Immunostaing of p53 associated with absence of mutations in TP53 gene in dogs with lymphoma

Sabendo-se da influência das mutações no gene TP53 no desenvolvimento das neoplasias e da discrepância entre os resultados obtidos pelas técnicas de sequenciamento e imunoistoquímica, esta pesquisa teve como objetivo relacionar a sequência do TP53 com a imunorreatividade da p53. Foram obtidas amostras de linfoma de 12 cães. O diagnóstico histopatológico foi determinado pela classificação de Kiel. O imunofenótipo e a imunomarcação da p53 foram determinados por imunoistoquímica. Para reação com a p53, utilizou-se anticorpo policlonal anti-p53 (CM1) na diluição de 1:500. A região do gene TP53 compreendida entre os exons quatro e nove foi amplificada por PCR e submetida ao sequenciamento. Apesar dos resultados obtidos pela imunoistoquímica, nenhuma mutação foi encontrada nas sequências analisadas. Conclui-se que a imunorreatividade da p53 pela imunoistoquímica não pode ser atribuída à presença de mutações no domínio central do gene TP53.TP53 mutations are usually involved in cancer, but sequencing and immunohistochemistry results are often controversial. Thus, the aim of this study was to associate TP53 sequence with p53 immunostaining in dogs with lymphoma. Tumor samples were collected from 12 dogs with lymphoma and were included in this study. Histopathological diagnosis was performed according to Kiel classification. Immunohistochemistry was performed to identify immunophenotype as well as p53 expression. Polyclonal antibody anti-p53 (CM1) was used at a 1:500 dilution. The region that encompasses exons 4-9 was amplified by f PCR reactions and sequencing was then performed. Nevertheless, gene mutations were not observed in any sequence. In conclusion, immunoreactivity of p53 by means of immunohistochemistry should not be indicator of presence of mutations in the core domain of TP53 gene

Serum concentration and immunostaining of vascular endothelial growth factor in dogs with multicentric lymphoma

O fator de crescimento do endotélio vascular (VEGF) é um dos mais específicos reguladores da angiogênese e da linfangiogênese tumoral, além de influenciar na tumorigênese mediante ativação de um circuito de sinalização autócrina que permite que as células neoplásicas estimulem seu próprio crescimento. O objetivo desse estudo foi investigar os níveis séricos e a imunomarcação do VEGF nos linfomas multicêntricos caninos e correlacionar esses parâmetros com fatores prognósticos da neoplasia. Foram avaliados 16 cães, sendo oito clinicamente sadios e oito com diagnóstico de linfoma multicêntrico. Os animais foram submetidos à coleta de sangue para mensuração da concentração sérica do VEGF e biopsia de linfonodo para avaliação da imunomarcação do VEGF. O escore de imunomarcação do VEGF foi superior (p=0,0003) nos linfonodos de cães com linfoma multicêntrico (8,50±2,33) em comparação ao escore dos linfonodos de cães sadios (1,87±1,80). Não houve diferença significativa entre as concentrações séricas do VEGF de cães sadios e de cães com linfoma (p=0,08). Imunofenótipo, estadiamento clínico e grau de malignidade influenciaram na sobrevida dos cães com linfoma. Os resultados obtidos permitiram concluir que o VEGF está expresso em quantidades elevadas nos linfonodos de cães com linfoma multicêntrico, podendo ser responsável pelo crescimento, sobrevivência e migração das células tumorais. Vascular endothelium growth factor (VEGF) is one of the most specific regulators of tumor angiogenesis and lymphangiogenesis. Moreover, VEGF plays a role in tumorigenesis through activation of an autocrine signaling pathway that enables cancer cells to stimulate their own growth. The aim of this study was to investigate VEGF serum levels and immunoexpression in canine multicentric lymphomas and correlate these parameters with the prognostic factors of this cancer. Sixteen dogs were evaluated (eight clinically healthy and eight diagnosed with multicentric lymphoma). The animals underwent blood sampling to measure VEGF serum concentrations and lymph node biopsy to evaluate VEGF immunoexpression. The VEGF immunoreactivity score was higher (p=0.0003) in the lymph nodes of dogs with multicentric lymphoma (8.50±2.33) than in those of healthy dogs (1.87±1.80). There was no significant difference in the VEGF serum concentrations between healthy dogs and dogs with lymphoma (p=0.08). Immunophenotype, clinical stage and grade of malignancy influenced the life expectancy of dogs with lymphoma. Our results showed that VEGF is expressed in high amounts in the lymph nodes of dogs with multicentric lymphoma and may be responsible for the growth, survival and migration of tumor cells

Meningoencefalite supurativa por Corynebacterium pseudotuberculosis em cabra com linfadenite caseosa: relato de caso

Suppurative lesions in the central nervous system of small ruminants may be caused by Corynebacterium pseudotuberculosis. This report describes a case of suppurative meningoencephalitis caused by C. pseudotuberculosis in goats treated in the Large Animal Clinic of FMVZ-UNESP/Botucatu, SP. The animal had anorexia, caseous lymphadenitis (CL), recumbency, opisthotonus, paddling movements and nystagmus. The CSF showed increased protein and lymphocytic pleocytosis. At necropsy there were localized abscess in the midbrain that after the microbiological culture, resulted in the isolation of C. pseudotuberculosis.Lesiones supurativas en el sistema nervioso central de los pequeños rumiates pueden ser causados por Corynebacterium pseudotuberculosis. En este informe se describe un caso de meningoencefalitis supurativa causada por C. pseudotuberculosis en cabras tratadas en el Large Animal Clinic FMVZ-UNESP/Botucatu, SP. El animal tenía anorexia, linfadenitis caseosa (CL), decúbito, opistótonos, movimientos de remo y nistagmo. El LCR mostró un aumento de proteínas y pleocitosis linfocitaria. En la necropsia había absceso localizado en el cerebro medio que después de que el cultivo microbiológico, dio como resultado el aislamiento de C. pseudotuberculosis.Lesões supurativas no sistema nervoso central de pequenos ruminantes podem ser causadas por Corynebacterium pseudotuberculosis. Este relato descreve um caso de meningoencefalite supurativa por C. pseudotuberculosis em cabra atendida na Clínica de Grandes Animais da FMVZ-UNESP/Botucatu, SP. O animal apresentava anorexia, linfadenite caseosa (LC), decúbito, opistótono, movimentos de pedalagem e nistagmo. O líquor evidenciou aumento de proteínas e pleocitose linfocítica. Na necropsia havia abscesso localizado no mesencéfalo que, após o cultivo microbiológico, resultou no isolamento de C. pseudotuberculosis

Immunophenotypic, immunocytochemistry, ultrastructural, and cytogenetic characterization of mesenchymal stem cells from equine bone marrow

The aim of this study was to isolate, culture, and characterize mesenchymal stem cells (MSCs) from horse bone marrow (BM) using the techniques of flow cytometry, immunocytochemistry, cytogenetics, and electron microscopy. Immunophenotypic analysis revealed the presence of MSCs with high expression of the CD90 marker, lower expression of the CD44 marker, and absent expression of the CD34 marker. In assays of differentiation, the positive response to osteogenic (OST), chondrogenic (CDG), and adipogenic (ADP) differentiation signals was observed and characterized by deposition of calcium-rich extracellular matrix (OST), proteoglycans and collagen II (CDG) and intracellular deposition of fat drops (ADP). In immunocytochemical characterization, MSCs were immunopositive for CD44, vimentin, and PCNA, and they were negative for CD13. In the ultrastructural analysis of MSCs, the most outstanding characteristic was the presence of rough endoplasmic reticulum with very dilated cisterns filled with a low electrodensity material. Additionally, MSCs had normal karyotypes (2n=64) as evidenced by cytogenetic analysis, and aneuploidy in metaphase was not observed. The protocols for isolating, culturing, and characterizing equine MSCs used in this study were shown to be appropriate for the production of a cell population with a good potential for differentiation and without aneuploidy that can be used to study future cellular therapies. © 2013 Wiley Periodicals, Inc