Differential regulations of AQP4 and Kir4.1 by triamcinolone acetonide and dexamethasone in the healthy and inflamed retina.

PURPOSE: Glucocorticoids are used to treat macular edema, although the mechanisms underlying this effect remain largely unknown. The authors have evaluated in the normal and endotoxin-induced uveitis (EIU) rats, the effects of dexamethasone (dex) and triamcinolone acetonide (TA) on potassium channel Kir4.1 and aquaporin-4 (AQP4), the two main retinal Müller glial (RMG) channels controlling retinal fluid movement. METHODS: Clinical as well as relatively low doses of dex and TA were injected in the vitreous of normal rats to evaluate their influence on Kir4.1 and AQP4 expression 24 hours later. The dose-dependent effects of the two glucocorticoids were investigated using rat neuroretinal organotypic cultures. EIU was induced by footpad lipopolysaccharide injection, without or with 100 nM intraocular dex or TA. Glucocorticoid receptor and channel expression levels were measured by quantitative PCR, Western blot, and immunohistochemistry. RESULTS: The authors found that dex and TA exert distinct and specific channel regulations at 24 hours after intravitreous injection. Dex selectively upregulated Kir4.1 (not AQP4) in healthy and inflamed retinas, whereas TA induced AQP4 (not Kir4.1) downregulation in normal retina and upregulation in EIU. The lower concentration (100 nM) efficiently regulated the channels. Moreover, in EIU, an inflammatory condition, the glucocorticoid receptor was downregulated in the retina, which was prevented by intravitreous injections of the low concentration of dex or TA. CONCLUSIONS: The results show that dex and TA are far from being equivalent to modulate RMG channels. Furthermore, the authors suggest that low doses of glucocorticoids may have antiedematous effects on the retina with reduced toxicity.

Direct binding of peptides to MHC class I molecules on living cells. Analysis at the single cell level.

To directly assess the binding of exogenous peptides to cell surface-associated MHC class I molecules at the single cell level, we examined the possibility of combining the use of biotinylated peptide derivatives with an immunofluorescence detection system based on flow cytometry. Various biotinylated derivatives of the adenovirus 5 early region 1A peptide 234-243, an antigenic peptide recognized by CTL in the context of H-2Db, were first screened in functional assays for their ability to bind efficiently to Db molecules on living cells. Suitable peptide derivatives were then tested for their ability to generate positive fluorescence signals upon addition of phycoerythrin-labeled streptavidin to peptide derivative-bearing cells. Strong fluorescent staining of Db-expressing cells was achieved after incubation with a peptide derivative containing a biotin group at the C-terminus. Competition experiments using the unmodified parental peptide as well as unrelated peptides known to bind to Kd, Kb, or Db, respectively, established that binding of the biotinylated peptide to living cells was Db-specific. By using Con A blasts derived from different H-2 congenic mouse strains, it could be shown that the biotinylated peptide bound only to Db among > 20 class I alleles tested. Moreover, binding of the biotinylated peptide to cells expressing the Dbm13 and Dbm14 mutant molecules was drastically reduced compared to Db. Binding of the biotinylated peptide to freshly isolated Db+ cells was readily detectable, allowing direct assessment of the relative amount of peptide bound to distinct lymphocyte subpopulations by three-color flow cytometry. While minor differences between peripheral T and B cells could be documented, thymocytes were found to differ widely in their peptide binding activity. In all cases, these differences correlated positively with the differential expression of Db at the cell surface. Finally, kinetic studies at different temperatures strongly suggested that the biotinylated peptide first associated with Db molecules available constitutively at the cell surface and then with newly arrived Db molecules.

Strain-transcending Fc-dependent killing of Plasmodium falciparum by merozoite surface protein 2 allele-specific human antibodies.

It is widely accepted that antibody responses against the human parasitic pathogen Plasmodium falciparum protect the host from the rigors of severe malaria and death. However, there is a continuing need for the development of in vitro correlate assays of immune protection. To this end, the capacity of human monoclonal and polyclonal antibodies in eliciting phagocytosis and parasite growth inhibition via Fcγ receptor-dependent mechanisms was explored. In examining the extent to which sequence diversity in merozoite surface protein 2 (MSP2) results in the evasion of antibody responses, an unexpectedly high level of heterologous function was measured for allele-specific human antibodies. The dependence on Fcγ receptors for opsonic phagocytosis and monocyte-mediated antibody-dependent parasite inhibition was demonstrated by the mutation of the Fc domain of monoclonal antibodies against both MSP2 and a novel vaccine candidate, peptide 27 from the gene PFF0165c. The described flow cytometry-based functional assays are expected to be useful for assessing immunity in naturally infected and vaccinated individuals and for prioritizing among blood-stage antigens for inclusion in blood-stage vaccines.

Direct Thy-1/alphaVbeta3 integrin interaction mediates neuron to astrocyte communication.

Thy-1 is an abundant neuronal glycoprotein of poorly defined function. We recently provided evidence indicating that Thy-1 clusters a beta3-containing integrin in astrocytes to induce tyrosine phosphorylation, RhoA activation and the formation of focal adhesions and stress fibers. To date, the alpha subunit partner of beta3 integrin in DI TNC1 astrocytes is unknown. Similarly, the ability of neuronal, membrane-bound Thy-1 to trigger astrocyte signaling via integrin engagement remains speculation. Here, evidence that alphav forms an alphavbeta3 heterodimer in DI TNC1 astrocytes was obtained. In neuron-astrocyte association assays, the presence of either anti-alphav or anti-beta3 integrin antibodies reduced cell-cell interaction demonstrating the requirement of both integrin subunits for this association. Moreover, anti-Thy-1 antibodies blocked stimulation of astrocytes by neurons but not the binding of these two cell types. Thus, neuron-astrocyte association involved binding between molecular components in addition to the Thy-1-integrin; however, the signaling events leading to focal adhesion formation in astrocytes depended exclusively on the latter interaction. Additionally, wild-type (RLD) but not mutated (RLE) Thy-1 was shown to directly interact with alphavbeta3 integrin by Surface Plasmon Resonance analysis. This interaction was promoted by divalent cations and was species-independent. Together, these results demonstrate that the alphavbeta3 integrin heterodimer interacts directly with Thy-1 present on neuronal cells to stimulate astrocytes.

Prevalencia de enfermedad celiaca en Latinoamérica: revisión sistemática de la literatura y meta-análisis

Introducción: La enfermedad celiaca (EC) es una enfermedad autoinmune (EA) intestinal desencadenada por la ingesta de gluten. Por la falta de información de la presencia de EC en Latinoamérica (LA), nosotros investigamos la prevalencia de la enfermedad en esta región utilizando una revisión sistemática de la literatura y un meta-análisis. Métodos y resultados: Este trabajo fue realizado en dos fases: La primera, fue un estudio de corte transversal de 300 individuos Colombianos. La segunda, fue una revisión sistemática y una meta-regresión siguiendo las guías PRSIMA. Nuestros resultados ponen de manifiesto una falta de anti-transglutaminasa tisular (tTG) e IgA anti-endomisio (EMA) en la población Colombiana. En la revisión sistemática, 72 artículos cumplían con los criterios de selección, la prevalencia estimada de EC en LA fue de 0,46% a 0,64%, mientras que la prevalencia en familiares de primer grado fue de 5,5 a 5,6%, y en los pacientes con diabetes mellitus tipo 1 fue de 4,6% a 8,7% Conclusión: Nuestro estudio muestra que la prevalencia de EC en pacientes sanos de LA es similar a la notificada en la población europea.

Transplacental transmission of Toxoplasma gondii in reinfected pregnant female canines

Twelve pregnant female canines, naturally infected with Toxoplasma gondii, were reinfected with T. gondii: three (GI) received tachyzoites subcutaneously (1.0 x 107), three (GII) were orally inoculated with oocysts (1.5 x 104), and six (GIII) were kept as a nonreinfected control group. All the reinfected female canines (GI and GII) miscarried or presented fetal death, while only one GIII female presented a stillborn in a litter of four pups (P < 0.01). Fever, lymphoadenopathy, miscarriage, and fetal death were the main clinical alterations observed. The highest serological titers detected through the indirect fluorescence antibody test (IFAT) were 1,024 (GI) and 4,096 (GII). In group III, the titers ranged between 64 and 256. By bioassays in mice, T. gondii was isolated in 17 organs of the reinfected adult canines, in 11 of the control group, and in 20 of the neonates. Positive immunostaining of cysts and/or tachyzoites were observed in 26 canine tissues (14 from GI and GII and ten from GIII). The agent was detected by immunohistochemistry in the encephalon of a neonate and in the spinal cord of a stillborn, thus, confirming that T. gondii infected canine fetuses, provoking miscarriages, even in bitches that presented primoinfection.

Canine ehrlichiosis: clinical, hematological, serological and molecular aspects

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

Antigenic characterization of Trypanosoma evansi using sera from experimentally and naturally infected bovines, equines, dogs, and coatis

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

VISCERAL LEISHMANIASIS IN A CAPTIVE CRAB-EATING FOX CERDOCYON THOUS

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

Occurrence of anti-Toxoplasma gondii and anti-Neospora caninum antibodies in cats with outdoor access in São Luís, Maranhão, Brazil

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Tracoma em crianças do ensino fundamental no município de Bauru: Estado de São Paulo, Brasil

OBJETIVO: Foi realizado um estudo transversal, por amostragem de escolares, frequentadores das escolas estaduais de Bauru, São Paulo, com o objetivo de detectar possíveis portadores de tracoma. MÉTODOS: Foram examinadas 1.749 crianças do ensino fundamental, de 6 a 14 anos, durante o ano de 2005. As escolas foram escolhidas por sorteio, de acordo com o número de estudantes e com a localização da escola dentro do setor da cidade (centro, região intermediária e periferia). O diagnóstico dos casos foi clínico, seguindo as normas da Organização Mundial de Saúde (OMS), confirmado laboratorialmente pelo método da imunofluorescência direta para detecção de Chlamydia trachomatis. RESULTADOS: A prevalência de tracoma inflamatório foi de 3,8%, tendo sido diagnosticado tracoma folicular em 3,7% e tracoma intenso em 0,06% das crianças examinadas. As crianças com tracoma moravam principalmente na região intermediária e periférica da cidade, áreas onde o fornecimento de água pode ser descontínuo e o tratamento do esgoto pode estar ausente. Um programa de controle foi desenvolvido e os comunicantes foram diagnosticados e tratados. CONCLUSÃO: A prevalência de tracoma inflamatório em Bauru - Estado de São Paulo - é de 3,8%. Além da oportunidade de reconhecer e tratar os portadores e contatantes, os autores realçam o fato positivo da mobilização da comunidade bauruense no sentido de prevenir a cegueira pelo tracoma.

Inquérito sorológico para toxoplasmose em equinos na região de Botucatu-SP

The frequency of anti-T.gondii IgG antibodies was evaluated and compared in 253 sera samples of equines by modified agglutination test (MAT) and indirect immunofluorescent antibody test (IFAT). Fifteen samples (5.9%) were positive by IFAT and 32 (12.6%) by MAT. Significant difference between the two methods was found when a cutoff titer of 16 was established. There was no correlation with breed, gender, and age of horses examined.

Surveillance of canine visceral leishmaniasis in a disease-free area

A leishmaniose é uma importante parasitose re-emergente observada no mundo, particularmente em países tropicais. Não há ainda relatos de casos autóctones no estado do Paraná. Não há até o momento referência de vigilância no reservatório canino, tais como Curitiba e região metropolitana do estado. O objetivo do estudo foi determinar a soroprevalência da leishmaniose visceral em cães entregues ao Centro de Controle de Zoonoses de São José dos Pinhais, Paraná para eutanásia. A detecção sorológica da presença de anticorpos contra Leishmania sp. foi realizada por (ELISA) indireto e pela Reação de Imunofluorescência Indireta (RIFI). Além disso, impressão de linfonodo poplíteo coletadas ao acaso de 50 cães com sinais clínicos suspeitos para leishmaniose visceral e analisados sob microscopia óptica para detecção de formas amastigotas, foram negativas. Amostras de soro de 364 animais foram testadas, e os resultados mostraram somente uma amostra positiva (0,0027%), reagente ao ELISA e negativa à RIFI, entretanto, o cão não apresentava sinais clínicos. A vigilância ao acaso em uma população de vários locais de uma área metropolitana pode ser uma forma de prevenção da disseminação da doença. Com base nos resultados observados, Curitiba e região metropolitana foram consideradas de baixo risco para a leishmaniose visceral.

Active surveillance of canine visceral leishmaniasis and american trypanossomiasis in rural dogs from non endemic area

A leishmaniose visceral canina (LVC) e a tripanossomíase americana são importantes zoonoses para a saúde pública que encontram no cão o principal reservatório doméstico para o homem. O trabalho procurou estimar a prevalência de anticorpos circulantes anti-Trypanosoma cruzi e anti-Leishmania spp., em amostras de cães provenientes da zona rural do município de Botucatu, SP. Durante a campanha de vacinação antirrábica canina da zona rural do município, foram coletadas 689 amostras de soro e processadas pela técnica de imunofluorescência indireta. Os testes sorológicos revelaram a ausência de anticorpos anti-Leishmania spp. e, na pesquisa dos anticorpos anti-T. cruzi, foram detectados 3 (0,4%) cães.

Detection and molecular analysis of Toxoplasma gondii and Neospora caninum from dogs with neurological disorders

Introduction: Toxoplasma gondii and Neospora caninum are related Apicomplexa parasites responsible for systemic diseases in many species of animals, including dogs. Methods: This study aimed to determine the occurrence of T. gondii and N. caninum infections in 50 dogs with neurological signs that were admitted to the Veterinary Hospital of Universidade Estadual Paulista, City of Botucatu, Brazil. All animals were screened for antibodies using an immunofluorescent antibody test for both parasites. Tissues of positive animals were bioassayed in mice (T. gondii) and gerbils (N. caninum), and DNA was analyzed using the polymerase chain reaction (PCR). Positive samples for T. gondii by PCR were typed using restriction fragment length polymorphism-PCR for 11 markers: SAG1, SAG2 (5'-3'-SAG2 and alt.SAG2), SAG3, Btub, GRA6, L358, c22-8, c29-6, PK1 and Apico, and CS3 marker for virulence analysis. Results: Specific antibodies were detected in 11/50 (22%; 95% confidence interval (CI95%), 12.8-35.3%) animals for T. gondii and 7/50 (14%; CI95%, 7.02-26.3%) for N. caninum. In the bioassay and PCR, 7/11 (63.6%; CI95%, 34.9-84.8%) samples were positive for T. gondii and 3/7 (42.9%; CI95% I, 15.7-75.5%) samples were positive for N. caninum. Three different genotypes were identified, but only 1 was unique. Conclusions: These data confirm the presence of T. gondii and N. caninum in dogs from Brazil, indicating the importance of this host as a sentinel of T. gondii for human beings, and the genotypic variation of this parasite in Brazil.