Proteção de bovinos contra Haemonchus placei e Haemonchus contortus após imunização com antígenos oriundos da membrana intestinal de H. contortus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Estudo de potenciais inibidores de inibidores de infecção causada por Vírus Sincicial Respiratório em cultura de célula

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Caracterização estrutural e das interações entre a Proteína G do hRSV e potenciais inibidores

Pós-graduação em Microbiologia - IBILCE

Estudo do perfil proteico nas membranas de espermatozóides congelados de caprinos

Pós-graduação em Medicina Veterinária - FMVZ

Efeitos da sexagem por citometria de fluxo sobre a composição protéica da membrana de espermatozóides bovinos

Pós-graduação em Medicina Veterinária - FMVZ

Vaccination of grazing calves with antigens from the intestinal membranes of Haemonchus contortus: effects against natural challenge with Haemonchus placei and Haemonchus similis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Fator de crescimento do endotélio vascular e do transportador de glicose 1 em cães com ceratose actínica e carcinoma de células escamosas

Pós-graduação em Medicina Veterinária - FCAV

Systematic analysis of protein-detergent complexes applying dynamic light scattering to optimize solutions for crystallization trials

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Attempts to vaccinate ewes and their lambs against natural infection with Haemonchus contortus in a tropical environment

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Effects of Yersinia enterocolitica O:3 derivatives on B lymphocyte activation in vivo

The potential sequelae of intestinal infection with Yersinia enterocolitica include reactive arthritis, erythema nodosum, Reiter's syndrome and other autoimmune diseases. The role of the immune response in the pathogenesis of these diseases has not been fully defined, but autoimmune manifestations may be a consequence of the increase in autoantibodies as a result of polyclonal B-cell activation induced by Yersinia. We investigated the effects of Y enterocolitica 0:3 derivatives on B lymphocyte activation in vivo. Groups of five specific pathogen free (SPF) Swiss mice were inoculated with bacterial cell extract, Yersinia outermembrane proteins (Yops) or lipopolysaccharide (LPS) obtained from Y enterocolitica 0:3 and their immunoglobulin-secreting spleen cells were detected by isotype-specific protein A plaque assay. The presence of specific anti-Yersinia antibodies and autoantibodies was determined in mouse sera by ELISA. In all experiments a marked increase in the number of secretory cells of different isotypes was observed as early as the third day after inoculation. IgG and IgM anti-Yersinia antibodies were detected in the sera of all inoculated mice, and autoantibodies against myosin in the sera of those inoculated with bacterial cell extract. The sera from animals stimulated with LPS reacted with myelin, actin and laminin, while the sera from mice inoculated with Yops reacted with myelin, thyroglobulin and cardiolipin. These results suggest that SPF Swiss mice inoculated with any one of the Y enterocolitica derivatives tested exhibited polyclonal activation of B lymphocytes as a result of stimulation by various bacterial components and not only LPS stimulation.

Funcionalidade das proteínas EF-Tu e 14-3-3 na virulência de Paracoccidioides brasiliensis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Immune receptors and adhesion molecules in human pulmonary leptospirosis

Pulmonary involvement in leptospirosis has been increasingly reported in the last 20 years, being related to the severity and mortality of the disease. The pathogenesis of pulmonary hemorrhage in leptospirosis is not understood. Lung endothelial cells have been proposed as targets in the pathogenesis of lung involvement in leptospirosis through the activation of Toll-like receptor 2 or the complement system, which stimulates the release of cytokines that lead to the activation of adhesion molecules. The aim of this study was to investigate the involvement of immune pathways and of the intercellular and vascular cell adhesion molecules (intercellular adhesion molecule and vascular cell adhesion molecule, respectively) in the lungs of patients with pulmonary involvement of leptospirosis. We studied the lungs of 18 patients who died of leptospirosis and compared them with 2 groups of controls: normal and noninfectious hemorrhagic lungs. Using immunohistochemistry and image analysis, we quantified the expression of the C3a anaphylatoxin receptor, intercellular adhesion molecule, vascular cell adhesion molecule, and Toll-like receptor 2 in small pulmonary vessels and in the alveolar septa. There was an increased expression of intercellular adhesion molecule (P <.03) and C3a anaphylatoxin receptor (P <.008) in alveolar septa in the leptospirosis group compared with the normal and hemorrhagic controls. In the vessels of the leptospirosis group, there was an increased expression of intercellular adhesion molecule (P=.004), vascular cell adhesion molecule (P=.030), and Toll-like receptor 2 (P=.042) compared with the normal group. Vascular cell adhesion molecule expression in vessels was higher in the leptospirosis group compared with the hemorrhagic group (P=.015). Our results indicate that immune receptors and adhesion molecules participate in the phenomena leading to pulmonary hemorrhage in leptospirosis. (C) 2012 Elsevier Inc. All rights reserved.

A novel protein refolding protocol for the solubilization and purification of recombinant peptidoglycan-associated lipoprotein from Xylella fastidiosa overexpressed in Escherichia coil

Xylella fastidiosa is a Gram-negative xylem-limited plant pathogenic bacterium responsible for several economically important crop diseases. Here, we present a novel and efficient protein refolding protocol for the solubilization and purification of recombinant X. fastidiosa peptidoglycan-associated lipoprotein (XfPal). Pal is an outer membrane protein that plays important roles in maintaining the integrity of the cell envelope and in bacterial pathogenicity. Because Pal has a highly hydrophobic N-terminal domain, the heterologous expression studies necessary for structural and functional protein characterization are laborious once the recombinant protein is present in inclusion bodies. Our protocol based on the denaturation of the XfPal-enriched inclusion bodies with 8 M urea followed by buffer-exchange steps via dialysis proved effective for the solubilization and subsequent purification of XfPal, allowing us to obtain a large amount of relatively pure and folded protein. In addition, XfPal was biochemically and functionally characterized. The method for purification reported herein is valuable for further research on the three-dimensional structure and function of Pal and other outer membrane proteins and can contribute to a better understanding of the role of these proteins in bacterial pathogenicity, especially with regard to the plant pathogen X. fastidiosa. (C) 2012 Elsevier Inc. All rights reserved.

Differential expression of toll-like receptor mRNAs in recurrent aphthous ulceration

BACKGROUND: Toll-like receptors (TLR) are membrane proteins that recognize conserved molecules derived from bacterial, viral, fungal or host tissues. They are responsible for promoting the production of cytokines and chemokines, increasing the expression of costimulatory molecules and influencing the T Helper response (Th) toward either a Th1 or Th2 profile, thereby modulating the regulatory T cell response and controlling the integrity of the epithelial barrier. The key factors responsible for increased susceptibility to recurrent aphthous ulceration (RAU) are unclear, and because TLRs are involved in both immune regulation and control of the epithelial barrier, a deficiency in TLR activity is likely to cause increased susceptibility. METHODS: We investigated the gene expression of TLRs one through 10 in tissue samples and peripheral blood mononuclear cells (PBMC) of RAU patients in comparison to healthy controls using real-time quantitative reverse transcription PCR. RESULTS: The analysis of mRNA expression levels in oral lesion showed significant (P < 0.01) overexpression of the TLR2(similar to 6-fold) gene and decreased expression of the TLR3 (similar to 5-fold) and TLR5 (similar to 6-fold) genes in comparison with healthy oral mucosa. The analysis of mRNA expression in PBMC indicated a down-regulation of TLR5 gene expression in the cells from RAU patients (P < 0.05; similar to 2-fold). CONCLUSION: Our results support the hypothesis that a subset of RAU patients has fewer TLR expression that have been tentatively implicated in antiinflammatory effects. This derangement of TLR gene expression may cause an overlay exuberant inflammation reaction in situations where normal individuals are resistant. J Oral Pathol Med (2012) 41: 8085

OmpL1 Is an Extracellular Matrix- and Plasminogen-Interacting Protein of Leptospira spp.

Leptospirosis is a zoonosis with multisystem involvement caused by pathogenic strains of the genus Leptospira. OmpL1 is an outer membrane protein of Leptospira spp. that is expressed during infection. In this work, we investigated novel features of this protein. We describe that OmpL1 is a novel leptospiral extracellular matrix (ECM)-binding protein and a plasminogen (PLG) receptor. The recombinant protein was expressed in Escherichia coli BL21(DE3) Star/pLysS as inclusion bodies, refolded, and purified by metal-chelating chromatography. The protein presented a typical beta-strand secondary structure, as evaluated by circular dichroism spectroscopy. The recombinant protein reacted with antibodies in serum samples from convalescent leptospirosis patients with a high specificity compared to serum samples from individuals with unrelated diseases. These data strengthen the usefulness of OmpL1 as a diagnostic marker of leptospirosis. The characterization of the immunogenicity of recombinant OmpL1 in inoculated BALB/c mice showed that the protein has the capacity to elicit humoral and cellular immune responses, as denoted by high antibody titers and the proliferation of lymphocytes. We demonstrate that OmpL1 has the ability to mediate attachment to laminin and plasma fibronectin, with KD (equilibrium dissociation constant) values of 2,099.93 +/- 871.03 nM and 1,239.23 +/- 506.85 nM, respectively. OmpL1 is also a PLG receptor, with a KD of 368.63 +/- 121.23 nM, capable of generating enzymatically active plasmin. This is the first report that shows and characterizes OmpL1 as an ECM-interacting and a PLG-binding protein of Leptospira spp. that may play a role in bacterial pathogenesis when expressed during infection.