Expressão gênica de interferon-gama, fator de necrose tumoral alfa e interleucinas 2 e 5 no baço de gatos naturalmente infectados por Leishmania spp: Karina Yukie Hirata. -

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

Francisella noatunensis orientalis em tilápias (Oreochromis niloticus) cultivadas em tanques-rede na bacia hidrográfica do rio Araguari-Minas Gerais

Pós-graduação em Medicina Veterinária - FMVZ 33004064022P3

Francisella noatunensis orientalis em tilápias (Oreochromis niloticus) cultivadas em tanques-rede na bacia hidrográfica do rio Araguari-Minas Gerais

Pós-graduação em Medicina Veterinária - FMVZ

Influence of biliary anastomosis on recovery from secondary biliary cirrhosis

Objective The influence of choledochoduodenostomy and choledochojejunostomy on the repair of hepatic lesions secondary to biliary obstruction is not well known. The aim of the present study was to compare the effects of choledochoduodenostomy and choledochojejunostomy on the recovery of these lesions in rats with biliary obstruction. Methods Rats subjected to 4 weeks of biliary obstruction underwent choledochoduodenostomy (n=10) or choledochojejunostomy (n=10). The following variables were measured: total bilirubin, alkaline phosphatase, aminotransferases, and albumin. Hepatic mitochondrial energy metabolism was evaluated by calculating the respiratory control ratio and the oxidative phosphorylation index. Hepatic morphometry was used to estimate the mass of the hepatocytes, bile ducts, and fibrosis, as well as the hepatic stellate cell count. Results After choledochoduodenostomy and choledochojejunostomy, there was a regression in cholestasis and a reduction in the oxidative phosphorylation index. However, the total bilirubin, alkaline phosphatase, albumin, and respiratory control ratio values improved only after choledochojejunostomy. The mass of the liver, spleen, and fibrosis was reduced after both choledochoduodenostomy and choledochojejunostomy, but the number of hepatic stellate cells increased. After choledochojejunostomy, the hepatic mass recovered completely, and the spleen mass was significantly reduced compared with that after choledochoduodenostomy. After both choledochoduodenostomy and choledochojejunostomy, enterobiliary reflux, biliary contamination, and an exacerbation in hepatic inflammation developed. Conclusion Choledochojejunostomy was more effective than choledochoduodenostomy, but both techniques induced enterobiliary reflux and biliary contamination, which may explain the maintenance of hepatic alterations, especially after choledochoduodenostomy. Eur J Gastroenterol Hepatol 24: 1039-1050 (C) 2012 Wolters Kluwer Health vertical bar Lippincott Williams & Wilkins.

Anti-CD25 Treatment Depletes Treg Cells and Decreases Disease Severity in Susceptible and Resistant Mice Infected with Paracoccidioides brasiliensis

Regulatory T (Treg) cells are fundamental in the control of immunity and excessive tissue pathology. In paracoccidioidomycosis, an endemic mycosis of Latin America, the immunoregulatory mechanisms that control the progressive and regressive forms of this infection are poorly known. Due to its modulatory activity on Treg cells, we investigated the effects of anti-CD25 treatment over the course of pulmonary infection in resistant (A/J) and susceptible (B10.A) mice infected with Paracoccidioides brasiliensis. We verified that the resistant A/J mice developed higher numbers and more potent Treg cells than susceptible B10.A mice. Compared to B10.A cells, the CD4(+)CD25(+)Foxp3(+) Treg cells of A/J mice expressed higher levels of CD25, CTLA4, GITR, Foxp3, LAP and intracellular IL-10 and TGF-beta. In both resistant and susceptible mice, anti-CD25 treatment decreased the CD4(+)CD25(+)Foxp3(+) Treg cell number, impaired indoleamine 2,3-dioxygenase expression and resulted in decreased fungal loads in the lungs, liver and spleen. In A/J mice, anti-CD25 treatment led to an early increase in T cell immunity, demonstrated by the augmented influx of activated CD4(+) and CD8(+) T cells, macrophages and dendritic cells to the lungs. At a later phase, the mild infection was associated with decreased inflammatory reactions and increased Th1/Th2/Th17 cytokine production. In B10.A mice, anti-CD25 treatment did not alter the inflammatory reactions but increased the fungicidal mechanisms and late secretion of Th1/Th2/Th17 cytokines. Importantly, in both mouse strains, the early depletion of CD25(+) cells resulted in less severe tissue pathology and abolished the enhanced mortality observed in susceptible mice. In conclusion, this study is the first to demonstrate that anti-CD25 treatment is beneficial to the progressive and regressive forms of paracoccidioidomycosis, potentially due to the anti-CD25-mediated reduction of Treg cells, as these cells have suppressive effects on the early T cell response in resistant mice and the clearance mechanisms of fungal cells in susceptible mice.

Experimental infection of suckling mice by subcutaneous inoculation with Oropouche virus

Oropouche virus, of the family Bunyaviridae, genus Orthobunyavirus, serogroup Simbu, is an important causative agent of arboviral febrile illness in Brazil. An estimated 500,000 cases of Oropouche fever have occurred in Brazil in the last 30 years, with recorded cases also in Panama, Peru, Suriname and Trinidad. We have developed an experimental model of Oropouche virus infection in neonatal BALB/c mouse by subcutaneous inoculation. The vast majority of infected animals developed disease on the 5th day post infection, characterized mainly by lethargy and paralysis, progressing to death within 10 days. Viral replication was documented in brain cells by in situ hybridization, immunohistochemistry and virus titration. Multi-step immunohistochemistry indicated neurons as the main target cells of OROV infection. Histopathology revealed glial reaction and astrocyte activation in the brain and spinal cord, with neuronal apoptosis. Spleen hyperplasia and mild meningitis were also found, without viable virus detected in liver and spleen. This is the first report of an experimental mouse model of OROV infection, with severe involvement of the central nervous system, and should become useful in pathogenesis studies, as well as in preclinical testing of therapeutic interventions for this emerging pathogen. (c) 2012 Elsevier B.V. All rights reserved.

Endostatin gene therapy stimulates upregulation of ICAM-1 and VCAM-1 in a metastatic renal cell carcinoma model

One of the greatest challenges in urological oncology is renal cell carcinoma (RCC), which is the third leading cause of death in genitourinary cancers. RCCs are highly vascularized and respond positively to antiangiogenic therapy. Endostatin (ES) is a fragment of collagen XVIII that possesses antiangiogenic activity. In this study, we examined the potential of ES-based antiangiogenic therapy to activate tumor-associated endothelial cells in metastatic RCC (mRCC). Balb/c-bearing Renca cells were treated with NIH/3T3-LendSN or, as a control, with NIH/3T3-LXSN cells. The T-cell subsets and lymphocyte populations of tumors, mediastinal lymph nodes and the spleen were assessed by flow cytometry. The expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) was assessed by real-time PCR, flow cytometry and immunohistochemistry analysis. ES gene therapy led to an increase in the percentage of infiltrating CD4-interferon (IFN)-gamma cells (P<0.05), CD8-IFN-gamma cells (P<0.01) and CD49b-tumor necrosis factor-alpha cells (P<0.01). In addition, ES therapy caused an increase at the mRNA level of ICAM-1 (1.4-fold; P<0.01) and VCAM-1 (1.5-fold) (control vs treated group; P<0.001). Through flow cytometry, we found a significant increase in the CD34/ICAM-1 cells (8.1-fold; P<0.001) and CD34/VCAM-1 cells (1.6-fold; P<0.05). ES gene therapy induced a significant increase in both T CD4 and CD8 cells in the lymph nodes and the spleen, suggesting that ES therapy may facilitate cell survival or clonal expansion. CD49b cells were also present in increased quantities in all of these organs. In this study, we demonstrate an antitumor inflammatory effect of ES in an mRCC model, and this effect is mediated by an increase in ICAM-1 and VCAM-1 expression in tumor-associated endothelial cells.

Chronic cold stress in mice induces a regulatory phenotype in macrophages: Correlation with increased 11 beta-hydroxysteroid dehydrogenase expression

Susceptibility to infections, autoimmune disorders and tumor progression is strongly influenced by the activity of the endocrine and nervous systems in response to a stressful stimulus. When the adaptive system is switched on and off efficiently, the body is able to recover from the stress imposed. However, when the system is activated repeatedly or the activity is sustained, as during chronic or excessive stress, an allostatic load is generated, which can lead to disease over long periods of time. We investigated the effects of chronic cold stress in BALB/c mice (4 degrees C/4 h daily for 7 days) on functions of macrophages. We found that chronic cold stress induced a regulatory phenotype in macrophages, characterized by diminished phagocytic ability, decreased TNF-alpha and IL-6 and increased IL-10 production. In addition, resting macrophages from mice exposed to cold stress stimulated spleen cells to produce regulatory cytokines, and an immunosuppressive state that impaired stressed mice to control Trypanosoma cruzi proliferation. These regulatory effects correlated with an increase in macrophage expression of 11 beta-hydroxysteroid dehydrogenase, an enzyme that converts inactive glucocorticoid into its active form. As stress is a common aspect of modern life and plays a role in the etiology of many diseases, the results of this study are important for improving knowledge regarding the neuro-immune-endocrine interactions that occur during stress and to highlight the role of macrophages in the immunosuppression induced by chronic stress. (C) 2011 Elsevier Inc. All rights reserved.

ELECTROCARDIOGRAPHIC PARAMETERS OF CAPTIVE TUFTED CAPUCHINS (CEBUS APELLA) UNDER CHEMICAL IMMOBILIZATION

This study presents the electrocardiogram findings from 97 captive tufted capuchin monkeys (Cebus apella) at the Sao Paulo Zoo (Sao Paulo, Brazil) while under ketamine anesthesia. The results did not differ greatly from data of domestic carnivores or other studied primate species. The most common rhythm recorded was normal sinus rhythm, followed by normal sinus rhythm with wandering pacemaker. Electrical axis varied from 0 degrees to -150 degrees but was most commonly between +60 degrees and +90 degrees. QRS complexes were predominantly positive in leads DI, DII, DIII, and AVF. These findings allow for the recognition of abnormal rhythms in these primate species and can contribute to future investigations into the cardiovascular diseases routinely diagnosed in primates and humans.

Identification of Leishmania infantum chagasi proteins in urine of patients with visceral leishmaniasis: a promising antigen discovery approach of vaccine candidates

Visceral leishmaniasis (VL) is a serious lethal parasitic disease caused by Leishmania donovani in Asia and by Leishmania infantum chagasi in southern Europe and South America. VL is endemic in 47 countries with an annual incidence estimated to be 500 000 cases. This high incidence is due in part to the lack of an efficacious vaccine. Here, we introduce an innovative approach to directly identify parasite vaccine candidate antigens that are abundantly produced in vivo in humans with VL. We combined RP-HPLC and mass spectrometry and categorized three L. infantum chagasi proteins, presumably produced in spleen, liver and bone marrow lesions and excreted in the patients urine. Specifically, these proteins were the following: Li-isd1 (XP_001467866.1), Li-txn1 (XP_001466642.1) and Li-ntf2 (XP_001463738.1). Initial vaccine validation studies were performed with the rLi-ntf2 protein produced in Escherichia coli mixed with the adjuvant BpMPLA-SE. This formulation stimulated potent Th1 response in BALB/c mice. Compared to control animals, mice immunized with Li-ntf2+ BpMPLA-SE had a marked parasite burden reduction in spleens at 40 days post-challenge with virulent L. infantum chagasi. These results strongly support the proposed antigen discovery strategy of vaccine candidates to VL and opens novel possibilities for vaccine development to other serious infectious diseases.

Histologia e ultraestrutura do rim e rim cefálico do pacu

O pacu, Piaractus mesopotamicus, é um teleósteo da Família Characidae, intensivamente cultivado no Brasil devido sua rusticidade, crescimento rápido e fácil adaptação. O conhecimento morfológico dos sistemas corpóreos, incluído órgãos linfóide, se faz necessário, para uma melhor produção no cultivo de peixes, fornecendo subsídios na manutenção dos estoques. O objetivo deste estudo foi descrever morfologicamente o rim e rim cefálico de Piaractus mesopotamicus, analisando os perfis celulares de cada órgão com o uso de microscopia de luz e microscopia eletrônica de transmissão. O resultado da análise macroscópica mostrou que a localização do rim e rim cefálico são as mesmas encontradas na maioria dos teleósteos. O rim apresentou uma forma em "H", onde a região média se expandia sobre a bexiga natatória. O rim cefálico se apresentou como uma dilatação na região cranial do rim, mostrando-se bem visível. Na microscopia eletrônica de transmissão também foram observadas similaridades ultraestruturais com outros teleósteos. Observando nossos resultados concluímos que histologicamente e ultraestruturalmente, os órgãos linfóides rim e rim cefálico de Piaractus mesopotamicus são similares aos de outros teleósteos.

Imunocompetent Mice Model for Dengue Virus Infection

Dengue fever is a noncontagious infectious disease caused by dengue virus (DENV). DENV belongs to the family Flaviviridae, genus Flavivirus, and is classified into four antigenically distinct serotypes: DENV-1, DENV-2, DENV-3, and DENV-4. The number of nations and people affected has increased steadily and today is considered the most widely spread arbovirus (arthropod-borne viral disease) in the world. The absence of an appropriate animal model for studying the disease has hindered the understanding of dengue pathogenesis. In our study, we have found that immunocompetent C57BL/6 mice infected intraperitoneally with DENV-1 presented some signs of dengue disease such as thrombocytopenia, spleen hemorrhage, liver damage, and increase in production of IFN gamma and TNF alpha cytokines. Moreover, the animals became viremic and the virus was detected in several organs by real-time RT-PCR. Thus, this animal model could be used to study mechanism of dengue virus infection, to test antiviral drugs, as well as to evaluate candidate vaccines.

Morfologia e topografia do fígado e pâncreas de emas Rhea americana

As emas são aves ratitas nativas do continente sul americano, são consideradas aves primitivas do ponto de vista filogenético que constituem um grupo altamente especializado. Este estudo buscou caracterizar macro e microscopicamente o fígado e pâncreas de emas. O material foi coletado no Centro de Multiplicação de Animais Silvestres (CEMAS), na cidade de Mossoró-RN, Brasil, (Registro IBAMA n° 14.78912). Utilizaram-se 20 animais jovens com idade entre dois e seis meses independente do sexo. Em emas, o fígado se relacionava cranialmente com o ápice do coração, dorsalmente com os pulmões, esôfago e o proventrículo gástrico, caudalmente, com o ventrículo gástrico, o baço, o duodeno e parte do jejuno. Apresentava coloração vermelha escura e possuía apenas dois lobos, sendo o direito ligeiramente menor que o esquerdo. Histologicamente, era revestido por uma cápsula de tecido conjuntivo delgada e cada lóbulo hepático pôde ser identificado pela presença evidente de veias centrais, com muitos sinusoides comunicando-se com elas. O pâncreas, ventralmente, apresentava-se como uma fita fina, formado por um lobo dorsal e um lobo ventral. Longitudinalmente o pâncreas em emas localiza-se no mesentério dorsal desde o fígado até a flexura cranial do duodeno, mantendo-se preso às alças duodenais por ligamentos. Histologicamente, era composto por uma cápsula delgada de tecido conjuntivo denso, com discretos lóbulos separados por tecido conjuntivo capsular, compostos por estruturas tubuloalveolares e ductos. O fígado e pâncreas de emas apresentam padrão morfológico similar ao descrito para aves domésticas.

Protection conferred by heterologous vaccination against tuberculosis is dependent on the ratio of CD4(+)/CD4(+) Foxp3(+) cells

CD4(+) Foxp3(+) regulatory T cells inhibit the production of interferon-?, which is the major mediator of protection against Mycobacterium tuberculosis infection. In this study, we evaluated whether the protection conferred by three different vaccines against tuberculosis was associated with the number of spleen and lung regulatory T cells. We observed that after homologous immunization with the 65 000 molecular weight heat-shock protein (hsp 65) DNA vaccine, there was a significantly higher number of spleen CD4(+) Foxp3(+) cells compared with non-immunized mice. Heterologous immunization using bacillus Calmette Guerin (BCG) to prime and DNA-hsp 65 to boost (BCG/DNA-hsp 65) or BCG to prime and culture filtrate proteins (CFP)-CpG to boost (BCG/CFP-CpG) induced a significantly higher ratio of spleen CD4(+)/CD4(+) Foxp3(+) cells compared with non-immunized mice. In addition, the protection conferred by either the BCG/DNA-hsp 65 or the BCG/CFP-CpG vaccines was significant compared with the DNA-hsp 65 vaccine. Despite the higher ratio of spleen CD4(+)/CD4(+) Foxp3(+) cells found in BCG/DNA-hsp 65-immunized or BCG/CFP-CpG-immunized mice, the lungs of both groups of mice were better preserved than those of DNA-hsp 65-immunized mice. These results confirm the protective efficacy of BCG/DNA-hsp 65 and BCG/CFP-CpG heterologous prime-boost vaccines and the DNA-hsp 65 homologous vaccine. Additionally, the prime-boost regimens assayed here represent a promising strategy for the development of new vaccines to protect against tuberculosis because they probably induce a proper ratio of CD4(+) and regulatory (CD4(+) Foxp3(+)) cells during the immunization regimen. In this study, this ratio was associated with a reduced number of regulatory cells and no injury to the lungs.

Acute heat stress impairs performance parameters and induces mild intestinal enteritis in broiler chickens: Role of acute hypothalamic-pituitary-adrenal axis activation

Studies on the environmental consequences of stress are relevant for economic and animal welfare reasons. We recently reported that long-term heat stressors (31 +/- 1 degrees C and 36 +/- 1 degrees C for 10 h/d) applied to broiler chickens (Gallus gallus domesticus) from d 35 to 42 of life increased serum corticosterone concentrations, decreased performance variables and the macrophage oxidative burst, and produced mild, multifocal acute enteritis. Being cognizant of the relevance of acute heat stress on tropical and subtropical poultry production, we designed the current experiment to analyze, from a neuroimmune perspective, the effects of an acute heat stress (31 +/- 1 degrees C for 10 h on d 35 of life) on serum corticosterone, performance variables, intestinal histology, and peritoneal macrophage activity in chickens. We demonstrated that the acute heat stress increased serum corticosterone concentrations and mortality and decreased food intake, BW gain, and feed conversion (P < 0.05). We did not find changes in the relative weights of the spleen, thymus, and bursa of Fabricius (P > 0.05). Increases in the basal and the Staphylococcus aureus-induced macrophage oxidative bursts and a decrease in the percentage of macrophages performing phagocytosis were also observed. Finally, mild, multifocal acute enteritis, characterized by the increased presence of lymphocytes and plasmocytes within the lamina propria of the jejunum, was also observed. We found that the stress-induced hypothalamic-pituitary-adrenal axis activation was responsible for the negative effects observed on chicken performance and immune function as well as for the changes in the intestinal mucosa. The data presented here corroborate with those presented in other studies in the field of neuroimmunomodulation and open new avenues for the improvement of broiler chicken welfare and production performance.