strategy to treat experimental paracoccidioidomycosis

The conventional treatment for paracoccidioidomycosis, the most prevalent mycosis in Latin America, involves long periods of therapy resulting in sequels and high frequency of relapses. The search for new alternatives of treatment is necessary. Previously, we have demonstrated that the hsp65 gene from Mycobacterium leprae shows prophylactic effects against murine paracoccidioidomycosis. Here, we tested the DNAhsp65 immunotherapy in BALB/c mice infected with Paracoccidioides brasiliensis, the agent of paracoccidioidomycosis. We observed an increase of Th1 cytokines accompanied by a reduction in fungal burden and pulmonary injury. These results provide new prospects for immunotherapy of paracoccidioidomycosis and other mycoses. (C) 2009 Elsevier Ltd. All rights reserved.Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior (CAPES)Conselho Nacional de Desenvolvimento Cientifico a Tecnologico (CNPq

Detrimental Effect of Fungal 60-kDa Heat Shock Protein on Experimental <i>Paracoccidioides brasiliensis</i> Infection

<div><p>The genus <i>Paracoccidioides</i> comprises species of dimorphic fungi that cause paracoccidioidomycosis (PCM), a systemic disease prevalent in Latin America. Here, we investigated whether administration of native 60-kDa heat shock protein of <i>P</i>. <i>brasiliensis</i> (nPbHsp60) or its recombinant counterpart (rPbHsp60) affected the course of experimental PCM. Mice were subcutaneously injected with nPbHsp60 or rPbHsp60 emulsified in complete’s Freund Adjuvant (CFA) at three weeks after intravenous injection of <i>P</i>. <i>brasiliensis</i> yeasts. Infected control mice were injected with CFA or isotonic saline solution alone. Thirty days after the nPbHsp60 or rPbHsp60 administration, mice showed remarkably increased fungal load, tissue inflammation, and granulomas in the lungs, liver, and spleen compared with control mice. Further, rPbHsp60 treatment <i>(i)</i> decreased the known protective effect of CFA against PCM and <i>(ii)</i> increased the concentrations of IL-17, TNF-α, IL-12, IFN-γ, IL-4, IL-10, and TGF-β in the lungs. Together, our results indicated that PbHsp60 induced a harmful immune response, exacerbated inflammation, and promoted fungal dissemination. Therefore, we propose that PbHsp60 contributes to the fungal pathogenesis.</p></div