The effect of Lacticaseibacillus casei on inflammatory cytokine (IL-8) gene expression induced by exposure to Shigella sonnei in Zebrafish (Danio rerio) / [O efeito do Lacticaseibacillus casei na citoquina inflamatória (IL-8) expressão genética induzida pela exposição a Shigella sonnei em Zebrafish (Danio rerio)]

This study aimed to evaluate the protective function of probiotics against Shigella sonnei pathogenicity. For this purpose, 400 zebrafish were divided into four groups with two replications: (T1): receiving Lacticaseibacillus casei for 27 days, (T2): receiving L. casei for 27 days followed by 72 hr exposure to S. sonnei, (T3): receiving basal diet for 27 days followed by 72 hr exposure to S. sonnei, and control group (C): receiving basal diet without exposure to the pathogen. According to the results, feeding with L. casei for 27 days reduced the interleukin-8 (IL-8) expression significantly (P<0.05). The results showed a decrease in IL-8 expression in the group exposed to the pathogen and fed with the probiotic compared to the group only fed with the basal diet (P<0.05). Considering the role of IL-8 as a pro-inflammatory cytokine, our results indicated that feeding with L. casei could modulate inflammatory responses.

Este estudo teve como objetivo avaliar a função protetora dos probióticos contra a patogenicidade Shigella sonnei. Para este fim, 400 zebrafish foram divididos em quatro grupos com duas réplicas: (T1): recebendo Lacticaseibacillus casei por 27 dias, (T2): recebendo L. casei por 27 dias seguido por 72 horas de exposição a S. sonnei, (T3): recebendo dieta basal por 27 dias seguido por 72 horas de exposição a S. sonnei, e grupo controle (C): recebendo dieta basal sem exposição ao patógeno. De acordo com os resultados, a alimentação com L. casei por 27 dias reduziu significativamente a expressão da interleucina-8 (IL-8) (P<0,05). Os resultados mostraram uma diminuição na expressão de IL-8 no grupo exposto ao patógeno e alimentado com o probiótico em comparação com o grupo alimentado apenas com a dieta basal (P<0,05). Considerando o papel da IL-8 como uma citocina pró-inflamatória, nossos resultados indicaram que a alimentação com L. casei poderia modular as respostas inflamatórias.

Detection of respiratory viruses and expression of inflammatory cytokines in patients with acute exacerbation chronic obstructive pulmonary disease in Mongolia China / Detecção de vírus respiratórios e expressão de citocinas inflamatórias em pacientes com doença pulmonar obstrutiva crônica de exacerbação aguda na Mongólia, China

Chronic obstructive pulmonary disease (COPD) was estimated to be the third cause of global mortality by 2020. Acute exacerbation COPD (AECOPD) is a sudden worsening of COPD symptoms and could be due to virus/bacterial infections and air pollution. Increased expression of inflammatory markers in patients with AECOPD is associated with viral infection. This study aimed to detect different viruses and analyze the expression of various inflammatory markers associated with AECOPD patients. Three hundred and forty-seven patients diagnosed with COPD according to GOLD criteria were included in this study. Swab samples and blood were collected for the detection of viruses by RT-PCR and expression of inflammatory markers, respectively. Of the swab samples, 113 (32.6%) of samples were positive for virus detection. Of these, HRV (39.8%) was the predominant virus detected followed by FluB (27.4%) and FluA (22.1%). The presence of HRV was significantly higher (p=0.044) among the other detected viruses. When compared to healthy controls the expression levels of TNF-α, IL-6 and IL-8 were significantly higher (p<0.05) in virus-positive patients. The IL-6 and IL-8 were the next predominantly expressed in markers among the samples. The higher expression rate of IL-8 was significantly (p<0.05) associated with patients having COPD GOLD III severity level and smoking history. Although HRV was the predominant virus detected the combined prevalence of Influenza A and B surpassing the rate of HRV. The high-level expression of well known inflammatory markers of AECOPD, TNF-α, IL-6 and IL-8 indicates a chronic severe illness. These markers play an important role and could be used as a marker for determining the severity of AECOPD.(AU)

Estima-se que a doença pulmonar obstrutiva crônica (DPOC) seja a terceira causa de mortalidade global em 2020. A exacerbação aguda DPOC (AECOPD) é um agravamento súbito dos sintomas da DPOC e pode ser devido a infecções por vírus/bactérias e poluição do ar. O aumento da expressão de marcadores inflamatórios em pacientes com AECOPD está associado à infecção viral. Este estudo teve como objetivo detectar diferentes vírus e analisar a expressão de vários marcadores inflamatórios associados a pacientes com AECOPD. Trezentos e quarenta e sete pacientes com diagnóstico de DPOC de acordo com os critérios GOLD foram incluídos neste estudo. Amostras de swab e sangue foram coletadas para detecção de vírus por RT-PCR e expressão de marcadores inflamatórios, respectivamente. Das amostras de esfregaço, 113 (32,6%) amostras foram positivas para detecção de vírus. Nestas, o HRV (39,8%) foi o vírus predominante detectado, seguido do FluB (27,4%) e do FluA (22,1%). A presença de VFC foi significativamente maior (p = 0,044) entre os demais vírus detectados. Quando comparados a controles saudáveis, os níveis de expressão de TNF-α, IL-6 e IL-8 foram significativamente maiores (p <0,05) em pacientes com vírus positivo. A IL-6 e a IL-8 foram as próximas predominantemente expressas em marcadores entre as amostras. A maior taxa de expressão de IL-8 foi significativamente (p <0,05) associada a pacientes com grau de gravidade GOLD III da DPOC e história de tabagismo. Embora o HRV tenha sido o vírus predominante, a prevalência combinada de Influenza A e B ultrapassou a taxa de HRV. O alto nível de expressão de marcadores inflamatórios bem conhecidos de AECOPD, TNF-α, IL-6 e IL-8 indica uma doença crônica grave. Esses marcadores desempenham um papel importante e podem ser usados como um marcador para determinar a gravidade da AECOPD.(AU)

Clonality test by PCR - PARR in real time of canine lymphomas

Lymphoma is a neoplasm of hematopoietic origin that affects canines. The proper establishment of prognosis and rapid institution of treatment are essential for a better quality of life, and immunophenotyping is one of the tools used for this purpose. The objective of this study was to perform a clonality test for immunophenotypic characterization of canine lymphomas using the polymerase chain reaction (PCR) for antigen receptor rearrangements (PARR) technique in real-time from samples fixed in formalin and embedded in paraffin. The 23 analyzed samples were fixed in formalin and embedded in paraffin canine lymphoma from the collection Laboratory of Histopathology of the Animal Pathology Area of the Departament of Veterinary Medicine - Federal Rural University of Pernambuco (UFRPE). Samples were processed, their DNA was extracted, quantified, diluted, and standardized at a concentration of 50 ng/µL. After extraction, all samples were subjected to conventional PCR for endogenous control (detection of the IgM target region), in which the extracted DNA was amplified in a final volume of 25 µL. The 128 bp amplified product was detected by 1.5% agarose gel electrophoresis. Of the 23 samples analyzed for the detection of the conserved region referring to the endogenous gene, 91.30% (21/23) amplified the conserved region Cµ by conventional PCR, and two samples 8.70% (2/23) were negative. Endogenous control positive samples were subjected to real-time PCR-PARR for detection of IgH Major and IgH Minor for B lymphocytes (LB), and TCRy for lymphocytes T (LT) target regions. All reactions were performed in duplicate to reduce the risk of false-positive or false-negative results due to technical errors. Samples previously confirmed by immunohistochemistry were used as positive controls for T cell and B cell lymphoma, and MilliQ water was used as a negative reaction control. After amplification, the melting curve gradually increased the temperature by 1o C/5 s to 95o C during continuous fluorescence monitoring. Of the 21 samples analyzed, 100.00% (21/21) demonstrated clonal amplification. Of these, 57.15% (12/21) were positive for phenotype B, and 42.85% (9/21) were positive for phenotype T. Due to the importance of researching and confirming samples from files fixed and embedded in paraffin samples in laboratories, PCR-PARR is a good tool for this purpose. In the present study, real-time PCR analysis demonstrated greater sensitivity in the characterization of the immunophenotype of lymphomas from old samples fixed in formalin and embedded in paraffin. The temperature of melting curve analysis may vary depending on the amount of DNA and its quality. In the present study, it was found that the average melting temperature in the samples varied between ± 3o C when compared to that in the control sample for LB and LT, 83.5o C and 80o C, respectively: in the literature, there is a relative difference in this temperature, which may vary up to 4o C. Real-time PCR-PARR was satisfactory in the characterization of the immunophenotype of canine lymphomas from formalin-fixed and paraffin-embedded samples; therefore, its use is recommended for both retrospective studies. The use of PCR-PARR associated with histopathological and/or cytopathological examination in cases of canine lymphomas strongly helps pathologists, provide a safe establishment of the immunophenotype, minimize errors, and optimize the diagnosis, thus directly contributing to the establishment of the prognosis.(AU)

Linfoma de células T na cavidade nasal de caprino / T-cell lymphoma in the nasal cavity in goat

Background: Lymphomas are considered uncommon in goats, being the multicentric form with the highest number of cases for the species. Primary intranasal lymphomas are often diagnosed in dogs, cats, and humans. In the literature, there is only a description of a multicentric case involving the frontal sinuses and mucosa of the nasal cavity in a goat; therefore, it is important to describe unusual cases of this disease for the inclusion of new clinical and pathological characteristics in the ruminant clinic medicine. The objective of this work is to describe a case of T-cell lymphoma in the nasal cavity of a young goat. Case: The animal had dyspnea and respiratory noise for 15 days. Clinical examination showed nodulation in the right nasal cavity associated with serosanguinous secretion. Tracheostomy was performed; however, after 30 days the animal was euthanized. A sagittal plane of the head showed a pinkish-gray mass in the right and left nasal cavity, with a smooth, multilobulated surface, smooth adhering to the rostral portion of the dorsal concha and occluding the dorsal nasal meatus. Submandibular lymph nodes were slightly enlarged. Histopathological examination of the nasal cavity revealed a non-encapsulated, poorly delimited and ulcerated tumor composed of round cells arranged in a mantle supported by a discrete fibrovascular stroma extending the mucosa and lamina propria. Cells were round with sparse, eosinophilic and poorly delimited cytoplasm. Nuclei varied from round to elongated with condensed chromatin and evident nucleoli. Occasionally, aberrant nuclei, reniform shape and multinucleated cells were seen. Pleomorphism was moderate characterized by anisocytosis and anisocariosis. Typical and atypical mitosis were frequent (0-4 per field of highest magnification [400x]). Amidst the neoplasm, there were multifocal areas of necrosis and hemorrhage associated with a mild lymphocytic inflammatory infiltrate. Immunohistochemistry showed positive immunostaining for Vimentin antibodies and CD3, and negative for pan CK and CD20. Discussion: The lymphomas immunophenotyping is little used when it comes to farm animals, and there are few studies that use this technique for the definitive diagnosis of these neoplasms for small ruminants. The use of this technique must be considered in each case, in order to determine the pathogenesis, the accurate diagnosis and the origin of the neoplastic lymphocytes. In goats, T-cell lymphomas are the most diagnosed, although cases of multicentric B-cell lymphomas with ocular involvement have been diagnosed. In view of the clinical picture of the case described, infectious rhinitis already described in goats, such as aspergillosis and protothecosis, should be included as differential diagnoses. However, the anatomopathological findings facilitate the direction of the diagnosis, since infectious rhinitis presents as nodules / ulcerated masses or focal areas of necrosis associated with purulent secretion and in the histopathological examination it is possible to observe the intralesional etiological agents. In addition, the enzootic ethmoidal tumor must be included, as it has similar clinical signs and affects young animals, but they are adenomas/adenocarcinomas that affect the ethmoidal nasal shells induced by a retrovirus. Lymphomas in the caprine species are rare in the Northeastern semi-arid, but that in the present diagnostic routine occasionally occurs, being important the first description of its nasal shape for its inclusion in the differential diagnoses of diseases that present with clinical obstruction and dyspnea for the species.

Histopathological and immunophenotypical characterization of canine multicentric lymphoma in Brazil: a study of 203 cases / Caracterização histopatológica e imunofenotípica do linfoma multicêntrico canino no Brasil: um estudo de 203 casos

The immunophenotype is regarded as an independent prognostic factor in high-grade lymphomas, seeing that lymphomas of T-cell origin are associated with shorter survival time. Although a number of studies have evaluated the immunophenotypical profile of lymphoma in the USA and Europe, Brazilian research on the matter remains scarce. Exact characterization of the histopathological type is crucial to establish proper treatment and prognosis. This study evaluated the database of immunohistochemistry laboratories that perform immunophenotyping of canine lymphoma in Brazil. A total of 203 cases of multicentric lymphoma were classified according to the WHO classification. Immunophenotyping was able to identify 71.4% lymphomas of B-cell line, 27.1% of T-cell line and 1.5% of non-B cells and non-T cell lines. Diffuse large B-cell lymphoma was the most common with 59.1% of the cases. Among T-cell lymphomas, lymphoblastic was the most common (11.33% of the cases). Even though canine lymphomas tend to be high-grade, indolent lymphomas comprised 11.82% of the cases and T-zone lymphoma was the most prevalent (8.86%). The immunophenotype of multicentric lymphoma in Brazil is similar to those in other parts of the world, which suggests similar etiologic factors to the development of this disease.(AU)

O imunofenótipo é considerado um fator prognóstico independente em linfomas de alto grau, visto que os linfomas de origem de células T estão associados a menor tempo de sobrevida. Apesar de vários estudos terem avaliado o perfil imunofenotípico do linfoma nos EUA e na Europa, a pesquisa brasileira sobre o assunto ainda é escassa. A caracterização exata do tipo histopatológico é crucial para estabelecer o tratamento e o prognóstico adequados. Este estudo avaliou a base de dados de laboratórios de imuno-histoquímica que realizam imunofenotipagem do linfoma canino no Brasil. Um total de 203 casos de linfoma multicêntrico foi classificado de acordo com a classificação da OMS. A imunofenotipagem foi capaz de identificar 71,4% dos linfomas da linhagem de células B, 27,1% da linhagem de células T e 1,5% das linhagens de células não B e não T. O linfoma difuso de grandes células B foi o mais comum em 59,1% dos casos. Entre os linfomas de células T, o linfoblástico foi o mais comum (11, 33% dos casos). Embora os linfomas caninos tendam a ser de alto grau, os linfomas indolentes representaram 11,82% dos casos e o linfoma da zona T foi o mais prevalente (8,86%). O imunofenótipo do linfoma multicêntrico no Brasil é semelhante ao de outras partes do mundo, o que sugere fatores etiológicos semelhantes ao desenvolvimento dessa doença.(AU)

Histopathological and immunophenotypical characterization of canine multicentric lymphoma in Brazil: a study of 203 cases / Caracterização histopatológica e imunofenotípica do linfoma multicêntrico canino no Brasil: um estudo de 203 casos

The immunophenotype is regarded as an independent prognostic factor in high-grade lymphomas, seeing that lymphomas of T-cell origin are associated with shorter survival time. Although a number of studies have evaluated the immunophenotypical profile of lymphoma in the USA and Europe, Brazilian research on the matter remains scarce. Exact characterization of the histopathological type is crucial to establish proper treatment and prognosis. This study evaluated the database of immunohistochemistry laboratories that perform immunophenotyping of canine lymphoma in Brazil. A total of 203 cases of multicentric lymphoma were classified according to the WHO classification. Immunophenotyping was able to identify 71.4% lymphomas of B-cell line, 27.1% of T-cell line and 1.5% of non-B cells and non-T cell lines. Diffuse large B-cell lymphoma was the most common with 59.1% of the cases. Among T-cell lymphomas, lymphoblastic was the most common (11.33% of the cases). Even though canine lymphomas tend to be high-grade, indolent lymphomas comprised 11.82% of the cases and T-zone lymphoma was the most prevalent (8.86%). The immunophenotype of multicentric lymphoma in Brazil is similar to those in other parts of the world, which suggests similar etiologic factors to the development of this disease.(AU)

O imunofenótipo é considerado um fator prognóstico independente em linfomas de alto grau, visto que os linfomas de origem de células T estão associados a menor tempo de sobrevida. Apesar de vários estudos terem avaliado o perfil imunofenotípico do linfoma nos EUA e na Europa, a pesquisa brasileira sobre o assunto ainda é escassa. A caracterização exata do tipo histopatológico é crucial para estabelecer o tratamento e o prognóstico adequados. Este estudo avaliou a base de dados de laboratórios de imuno-histoquímica que realizam imunofenotipagem do linfoma canino no Brasil. Um total de 203 casos de linfoma multicêntrico foi classificado de acordo com a classificação da OMS. A imunofenotipagem foi capaz de identificar 71,4% dos linfomas da linhagem de células B, 27,1% da linhagem de células T e 1,5% das linhagens de células não B e não T. O linfoma difuso de grandes células B foi o mais comum em 59,1% dos casos. Entre os linfomas de células T, o linfoblástico foi o mais comum (11, 33% dos casos). Embora os linfomas caninos tendam a ser de alto grau, os linfomas indolentes representaram 11,82% dos casos e o linfoma da zona T foi o mais prevalente (8,86%). O imunofenótipo do linfoma multicêntrico no Brasil é semelhante ao de outras partes do mundo, o que sugere fatores etiológicos semelhantes ao desenvolvimento dessa doença.(AU)

Intradural extramedullary plasmacytoma in a dog

Background: Plasmacytoma is a neoplasm originating in plasma cells, derived from B lymphocytes. Extramedullary presentation is the most common form of plasmacytoma, mainly in the skin and rarely in other tissues, including the vertebral canal. Extramedullary plasmacytoma (EMP) occurs more frequently in senile dogs and rarely in cats and some dog breeds have predisposition for this type of neoplasm. The aim of this study was to report plasmacytoma in a dog located adjacent to the thoracic vertebrae. Case: An approximately 5-year-old mongrel male with sudden paraplegia resulting from upper motor neuron injury was admitted to a University Hospital. The dog presented a six-centimeter diameter mass in the dorsal region, adjacent to the thoracic vertebrae. The leucogram presented mild leukopenia by eosinopenia and lymphopenia. Myelogram associated with epidurography showed a fill failure in the contrast column between the seventh and ninth thoracic vertebrae, and there were no osteolysis points in the thoracic vertebrae, adjacent to the neoplastic mass. The evaluation of the intervertebral spaces between the fourth to ninth thoracic vertebrae presented no increase in radiopacity, nor alterations in the size of intervertebral spaces, indicating intervertebral disc extrusion. The laterolateral radiographs showed an alignment of the vertebrae and vertebral canal, without presence of bone neoformations on the ventral face of the vertebral bodies. The animal was euthanized and fragments of mass were collected for histological analysis. Macroscopic findings presented a non-encapsulated well-delimited mass, with color ranging from white to reddish. Microscopically, there was proliferation of moderately differentiated round cells in bone and muscle tissues, arranged in mantle

Intradural extramedullary plasmacytoma in a dog

Background: Plasmacytoma is a neoplasm originating in plasma cells, derived from B lymphocytes. Extramedullary presentation is the most common form of plasmacytoma, mainly in the skin and rarely in other tissues, including the vertebral canal. Extramedullary plasmacytoma (EMP) occurs more frequently in senile dogs and rarely in cats and some dog breeds have predisposition for this type of neoplasm. The aim of this study was to report plasmacytoma in a dog located adjacent to the thoracic vertebrae. Case: An approximately 5-year-old mongrel male with sudden paraplegia resulting from upper motor neuron injury was admitted to a University Hospital. The dog presented a six-centimeter diameter mass in the dorsal region, adjacent to the thoracic vertebrae. The leucogram presented mild leukopenia by eosinopenia and lymphopenia. Myelogram associated with epidurography showed a fill failure in the contrast column between the seventh and ninth thoracic vertebrae, and there were no osteolysis points in the thoracic vertebrae, adjacent to the neoplastic mass. The evaluation of the intervertebral spaces between the fourth to ninth thoracic vertebrae presented no increase in radiopacity, nor alterations in the size of intervertebral spaces, indicating intervertebral disc extrusion. The laterolateral radiographs showed an alignment of the vertebrae and vertebral canal, without presence of bone neoformations on the ventral face of the vertebral bodies. The animal was euthanized and fragments of mass were collected for histological analysis. Macroscopic findings presented a non-encapsulated well-delimited mass, with color ranging from white to reddish. Microscopically, there was proliferation of moderately differentiated round cells in bone and muscle tissues, arranged in mantle…(AU)

Effects of Methionine Deficiency on the B Lymphocyte and Immunoglobulins of Cecal Tonsil in Cobb Broilers

The present study was conducted on one-day-old broilers to investigate the effects of methionine deficiency on B lymphocytes and immunoglobulins (sIgA, IgA, IgG, IgM) in the cecal tonsil of Cobb broiler chicken. Methods including immunohistochemistry (IHC), western blot (WB), quantitative real-time reverse transcriptase-polymerase chain reaction (qRT-PCR) and Enzyme-Linked Immuno Sorbent Assay (ELISA) were used, and the study lasted for 6 weeks. It was found that the IgA+ B lymphocytes mainly existed in the diffuse lymphoid tissues and lymphoid follicles of the cecal tonsil. The data indicated that the number of IgA+ B lymphocytes was decreased in the methionine deficiency group, and the WB and qRT-PCR results suggested that the protein and mRNA expression of CD19 were reduced in the methionine deficiency group. Besides, the contents of sIgA, IgA, IgG and IgM determined by ELISA decreased in the methionine deficiency group. It could be concluded that methionine deficiency exerts significantly negative effects on the humoral immune function of the intestinal mucosal immunity.(AU)

Effects of Methionine Deficiency on the B Lymphocyte and Immunoglobulins of Cecal Tonsil in Cobb Broilers

The present study was conducted on one-day-old broilers to investigate the effects of methionine deficiency on B lymphocytes and immunoglobulins (sIgA, IgA, IgG, IgM) in the cecal tonsil of Cobb broiler chicken. Methods including immunohistochemistry (IHC), western blot (WB), quantitative real-time reverse transcriptase-polymerase chain reaction (qRT-PCR) and Enzyme-Linked Immuno Sorbent Assay (ELISA) were used, and the study lasted for 6 weeks. It was found that the IgA+ B lymphocytes mainly existed in the diffuse lymphoid tissues and lymphoid follicles of the cecal tonsil. The data indicated that the number of IgA+ B lymphocytes was decreased in the methionine deficiency group, and the WB and qRT-PCR results suggested that the protein and mRNA expression of CD19 were reduced in the methionine deficiency group. Besides, the contents of sIgA, IgA, IgG and IgM determined by ELISA decreased in the methionine deficiency group. It could be concluded that methionine deficiency exerts significantly negative effects on the humoral immune function of the intestinal mucosal immunity.

Canine lymphomas diagnosed in southern Brazil from 2000 to 2017: epidemiology and immunophenotype / Linfomas caninos diagnosticados na região Sul do Rio Grande do Sul de 2000 a 2017: epidemiologia e imunofenótipo

Lymphoma is a neoplasm that originates from solid hematopoietic tissues and is one of the most common tumors in dogs. The goal of the present study was to perform a retrospective study of canine lymphomas diagnosed at the "Laboratório Regional de Diagnóstico", at the "Faculdade de Veterinária" of the "Universidade Federal de Pelotas" (LRD-UFPel) from 2000 to 2017, to determine the epidemiology and anatomical distribution, and to evaluate the histopathological and immunohistochemical aspects of each case according to the adapted Kiel classification. The protocols for necropsies and biopsies in the laboratory were reviewed. Lymphoma was diagnosed in 77 dogs. Approximately 37.7% (29/77) of affected dogs had no defined breed, while dogs with defined breeds accounted for 58.4% (45/77) of the diagnoses. The occurrence in males (40/77) was slightly higher than that in females (36/77), and the mean age was 8.1 years (1.4-17 years). The most affected age group was between six and 10 years of age with 31 cases (40.2%). Regarding the anatomical classification, the multicentric form was the most prevalent, accounting for 71.4% (55/77) of the diagnoses. In 40 cases that immunophenotyping was performed, B-cell lymphomas represented 62.5% of the diagnoses (25/40), while T-cell lymphomas corresponded to 37.5% of the diagnoses (15/40). The degree of malignancy according to the modified Kiel classification was low in 35% of lymphomas (14/40) and high in 65% of cases (26/40). The multicentric form was more frequent in the region of influence of the LRD-UFPel. Identification of the immunophenotype can improve the quality of life and survival in affected dogs since it allows the most appropriate treatment for each patient.(AU)

O linfoma é uma neoplasia com origem nos tecidos hematopoiéticos sólidos e é um dos tumores mais frequentes em cães. O objetivo do presente trabalho foi efetuar um estudo retrospectivo dos linfomas caninos recebidos no Laboratório Regional de Diagnóstico, da Faculdade de Veterinária da Universidade Federal de Pelotas (LRD-UFPel) de 2000 a 2017, determinando a epidemiologia e a distribuição anatômica, bem como os aspectos histopatológicos e imuno-histoquímicos de cada caso de acordo com a classificação de Kiel adaptada. Foram revisados os protocolos de necropsias e biópsias recebidos no laboratório identificando-se 77 casos de cães com diagnóstico de linfoma. A doença afetou cães sem raça definida em 37,7% (29/77) dos casos, enquanto os cães com raças definidas tiveram 58,4% (45/77) dos diagnósticos. A ocorrência em machos (40/77) foi discretamente maior do que em fêmeas (36/77) e a idade média foi de 8,1 anos (1,4-17 anos). A faixa etária mais acometida foi entre seis e 10 anos de idade com 31 casos (40,2%). Quanto à classificação anatômica a forma multicêntrica foi a mais prevalente atingindo 71,4% (55/77) dos diagnósticos. Em 40 casos em que a imunofenotipagem foi realizada, os linfomas de células B representaram 62,5% dos casos (25/40), enquanto os linfomas de células T equivaleram a 37,5% dos diagnósticos (15/40). O grau de malignidade de acordo com a classificação de Kiel modificada foi baixo em 35% dos linfomas (14/40) e alto em 65% dos casos (26/40). Conclui-se que a forma multicêntrica é mais frequente na região de influência do LRD-UFPel e que a identificação do imunofenótipo pode melhorar a qualidade de vida e dar maior sobrevida aos cães afetados uma vez que permite o tratamento mais adequado para cada caso.(AU)

Canine lymphomas diagnosed in southern Brazil from 2000 to 2017: epidemiology and immunophenotype / Linfomas caninos diagnosticados na região Sul do Rio Grande do Sul de 2000 a 2017: epidemiologia e imunofenótipo

Lymphoma is a neoplasm that originates from solid hematopoietic tissues and is one of the most common tumors in dogs. The goal of the present study was to perform a retrospective study of canine lymphomas diagnosed at the "Laboratório Regional de Diagnóstico", at the "Faculdade de Veterinária" of the "Universidade Federal de Pelotas" (LRD-UFPel) from 2000 to 2017, to determine the epidemiology and anatomical distribution, and to evaluate the histopathological and immunohistochemical aspects of each case according to the adapted Kiel classification. The protocols for necropsies and biopsies in the laboratory were reviewed. Lymphoma was diagnosed in 77 dogs. Approximately 37.7% (29/77) of affected dogs had no defined breed, while dogs with defined breeds accounted for 58.4% (45/77) of the diagnoses. The occurrence in males (40/77) was slightly higher than that in females (36/77), and the mean age was 8.1 years (1.4-17 years). The most affected age group was between six and 10 years of age with 31 cases (40.2%). Regarding the anatomical classification, the multicentric form was the most prevalent, accounting for 71.4% (55/77) of the diagnoses. In 40 cases that immunophenotyping was performed, B-cell lymphomas represented 62.5% of the diagnoses (25/40), while T-cell lymphomas corresponded to 37.5% of the diagnoses (15/40). The degree of malignancy according to the modified Kiel classification was low in 35% of lymphomas (14/40) and high in 65% of cases (26/40). The multicentric form was more frequent in the region of influence of the LRD-UFPel. Identification of the immunophenotype can improve the quality of life and survival in affected dogs since it allows the most appropriate treatment for each patient.(AU)

O linfoma é uma neoplasia com origem nos tecidos hematopoiéticos sólidos e é um dos tumores mais frequentes em cães. O objetivo do presente trabalho foi efetuar um estudo retrospectivo dos linfomas caninos recebidos no Laboratório Regional de Diagnóstico, da Faculdade de Veterinária da Universidade Federal de Pelotas (LRD-UFPel) de 2000 a 2017, determinando a epidemiologia e a distribuição anatômica, bem como os aspectos histopatológicos e imuno-histoquímicos de cada caso de acordo com a classificação de Kiel adaptada. Foram revisados os protocolos de necropsias e biópsias recebidos no laboratório identificando-se 77 casos de cães com diagnóstico de linfoma. A doença afetou cães sem raça definida em 37,7% (29/77) dos casos, enquanto os cães com raças definidas tiveram 58,4% (45/77) dos diagnósticos. A ocorrência em machos (40/77) foi discretamente maior do que em fêmeas (36/77) e a idade média foi de 8,1 anos (1,4-17 anos). A faixa etária mais acometida foi entre seis e 10 anos de idade com 31 casos (40,2%). Quanto à classificação anatômica a forma multicêntrica foi a mais prevalente atingindo 71,4% (55/77) dos diagnósticos. Em 40 casos em que a imunofenotipagem foi realizada, os linfomas de células B representaram 62,5% dos casos (25/40), enquanto os linfomas de células T equivaleram a 37,5% dos diagnósticos (15/40). O grau de malignidade de acordo com a classificação de Kiel modificada foi baixo em 35% dos linfomas (14/40) e alto em 65% dos casos (26/40). Conclui-se que a forma multicêntrica é mais frequente na região de influência do LRD-UFPel e que a identificação do imunofenótipo pode melhorar a qualidade de vida e dar maior sobrevida aos cães afetados uma vez que permite o tratamento mais adequado para cada caso.(AU)

Hematological and immunological development from birth to six months of age in Holstein calves / Desenvolvimento hematológico e imunológico em bezerros Holandeses do nascimento aos seis meses de idade

The hematological and immunological development of calves from birth to 6 months of age was performed by hemogram and cellular phenotype. Ten male Holstein calves were assessed in 13 moments: before colostrum intake (D0), every 2 days until the 10th day of life (D2 to D10), at the 15th day after birth (D15), and then monthly up to 6 months (D180). Calves presented hemoconcentration on day (D) 0 and showed a gradual decrease in hematimetric rates until D180. The inversion of the neutrophilic for lymphocytic profile was observed on D4. The percentage of CD3+ cells on D10 was higher than D30 up to D180. The number of CD4+ and CD8+ cells did not change between time points. The number of CD21+ lymphocytes was significantly higher at early time points of D0 up to D15, compared on D30 until D150. In conclusion, the neonatal period was marked by stress leukogram in the first 4 days, and low number of B lymphocytes. These might be risk factors for bacterial infections responsible for navel inflammation and diarrhea. The increase in the number of B cells from 30 days of age demonstrated that the calves were functional and able to generate an immune response.(AU)

O desenvolvimento hematológico e o imunológico de bezerros do nascimento aos seis meses de idade foram avaliados por meio hemograma e imunofenotipagem dos linfócitos. Para tanto, 10 bezerros machos Holandeses foram avaliados em 13 momentos: antes da ingestão do colostro (D0), a cada dois dias até o 10º dia (D2 a D10), 15º dia (D15) e, em seguida, mensalmente até seis meses de idade (D180). Os bezerros apresentaram hemoconcentração em D0 e decréscimo gradual dos índices hematimétricos até D180. A inversão do perfil neutrofílico para linfocítico foi observada em D4. Os valores de CD3+ (%) em D10 foram maiores que aqueles observados em D30 até D180. CD4+ e CD8+ não apresentaram alterações entre os momentos. Os linfócitos B CD21+ aumentaram significativamente a partir dos primeiros momentos D0 até D15, em comparação com os momentos subsequentes D30 até D150. Em conclusão, o período neonatal foi caracterizado pelo leucograma de estresse nos primeiros quatro dias e o baixo número de linfócitos B. Esses fatos podem ser considerados como fatores de risco para infecções bacterianas responsáveis pelas inflamações umbilicais e pela diarreia. Por outro lado, o aumento de células B após os 30 dias de vida demonstrou que os bezerros foram capazes de gerar uma resposta imune.(AU)

Hematological and immunological development from birth to six months of age in Holstein calves / Desenvolvimento hematológico e imunológico em bezerros Holandeses do nascimento aos seis meses de idade

The hematological and immunological development of calves from birth to 6 months of age was performed by hemogram and cellular phenotype. Ten male Holstein calves were assessed in 13 moments: before colostrum intake (D0), every 2 days until the 10th day of life (D2 to D10), at the 15th day after birth (D15), and then monthly up to 6 months (D180). Calves presented hemoconcentration on day (D) 0 and showed a gradual decrease in hematimetric rates until D180. The inversion of the neutrophilic for lymphocytic profile was observed on D4. The percentage of CD3+ cells on D10 was higher than D30 up to D180. The number of CD4+ and CD8+ cells did not change between time points. The number of CD21+ lymphocytes was significantly higher at early time points of D0 up to D15, compared on D30 until D150. In conclusion, the neonatal period was marked by stress leukogram in the first 4 days, and low number of B lymphocytes. These might be risk factors for bacterial infections responsible for navel inflammation and diarrhea. The increase in the number of B cells from 30 days of age demonstrated that the calves were functional and able to generate an immune response.(AU)

O desenvolvimento hematológico e o imunológico de bezerros do nascimento aos seis meses de idade foram avaliados por meio hemograma e imunofenotipagem dos linfócitos. Para tanto, 10 bezerros machos Holandeses foram avaliados em 13 momentos: antes da ingestão do colostro (D0), a cada dois dias até o 10º dia (D2 a D10), 15º dia (D15) e, em seguida, mensalmente até seis meses de idade (D180). Os bezerros apresentaram hemoconcentração em D0 e decréscimo gradual dos índices hematimétricos até D180. A inversão do perfil neutrofílico para linfocítico foi observada em D4. Os valores de CD3+ (%) em D10 foram maiores que aqueles observados em D30 até D180. CD4+ e CD8+ não apresentaram alterações entre os momentos. Os linfócitos B CD21+ aumentaram significativamente a partir dos primeiros momentos D0 até D15, em comparação com os momentos subsequentes D30 até D150. Em conclusão, o período neonatal foi caracterizado pelo leucograma de estresse nos primeiros quatro dias e o baixo número de linfócitos B. Esses fatos podem ser considerados como fatores de risco para infecções bacterianas responsáveis pelas inflamações umbilicais e pela diarreia. Por outro lado, o aumento de células B após os 30 dias de vida demonstrou que os bezerros foram capazes de gerar uma resposta imune.(AU)

Mieloma múltiplo em cão: Relato de Caso / Multiple myeloma in dog: Case Report

Multiple myeloma is a neoplasm characterized by the proliferation of clonal Blymphocytes in the bone marrow, resulting in disordered production ofimmunoglobulins, progressively promoting bone destruction, renal failure,hematopoietic suppression, resulting in several clinical signs in dogs. This work aims toreport a case of multiple myeloma in a 11 - year - old female poodle, who was attendedat a veterinary clinic In the radiographic examination, bone lysis was observed in thedistal third of the femur, which led to the clinical suspicion of bone neoplasia, initiallyconfirmed by cytological analysis of the bone marrow of the site affected by thepresence of atypical plasma cells. The animal was then referred for surgery to performovarian resection (OSH) and removal of a bone biopsy fragment, which can not beperformed. The clinical and cytological diagnoses associated with the surgicalprocedure were essential for the prognosis of the patient.

Mieloma múltiplo em cão: Relato de Caso / Multiple myeloma in dog: Case Report

Multiple myeloma is a neoplasm characterized by the proliferation of clonal Blymphocytes in the bone marrow, resulting in disordered production ofimmunoglobulins, progressively promoting bone destruction, renal failure,hematopoietic suppression, resulting in several clinical signs in dogs. This work aims toreport a case of multiple myeloma in a 11 - year - old female poodle, who was attendedat a veterinary clinic In the radiographic examination, bone lysis was observed in thedistal third of the femur, which led to the clinical suspicion of bone neoplasia, initiallyconfirmed by cytological analysis of the bone marrow of the site affected by thepresence of atypical plasma cells. The animal was then referred for surgery to performovarian resection (OSH) and removal of a bone biopsy fragment, which can not beperformed. The clinical and cytological diagnoses associated with the surgicalprocedure were essential for the prognosis of the patient.(AU)

Isolation, culture, characterization and cryopreservation of stem cells derived from amniotic mesenchymal layer and umbilical cord tissue of bovine fetuses / Isolamento, cultura, caracterização e criopreservação de células tronco derivadas de camada amniótica mesenquimal e de tecido do cordão umbilical de fetuses bovinos

Stem cells are undifferentiated cells with a high proliferation potential. These cells can be characterized by their in vivo ability to self-renew and to differentiate into specialized cell lines. The most used stem cell types, in both human and veterinary fields, are the mesenchymal stem cells (MSC) derived from bone marrow and adipose tissue. Nowadays, there is a great interest in using stem cells derived from fetal tissues, such as amniotic membrane (AM) and umbilical cord tissue (UCT), which can be obtained non-invasively at delivery time. Due to the scarcity of studies in bovine species, the aim of this study was to isolate, characterize, differentiate and cryopreserve MSC derived from the mesenchymal layer of amniotic membrane (AM), for the first time, and umbilical cord tissue (UCT) of dairy cow neonates after assisted delivery (AD) and from fetus at initial third of pregnancy (IT) obtained in slaughterhouse. Cells were isolated by enzymatic digestion of the tissue fragments with 0.1% collagenase solution. Six samples of AM and UCT at delivery time and six samples of AM and UCT at first trimester of pregnancy were subjected to morphology evaluation, imunophenotype characterization, in vitro osteogenic, adipogenic and chondrogenic differentiation and viability analysis after cryopreservation. All samples showed adherence to plastic and fibroblast-like morphology. Immunocytochemistry revealed expression of CD 44, NANOG and OCT-4 and lack of expression of MHC II in MSC from all samples. Flow cytometry demonstrated that cells from all samples expressed CD 44, did not or low expressed CD 34 (AM: IT-0.3%a, AD-3.4%b; UCT: 0.4%, 1.4%) and MHC II (AM: IT-1.05%a, AD-9.7%b; UCT: IT-0.7%a, AD-5.7%b). They were also capable of trilineage mesenchymal differentiation and showed 80% viability after cryopreservation. According to the results, bovine AM and UCT-derived cells, either obtained at delivery time or from slaughterhouse, are a painless and non-invasive source of MSC and can be used for stem cell banking.(AU)

As células tronco mesenquimais (CTMs) estão presentes na maioria dos tecidos adultos e possuem grande capacidade de multiplicação. Quando cultivadas in vitro são capazes de se auto renovar e dar origem a novos tipos celulares. As células tronco mais utilizadas, tanto na medicina humana como na medicina veterinária são as células tronco mesenquimais derivadas da medula óssea e do tecido adiposo. Atualmente, uma grande tendência para a utilização de CTMs obtidas de tecidos fetais, como a membrana amniótica (MA), matriz extravascular do cordão umbilical (TCU) e sangue do cordão umbilical (SCU) pode ser observada, já que estas fontes podem ser colhidas no momento do parto por uma técnica não invasiva. Dessa forma, o objetivo deste estudo foi isolar, caracterizar, diferenciar e criopreservar CTMs obtidas de MA e TCU de fetos bovinos colhidos no momento do parto e de fetos do terço inicial da gestação em abatedouro-frigorífico. As células foram recuperadas por meio de digestão enzimática tecidual, realizada com solução de colagenase 0,1%. Foram colhidas amostras de MA e TCU no momento do parto (n=6) e de MA e TCU no terço inicial de gestação (n=6), as quais foram submetidas às análises morfológicas, imunofenotípica por imunocitoquímica e citometria de fluxo, diferenciações in vitro nas linhagens osteogênica, adipogênica e condrogênica e ainda, avaliação da viabilidade após a criopreservação por citometria de fluxo. Todas as amostras dos diferentes grupos demonstraram adesão ao plástico e morfologia fibroblastóide. No ensaio imunocitoquímico todas as amostras foram imunomarcadas para CD44, NANOG e Oct-4, com ausência de marcação para MHC II. Na análise imunofenotipica por citometria de fluxo, todas as amostras apresentaram marcação para CD44, ausência de marcação para ou baixíssima expressão de CD34 (MA: TI-0,3%a, PA-3.4%b; TCU: TI-0,4%, PA-1.4%) e nula ou baixa expressão de MHC II (MA: TI-1.5%a, PA-9.7%b; UCT: TI-0.7%a, PA-5.7%b. Apresentaram também capacidade de diferenciação in vitro nas três linhagens mesodermais e quando analisadas pós criopreservação por citometria de fluxo, todas as amostras apresentaram viabilidade de 80%. Estes resultados indicam que MA e TCU, obtidos tanto no momento de parto como em abatedouro, de fetos bovinos podem ser utilizados como fonte não invasiva e indolor de CTMs e possibilitam a formação de bancos de armazenamento de células.(AU)

Isolation, culture, characterization and cryopreservation of stem cells derived from amniotic mesenchymal layer and umbilical cord tissue of bovine fetuses / Isolamento, cultura, caracterização e criopreservação de células tronco derivadas de camada amniótica mesenquimal e de tecido do cordão umbilical de fetuses bovinos

Stem cells are undifferentiated cells with a high proliferation potential. These cells can be characterized by their in vivo ability to self-renew and to differentiate into specialized cell lines. The most used stem cell types, in both human and veterinary fields, are the mesenchymal stem cells (MSC) derived from bone marrow and adipose tissue. Nowadays, there is a great interest in using stem cells derived from fetal tissues, such as amniotic membrane (AM) and umbilical cord tissue (UCT), which can be obtained non-invasively at delivery time. Due to the scarcity of studies in bovine species, the aim of this study was to isolate, characterize, differentiate and cryopreserve MSC derived from the mesenchymal layer of amniotic membrane (AM), for the first time, and umbilical cord tissue (UCT) of dairy cow neonates after assisted delivery (AD) and from fetus at initial third of pregnancy (IT) obtained in slaughterhouse. Cells were isolated by enzymatic digestion of the tissue fragments with 0.1% collagenase solution. Six samples of AM and UCT at delivery time and six samples of AM and UCT at first trimester of pregnancy were subjected to morphology evaluation, imunophenotype characterization, in vitro osteogenic, adipogenic and chondrogenic differentiation and viability analysis after cryopreservation. All samples showed adherence to plastic and fibroblast-like morphology. Immunocytochemistry revealed expression of CD 44, NANOG and OCT-4 and lack of expression of MHC II in MSC from all samples. Flow cytometry demonstrated that cells from all samples expressed CD 44, did not or low expressed CD 34 (AM: IT-0.3%a, AD-3.4%b; UCT: 0.4%, 1.4%) and MHC II (AM: IT-1.05%a, AD-9.7%b; UCT: IT-0.7%a, AD-5.7%b). They were also capable of trilineage mesenchymal differentiation and showed 80% viability after cryopreservation. According to the results, bovine AM and UCT-derived cells, either obtained at delivery time or from slaughterhouse, [...](AU)

As células tronco mesenquimais (CTMs) estão presentes na maioria dos tecidos adultos e possuem grande capacidade de multiplicação. Quando cultivadas in vitro são capazes de se auto renovar e dar origem a novos tipos celulares. As células tronco mais utilizadas, tanto na medicina humana como na medicina veterinária são as células tronco mesenquimais derivadas da medula óssea e do tecido adiposo. Atualmente, uma grande tendência para a utilização de CTMs obtidas de tecidos fetais, como a membrana amniótica (MA), matriz extravascular do cordão umbilical (TCU) e sangue do cordão umbilical (SCU) pode ser observada, já que estas fontes podem ser colhidas no momento do parto por uma técnica não invasiva. Dessa forma, o objetivo deste estudo foi isolar, caracterizar, diferenciar e criopreservar CTMs obtidas de MA e TCU de fetos bovinos colhidos no momento do parto e de fetos do terço inicial da gestação em abatedouro-frigorífico. As células foram recuperadas por meio de digestão enzimática tecidual, realizada com solução de colagenase 0,1%. Foram colhidas amostras de MA e TCU no momento do parto (n=6) e de MA e TCU no terço inicial de gestação (n=6), as quais foram submetidas às análises morfológicas, imunofenotípica por imunocitoquímica e citometria de fluxo, diferenciações in vitro nas linhagens osteogênica, adipogênica e condrogênica e ainda, avaliação da viabilidade após a criopreservação por citometria de fluxo. Todas as amostras dos diferentes grupos demonstraram adesão ao plástico e morfologia fibroblastóide. No ensaio imunocitoquímico todas as amostras foram imunomarcadas para CD44, NANOG e Oct-4, com ausência de marcação para MHC II. Na análise imunofenotipica por citometria de fluxo, todas as amostras apresentaram marcação para CD44, ausência de marcação para ou baixíssima expressão de CD34 (MA: TI-0,3%a, PA-3.4%b; TCU: TI-0,4%, PA-1.4%) e nula ou baixa expressão de MHC II (MA: TI-1.5%a, PA-9.7%b; UCT: TI-0.7%a, PA-5.7%b. [...](AU)

Quantificação de subpopulações linfocitárias no sangue periférico de cães sadios adultos / Lymphocyte subsets in peripheral blood of adult healthy dogs

A citometria de fluxo vem se firmando como uma ferramenta útil na prática médico-veterinária, particularmente na clínica de pequenos animais. Tal conhecimento tem ensejado o estabelecimento de valores fisiológicos para diferentes subpopulações linfocitárias, indispensáveis à compreensão da dinâmica da celularidade linfóide, em diversas situações patológicas. Assim sendo, o presente ensaio teve como objetivo imunomarcar, por intermédio da técnica citométrica, as subpopulações linfocitárias CD5+CD4+, CD5+CD8+ e CD21+, em quatro diferentes raças de cães domésticos e sadios, de tal forma a agregar informações sobre o perfil imunológico das diferentes raças. Foram utilizados 40 cães adultos (2-7 anos), machos e fêmeas, das raças Beagle (G1, n=10), Golden Retriever (G2, n=10), Bulldog Inglês (G3, n=10) e sem raça definida (SRD) (G4, n=10). As colheitas de sangue foram realizadas por venipunção jugular, utilizando-se sistema de frascos a vácuo (K2-EDTA). O hemograma e o processamento das amostras para citometria de fluxo foram realizados num prazo máximo de 24 horas após a colheita do sangue. As amostras foram analisadas no citofluorômetro FACSCANTO (Becton Dickinson, San Jose, CA, USA). Utilizou-se o programa FACSDiva (Becton Dickinson, San Jose, CA, USA), para identificar e quantificar as células CD5+CD4+, CD5+CD8+ e CD21+, que forneceu o histograma e respectiva tabela com a quantidade de células detectadas pela imunofenotipagem. Os dados obtidos para contagem de linfócitos T auxiliares, T citotóxicos/supressores e linfócitos B foram tabulados e submetidos a Análise de Variância pelo teste F. O teste de Tukey a 5% de probabilidade foi utilizado para comparação das médias entre as diferentes raças de cães. Os valores médios de contagens de células CD5+CD8+ no sangue periférico do G1, G2, G3 e G4 foram de 155, 206, 544 e 503 células/µL, respectivamente. Os valores médios de contagens de células CD5+CD4+ no sangue periférico do G1, G2, G3 e G4 foram de 746, 642, 1101 e 855 células/µL, respectivamente. Os valores médios de contagens de células CD21+ no sangue periférico do G1, G2, G3 e G4 foram de 171, 299, 494 e 403 células/µL, respectivamente. Assim sendo, o número médio de células obtido para a subpopulação de linfócitos T citotóxicos foi significativamente maior (p<0,05) nos Bulldogs Ingleses e cães SRD, comparativamente aqueles encontrados nos grupos de Beagles e Golden Retrievers. Ademais, observou-se que o número médio de células obtido para a subpopulação de linfócitos B foi significavamente maior (p<0,05) nos Bulldogs Ingleses, quando comparado àquele dos Beagles.(AU)

Flow cytometry has established itself as a useful tool in veterinary practice, particularly for small animals practice. Such knowledge has made necessary the establishment of physiological values for different lymphocyte subpopulations, indispensable to understanding the dynamics of lymphoid cellularity in various pathological conditions. In this sense, the objective of this study was to determine, through cytometric technique, lymphocyte subsets of CD5+CD4+, CD5+CD8+ and CD21+ in four different breeds of domestic dogs, so to add information about the immunological profile of different breeds. A total of 40 adult dogs were used (2-7 years), being males and females of Beagles (G1, n=10), Golden Retrievers (G2, n=10), English Bulldogs (G3, n=10) and crossbreed dogs (G4, n=10). Blood samples were collected by jugular venipuncture, using vacuum flasks system (K2-EDTA). Hemogram and processing of samples for flow cytometry were performed within a maximum of 24 hours after blood collection. Samples were analyzed using FACSCanto device (Becton Dickinson, San Jose, CA, USA). FACSDiva software (Becton Dickinson, San Jose, CA, USA) was used to identify and quantify the CD5+ CD4+, CD5+CD8+, and CD21+, which provided the histogram and the respective table with the number of cells identified by immunophenotyping. The data obtained for T helper lymphocyte count, T cytotoxic/suppressor lymphocyte count and B lymphocytes were tabulated and submitted to analysis of variance by F test. Tukey test at 5% probability was used to compare means between the different breeds of dogs. The average values for CD5+CD8+ cell count in peripheral blood in G1, G2, G3 and G4 were of 155, 206, 544 and 503 cells/uL, respectively. The average values for CD5+CD4+ cell count in peripheral blood in G1, G2, G3 and G4 were of 746, 642, 855 and 1101 cells/uL, respectively. The average values for CD21 + cell count in peripheral blood for G1, G2, G3 and G4 were of 171, 299, 494 and 403 cells/uL, respectively. Therefore, The average number of cells obtained for the subsets of cytotoxic T lymphocytes was significantly higher (p<0,05) in the British Bulldogs and crossbreed dogs compared to those found in Beagles and Golden Retrievers. Furthermore, it was observed that the average number of cells obtained for the subsets of B lymphocytes was significantly higher (p<0,05) in English Bulldogs compared to that of Beagles.(AU)

Quantificação de subpopulações linfocitárias no sangue periférico de cães sadios adultos / Lymphocyte subsets in peripheral blood of adult healthy dogs

A citometria de fluxo vem se firmando como uma ferramenta útil na prática médico-veterinária, particularmente na clínica de pequenos animais. Tal conhecimento tem ensejado o estabelecimento de valores fisiológicos para diferentes subpopulações linfocitárias, indispensáveis à compreensão da dinâmica da celularidade linfóide, em diversas situações patológicas. Assim sendo, o presente ensaio teve como objetivo imunomarcar, por intermédio da técnica citométrica, as subpopulações linfocitárias CD5+CD4+, CD5+CD8+ e CD21+, em quatro diferentes raças de cães domésticos e sadios, de tal forma a agregar informações sobre o perfil imunológico das diferentes raças. Foram utilizados 40 cães adultos (2-7 anos), machos e fêmeas, das raças Beagle (G1, n=10), Golden Retriever (G2, n=10), Bulldog Inglês (G3, n=10) e sem raça definida (SRD) (G4, n=10). As colheitas de sangue foram realizadas por venipunção jugular, utilizando-se sistema de frascos a vácuo (K2-EDTA). O hemograma e o processamento das amostras para citometria de fluxo foram realizados num prazo máximo de 24 horas após a colheita do sangue. As amostras foram analisadas no citofluorômetro FACSCANTO (Becton Dickinson, San Jose, CA, USA). Utilizou-se o programa FACSDiva (Becton Dickinson, San Jose, CA, USA), para identificar e quantificar as células CD5+CD4+, CD5+CD8+ e CD21+, que forneceu o histograma e respectiva tabela com a quantidade de células detectadas pela imunofenotipagem. Os dados obtidos para contagem de linfócitos T auxiliares, T citotóxicos/supressores e linfócitos B foram tabulados e submetidos a Análise de Variância pelo teste F. O teste de Tukey a 5% de probabilidade foi utilizado para comparação das médias entre as diferentes raças de cães. Os valores médios de contagens de células CD5+CD8+ no sangue periférico do G1, G2, G3 e G4 foram de 155, 206, 544 e 503 células/µL, respectivamente. Os valores médios de contagens de células CD5+CD4+ no sangue periférico do G1, G2, G3 e G4 foram de 746, 642, 1101 e 855 células/µL, respectivamente. Os valores médios de contagens de células CD21+ no sangue periférico do G1, G2, G3 e G4 foram de 171, 299, 494 e 403 células/µL, respectivamente. Assim sendo, o número médio de células obtido para a subpopulação de linfócitos T citotóxicos foi significativamente maior (p<0,05) nos Bulldogs Ingleses e cães SRD, comparativamente aqueles encontrados nos grupos de Beagles e Golden Retrievers. Ademais, observou-se que o número médio de células obtido para a subpopulação de linfócitos B foi significavamente maior (p<0,05) nos Bulldogs Ingleses, quando comparado àquele dos Beagles.(AU)

Flow cytometry has established itself as a useful tool in veterinary practice, particularly for small animals practice. Such knowledge has made necessary the establishment of physiological values for different lymphocyte subpopulations, indispensable to understanding the dynamics of lymphoid cellularity in various pathological conditions. In this sense, the objective of this study was to determine, through cytometric technique, lymphocyte subsets of CD5+CD4+, CD5+CD8+ and CD21+ in four different breeds of domestic dogs, so to add information about the immunological profile of different breeds. A total of 40 adult dogs were used (2-7 years), being males and females of Beagles (G1, n=10), Golden Retrievers (G2, n=10), English Bulldogs (G3, n=10) and crossbreed dogs (G4, n=10). Blood samples were collected by jugular venipuncture, using vacuum flasks system (K2-EDTA). Hemogram and processing of samples for flow cytometry were performed within a maximum of 24 hours after blood collection. Samples were analyzed using FACSCanto device (Becton Dickinson, San Jose, CA, USA). FACSDiva software (Becton Dickinson, San Jose, CA, USA) was used to identify and quantify the CD5+ CD4+, CD5+CD8+, and CD21+, which provided the histogram and the respective table with the number of cells identified by immunophenotyping. The data obtained for T helper lymphocyte count, T cytotoxic/suppressor lymphocyte count and B lymphocytes were tabulated and submitted to analysis of variance by F test. Tukey test at 5% probability was used to compare means between the different breeds of dogs. The average values for CD5+CD8+ cell count in peripheral blood in G1, G2, G3 and G4 were of 155, 206, 544 and 503 cells/uL, respectively. The average values for CD5+CD4+ cell count in peripheral blood in G1, G2, G3 and G4 were of 746, 642, 855 and 1101 cells/uL, respectively. The average values for CD21 + cell count in peripheral blood for G1, G2, G3 and G4 were of 171, 299, 494 and 403 cells/uL, respectively. Therefore, The average number of cells obtained for the subsets of cytotoxic T lymphocytes was significantly higher (p<0,05) in the British Bulldogs and crossbreed dogs compared to those found in Beagles and Golden Retrievers. Furthermore, it was observed that the average number of cells obtained for the subsets of B lymphocytes was significantly higher (p<0,05) in English Bulldogs compared to that of Beagles.(AU)