Expression of acetylated histones H3 and H4 and histone deacetylase enzymes HDAC1, HDAC2 and HDAC6 in simple mammary carcinomas of female dogs.

Histone deacetylation is an important mechanism involved in human breast cancer tumorigenesis and recent veterinary oncology studies also demonstrate a similar relationship in some canine neoplasms. The use of HDAC inhibitors in vitro and in vivo has demonstrated antitumor action on several strains of human and animal cancers. The present study aims to correlate the expression of H3K9Ac, H4K12Ac, HDAC1, HDAC2 and HDAC6 in simple mammary carcinomas in dogs with clinicopathological parameters and overall survival time. To this end, 61 samples of simple breast carcinomas were analyzed by the immunohistochemistry technique with subsequent validation of the antibodies by the Western Blot technique. The expressions obtained via a semi-quantitative way were categorized by assigning scores and classified into high or low expressions according to the given score, except for HDAC6, when the marking percentage was considered and subdivided into high and low expressions using the median value. For statistical analysis, the chi-square test or Fisher exact test were used as univariate analysis and correspondence analysis as a multivariate test, in addition to the Kaplan-Meier survival analysis. In the studied samples, the highest frequencies were determined for the high expression proteins H4K12Ac (88.5%), HDAC2 (65.6%) and HDAC6 (56.7%) and the low expression proteins H3K9Ac (73.8%) and HDAC1 (54.1%). An association between the low expression of HDAC1 and the presence of lymph node metastasis (p = 0.035) was indicated by univariate analysis while the high expression of HDAC1 was associated with favorable prognostic factors, such as the absence of lymph node metastasis and low mitotic index by multivariate analysis. Also, by multivariate analysis, the low expression of HDAC6 was correlated with the low expression of Ki67, smaller tumors, and better prognosis factors as well. Protein expression was not correlated with patients' overall survival time (p > 0.05). The high expressions of HDAC2 and HDAC6 in mammary carcinomas in female dogs may be useful information for research involving therapeutic targets with iHDACs since their inhibition favors hyperacetylation and transcription of tumor suppressor genes.

Detection of Trypanosoma vivax in tissues of experimentally infected goats: what is the role of adipose tissue in the life cycle of this protozoon?

Trypanosomiasis, caused by Trypanosoma vivax, is responsible for great economic losses among livestock in Africa and South America. During the life cycle of these parasites, they may present different morphological, metabolic and physiological characteristics depending on the interactions that are encountered at each point of their life cycle. Although T. vivax is frequently reported in the circulation of its mammalian hosts, it has the ability to migrate to the tissues of these individuals. However, this characteristic is poorly understood. In this context, we aimed to investigate the presence of T. vivax and the changes caused in different tissues of experimentally infected goats. Despite the animals were not perfused before tissues collection, using different approaches, we demonstrated its presence in different samples, including in the adipose tissue and skin of infected animals. In addition, a mononuclear inflammatory reaction, mostly characterized by an infiltrate of lymphocytes, plasma cells and macrophages were observed. The results highlight the possibility that, like other trypanosomatids, T. vivax may use these tissues during its life cycle. Future studies aiming to elucidate the length of time for which T. vivax remains active in these sites, and whether it uses these sites as a refuge from trypanocidal drugs, and whether it is capable of recolonizing the blood circulation, are much needed.

Detection of Trypanosoma vivax in tissues of experimentally infected goats: what is the role of adipose tissue in the life cycle of this protozoon? / Detecção de Trypanosoma vivax em tecidos de caprinos experimentalmente infectados: qual é o papel do tecido adiposo no ciclo de vida desse protozoário?

Abstract Trypanosomiasis, caused by Trypanosoma vivax, is responsible for great economic losses among livestock in Africa and South America. During the life cycle of these parasites, they may present different morphological, metabolic and physiological characteristics depending on the interactions that are encountered at each point of their life cycle. Although T. vivax is frequently reported in the circulation of its mammalian hosts, it has the ability to migrate to the tissues of these individuals. However, this characteristic is poorly understood. In this context, we aimed to investigate the presence of T. vivax and the changes caused in different tissues of experimentally infected goats. Despite the animals were not perfused before tissues collection, using different approaches, we demonstrated its presence in different samples, including in the adipose tissue and skin of infected animals. In addition, a mononuclear inflammatory reaction, mostly characterized by an infiltrate of lymphocytes, plasma cells and macrophages were observed. The results highlight the possibility that, like other trypanosomatids, T. vivax may use these tissues during its life cycle. Future studies aiming to elucidate the length of time for which T. vivax remains active in these sites, and whether it uses these sites as a refuge from trypanocidal drugs, and whether it is capable of recolonizing the blood circulation, are much needed.

Resumo A tripanossomíase, causada por Trypanosoma vivax, é responsável por grandes perdas econômicas na bovinocultura da África e da América do Sul. Durante seu ciclo de vida, o parasita pode apresentar diferentes características morfológicas, metabólicas e fisiológicas em função das interações que ele encontra em cada ponto do seu ciclo. Embora o T. vivax seja reportado, frequentemente, na circulação dos seus hospedeiros mamíferos, o protozoário tem a capacidade de migrar para os tecidos desses indivíduos. Entretanto, essa característica é pobremente conhecida. Neste contexto, o objetivo foi verificar a presença, assim como as alterações causadas pelo T. vivax nos diferentes tecidos de caprinos experimentalmente infectados. Apesar dos animais não terem sido perfundidos antes da coleta dos tecidos, utilizando-se diferentes abordagens, foi evidenciada a presença do T. vivax em diferentes amostras teciduais, incluindo no tecido adiposo e pele dos animais infectados. Além disso, foi observada reação inflamatória mononuclear, caracterizada majoritariamente por infiltrado de linfócitos, plasmócitos e macrófagos. Os resultados evidenciam a possibilidade de que, assim como outros tripanossomatídeos, T. vivax pode usar esses tecidos durante o seu ciclo de vida. São necessários futuros estudos, objetivando elucidar o período em que o T. vivax permanece ativo nesses sítios, se ele utiliza esses locais como refúgio das drogas tripanocidas, e se ele é capaz de recolonizar a circulação sanguínea.

Toxicity of a methotrexate metronomic schedule in Wistar rats.

Metronomic chemotherapy is a relevant strategy that uses low doses of antineoplastic drugs for sustained periods to control tumor growth, an alternative frequently utilized in veterinary patients. This work aimed to evaluate the toxic effects of a metronomic oral dose of methotrexate (MTX) for 45 days in tumor-free Wistar rats when compared with control animals. Clinical alterations, body weight, food, and water intake were monitored daily, and bone marrow suppression, hematological, biochemical, and histopathological analyses were performed at three points (days 30, 45, and 60). MTX-treated animals did not demonstrate severe systemic involvement. At 30 days, compared with control animals, MTX-treated animals showed significant leukocytosis (11.9 ± 2.3 vs. 7.8 ± 0.2 106/µL; P < .05) and augmentation of immature myeloid populations from bone marrow (9.0 ± 0.8 vs. 6.5 ± 1.5%; P < .05), and at 60 days, treated animals showed significant neutrophilia (35.0 ± 11.0 vs. 23.00 ± 3.0%; P < .05), depletion of bone marrow lymphocytes (8.2 ± 0.7 vs. 11.5 ± 1.9%; P < .05), and immature myeloid populations (7.2 ± 0.7 vs. 8.3 ± 0.6%; P < .05). At a histopathological level, splenic hypoplasia and respiratory inflammatory lesions were significant when compared with control animals, presenting mild to moderate myelotoxicity, immune suppression, and associated clinical compromise that persisted beyond treatment withdrawal. This suggested that MTX metronomic toxicity should not be neglected owing to the observed residual side-effects and special care should be taken regarding myelosuppression.

Aortic valve endocarditis due to Bartonella clarridgeiae in a dog in Brazil / Endocardite de válvula aórtica por Bartonella clarridgeiae em um cão no Brasil

Abstract We report the first documented case of endocarditis associated with Bartonella clarridgeiae in a dog in Latin America. Infective vegetative valvular aortic endocarditis was diagnosed in a 10-year-old male mixed breed dog. The dog presented grade V/VI systolic and diastolic murmur, hyperthermia, and progressive weight loss. Cardiomegaly and presence of diffuse alveolar pattern in the lung fields were observed in the thorax radiography evaluation. Irregular and hyperechogenic structures adhered to the aortic leaflets, causing obstruction of the left ventricular outflow tract and severe aortic insufficiency, were observed in the echocardiography evaluation. A vegetative, whitish, hardened structure measuring 1.0 cm in diameter was observed in aortic semilunar valve at necropsy. Based on a combination of pre-enrichment insect-based medium liquid culture, quantitative real-time and conventional PCR assays based on nuoG and gltA genes, respectively, followed by sequencing and phylogenetic inferences, B. clarridgeiae DNA was detected in the patient's aortic valve lesions. Clinical, echocardiographic, anatomopathologic and molecular features supported the diagnosis of severe aortic vegetative endocarditis possibly caused by B. clarridgeiae in a dog in Brazil.

Resumo Relatamos o primeiro caso documentado de endocardite associada à Bartonella clarridgeiae em um cão na América Latina. Endocardite aórtica valvar vegetativa infecciosa foi diagnosticada em um cão sem raça definida de 10 anos de idade. O cão apresentou sopro sistólico e diastólico de grau V / VI, hipertermia e perda progressiva de peso. Cardiomegalia e presença de padrão alveolar difuso nos campos pulmonares foram observados na avaliação radiográfica do tórax. Estruturas irregulares e hiperecogênicas aderidas aos folhetos aórticos, causando obstrução da via de saída do ventrículo esquerdo e insuficiência aórtica grave, foram observadas na avaliação ecocardiográfica. À necropsia, foi observada uma estrutura vegetativa, esbranquiçada e endurecida medindo 1,0 cm de diâmetro na válvula semilunar aórtica. Por meio de uma combinação de cultura líquida baseada em meio de pré-enriquecimento de inseto, ensaios de PCR quantitativa em tempo real e convencional baseados nos genes nuoG e gltA, respectivamente, seguidos de sequenciamento e inferências filogenéticas, DNA de B. clarridgeiae foi detectado no tecido valvular lesionado do paciente. O diagnóstico de endocardite vegetativa aórtica grave, possivelmente causado por B. clarridgeiae em um cão no Brasil, foi apoiado por características clínicas, ecocardiográficas, anatomopatológicas e moleculares.

Aortic valve endocarditis due to Bartonella clarridgeiae in a dog in Brazil.

We report the first documented case of endocarditis associated with Bartonella clarridgeiae in a dog in Latin America. Infective vegetative valvular aortic endocarditis was diagnosed in a 10-year-old male mixed breed dog. The dog presented grade V/VI systolic and diastolic murmur, hyperthermia, and progressive weight loss. Cardiomegaly and presence of diffuse alveolar pattern in the lung fields were observed in the thorax radiography evaluation. Irregular and hyperechogenic structures adhered to the aortic leaflets, causing obstruction of the left ventricular outflow tract and severe aortic insufficiency, were observed in the echocardiography evaluation. A vegetative, whitish, hardened structure measuring 1.0 cm in diameter was observed in aortic semilunar valve at necropsy. Based on a combination of pre-enrichment insect-based medium liquid culture, quantitative real-time and conventional PCR assays based on nuoG and gltA genes, respectively, followed by sequencing and phylogenetic inferences, B. clarridgeiae DNA was detected in the patient's aortic valve lesions. Clinical, echocardiographic, anatomopathologic and molecular features supported the diagnosis of severe aortic vegetative endocarditis possibly caused by B. clarridgeiae in a dog in Brazil.

Squamous cell carcinoma of unknown primary origin in a dog presenting with bone metastasis.

A 10-year-old female American Pit Bull dog was diagnosed with metastatic undifferentiated carcinoma of the scapula. Immunohistochemistry showed positive immunoexpression for cytokeratins (AE1/AE3, 34BE12, CK7) and vimentin, confirming squamous cell carcinoma. No evidence of nodules was found in the complete physical examination and imaging procedures conducted. The patient was diagnosed with carcinoma of unknown primary origin. Amputation and adjuvant chemotherapy with doxorubicin and piroxicam were performed, but the patient died of respiratory failure after 737 days of diagnosis. Necropsy confirmed undifferentiated carcinoma infiltrating the lungs and kidneys, and showing the same immunoexpression as the tumor in the scapula. Amputation associated with chemotherapy extended the overall survival time of this patient.

Expression of matrix metalloproteinase-2 and metalloproteinase-9 in the skin of dogs with visceral leishmaniasis.

The skin is the first organ to be infected by the parasite in canine visceral leishmaniasis. The enzyme matrix metalloproteinase (MMP) acts towards degradation of the extracellular matrix (ECM) and modulation of the inflammatory response against many kinds of injuries. The aims of this study were to evaluate the expression of MMP-2 and MMP-9 through immunohistochemistry and zymography on the skin (muzzle, ears, and abdomen) of dogs that were naturally infected by Leishmania spp. and to compare these results with immunodetection of the parasite and with alterations to the dermal ECM. Picrosirius red staining was used to differentiate collagen types I and III in three regions of the skin. The parasite load, intensity of inflammation, and production of MMP-2 (latent) and MMP-9 (active and latent) were higher in the ear and muzzle regions. MMP-9 (active) predominated in the infected group of dogs and its production was significantly different to that of the control group. Macrophages, lymphocytes, and plasma cells predominated in the dermal inflammation and formed granulomas in association with degradation of mature collagen (type I) and with discrete deposition of young collagen (type III). This dermal change was more pronounced in dogs with high parasite load in the skin. Therefore, it was concluded that the greater parasite load and intensity of inflammation in the skin led consequently to increased degradation of mature collagen, caused by increased production of MMPs, particularly active MMP-9, in dogs with visceral leishmaniasis. This host response profile possibly favors systemic dissemination of the parasite.

Immunodetection of hepatic stellate cells in dogs with visceral leishmaniasis.

Hepatic stellate cells (HSC), or Ito cells, store vitamin A when at rest but undergo phenotypic changes in situations of liver injury, which may induce fibrosis, and they may participate in the immune response in the liver. The objective of the present study was to investigate the role of HSC in the livers of dogs with visceral leishmaniasis (VL). Twenty-eight livers from dogs infected with VL that were living in an area endemic for the disease were evaluated, among which 13 were asymptomatic (A) and 15 were symptomatic (S). A control group (C) was formed by five dogs from an area that was not endemic for VL. These organs were subjected to histopathological analysis (Masson's trichrome for fibrosis) and immunohistochemical analysis (Leishmania, smooth-muscle α-actin and TGF-ß). In the livers from the symptomatic dogs, a moderate to severe granulomatous inflammatory reaction was observed in the capsule and in the portal, centrilobular and intralobular regions. In the asymptomatic dogs, there was slight to moderate presence of granulomas, and these were even absent in some dogs. The intensity of hepatic fibrosis was predominantly low in the infected dogs (A and S), and fibrosis was absent in the control group. The immunomarking of HSC in the infected groups (A and S) differed significantly (P = 0.0153) from that of the control group. The symptomatic dogs presented the largest number of positive cells. This group also presented a larger number of parasitized macrophages, but did not differ statistically from the asymptomatic group (P > 0.05). The cytokine TGF-ß was only detected at low levels, and only in the infected animals, but this did not differ from the control group. Immunomarking for HSC was observed mainly in the nuclei of cells present in the hepatic granulomas of symptomatic dogs and in the sinusoids of the asymptomatic dogs. It was concluded that in the livers of dogs with VL, the HSC are activated and participate in the hepatic response to the parasite. The cytokine TGF-ß may be involved in this activation, but in the chronic phase of the infection, this cytokine was detected at lower proportions. It is possible that HSC may also contribute towards chemotaxis of leukocytes for the hepatic compartment, along with other cell types such as Kupffer cells.

Influence of apoptosis on liver and spleen resistance in dogs with visceral leishmaniosis.

The aim of this study was to evaluate apoptosis and parasite load in the liver and spleen of dogs with visceral leishmaniosis (VL), using immunohistochemistry. Liver and spleen samples from 71 dogs with VL were used. The parasite load in the spleen and liver showed significant difference between organs in infected group (P=0.0219). The density of the parasite load in the spleen (median=2.4) was higher than liver (median=0.8). Immunodetection of apoptotic cells was predominant in lymphocytes and differ between the infected and control group in spleen (P=0.0307) and liver (P=0.0346). There was a significant correlation between apoptosis and parasite load (P = 0.0084; r=0.3104) only in the spleen of the infected group, where it was observed that, when increasing the number of apoptotic cells increases the parasitic load. It was concluded that the liver and spleen of infected dogs presented greater numbers of cells undergoing apoptosis (lymphocytes) than the control group, thus suggesting that this process may be contributing towards the survival of Leishmania in these organs, because lymphocyte in apoptosis did not have the ability to present and recognize the antigen, allowing the survival of the parasite.

Influence of apoptosis on liver and spleen resistance in dogs with visceral leishmaniosis / Influência da apoptose na resistência hepática e esplênica de cães com leishmaniose visceral

Abstract The aim of this study was to evaluate apoptosis and parasite load in the liver and spleen of dogs with visceral leishmaniosis (VL), using immunohistochemistry. Liver and spleen samples from 71 dogs with VL were used. The parasite load in the spleen and liver showed significant difference between organs in infected group (P=0.0219). The density of the parasite load in the spleen (median=2.4) was higher than liver (median=0.8). Immunodetection of apoptotic cells was predominant in lymphocytes and differ between the infected and control group in spleen (P=0.0307) and liver (P=0.0346). There was a significant correlation between apoptosis and parasite load (P = 0.0084; r=0.3104) only in the spleen of the infected group, where it was observed that, when increasing the number of apoptotic cells increases the parasitic load. It was concluded that the liver and spleen of infected dogs presented greater numbers of cells undergoing apoptosis (lymphocytes) than the control group, thus suggesting that this process may be contributing towards the survival of Leishmania in these organs, because lymphocyte in apoptosis did not have the ability to present and recognize the antigen, allowing the survival of the parasite.

Resumo O objetivo deste estudo foi avaliar a apoptose e a carga parasitária no fígado e baço de cães com leishmaniose visceral (LV), pela técnica de imuno-histoquímica. Foram utilizadas amostras de fígado e baço de 71 cães com LV. A carga parasitária no baço e fígado mostrou diferença significativa entre os órgãos no grupo infectado (P=0,0219). A densidade da carga de parasita no baço (média=2,4) foi maior do que no fígado (média=0,8). A imunodetecção de células em apoptose foi predominante nos linfócitos, com diferenças entre o grupo infectado e controle no baço (P=0,0307) e fígado (P=0,0346). Houve uma correlação positiva fraca entre apoptose e carga parasitária (P=0,0084; r=0,3104) apenas no baço do grupo infectado, onde observou-se que quando aumentava o número de células em apoptose aumentava a carga parasitária. Concluiu-se que o fígado e o baço de cães infectados apresentam um maior número de células que sofrem apoptose (linfócitos) do que o grupo controle, sugerindo que este processo possa contribuir para a sobrevivência de Leishmania nestes órgãos, pois os linfócitos em apoptose não tiveram a capacidade de apresentar e reconhecer o antígeno, permitindo a sobrevivência do parasita.

Influence of apoptosis on liver and spleen resistance in dogs with visceral leishmaniosis / Influência da apoptose na resistência hepática e esplênica de cães com leishmaniose visceral

Abstract The aim of this study was to evaluate apoptosis and parasite load in the liver and spleen of dogs with visceral leishmaniosis (VL), using immunohistochemistry. Liver and spleen samples from 71 dogs with VL were used. The parasite load in the spleen and liver showed significant difference between organs in infected group (P=0.0219). The density of the parasite load in the spleen (median=2.4) was higher than liver (median=0.8). Immunodetection of apoptotic cells was predominant in lymphocytes and differ between the infected and control group in spleen (P=0.0307) and liver (P=0.0346). There was a significant correlation between apoptosis and parasite load (P = 0.0084; r=0.3104) only in the spleen of the infected group, where it was observed that, when increasing the number of apoptotic cells increases the parasitic load. It was concluded that the liver and spleen of infected dogs presented greater numbers of cells undergoing apoptosis (lymphocytes) than the control group, thus suggesting that this process may be contributing towards the survival of Leishmania in these organs, because lymphocyte in apoptosis did not have the ability to present and recognize the antigen, allowing the survival of the parasite.

Resumo O objetivo deste estudo foi avaliar a apoptose e a carga parasitária no fígado e baço de cães com leishmaniose visceral (LV), pela técnica de imuno-histoquímica. Foram utilizadas amostras de fígado e baço de 71 cães com LV. A carga parasitária no baço e fígado mostrou diferença significativa entre os órgãos no grupo infectado (P=0,0219). A densidade da carga de parasita no baço (média=2,4) foi maior do que no fígado (média=0,8). A imunodetecção de células em apoptose foi predominante nos linfócitos, com diferenças entre o grupo infectado e controle no baço (P=0,0307) e fígado (P=0,0346). Houve uma correlação positiva fraca entre apoptose e carga parasitária (P=0,0084; r=0,3104) apenas no baço do grupo infectado, onde observou-se que quando aumentava o número de células em apoptose aumentava a carga parasitária. Concluiu-se que o fígado e o baço de cães infectados apresentam um maior número de células que sofrem apoptose (linfócitos) do que o grupo controle, sugerindo que este processo possa contribuir para a sobrevivência de Leishmania nestes órgãos, pois os linfócitos em apoptose não tiveram a capacidade de apresentar e reconhecer o antígeno, permitindo a sobrevivência do parasita.

Correlation between systolic function and presence of myofibroblasts in doxorubicin-induced cardiomyopathy / Correlação entre função sistólica e presença de miofibroblastos na cardiomiopatia induzida pela doxorrubicina

ABSTRACT: Cardiotoxicity induced by doroxubicin generates systolic disfunction and myocardial remodeling with presence of myofibroblasts. These cells are thought to be attracted to the injured heart to avoid the development of congestive heart failure. The objective of this study was to evaluate the systolic dysfunction generated by doxorubicin through Doppler echocardiography, and its correlation with the presence of myofibroblasts in the myocardium. Twenty-five New Zealand White rabbits were divided into two groups (control, and treated with doxorubicin). The drug was administered for six weeks; Doppler echocardiography was performed before the first, and after the last administration of doxorubicin. Immuno detection of myofibroblasts was performed by immunohistochemistry. The treated group exhibited significant reduction in systolic function as assessed by Doppler echocardiography, and increased frequency of myofibroblasts, which were present in similar amounts in the left ventricle, interventricular septum, and right ventricle. There was a significant negative correlation between number of myofibroblasts in the interventricular septum and in the left ventricle with systolic function indices, which reveals that the higher the number of fibroblasts, the worst systolic function is in rabbits treated with doxorubicin. Increase in myofibroblast numbers was not sufficient to preserve systolic function.

RESUMO: A cardiotoxicidade induzida pela doxorrubicina gera disfunção sistólica e remodelamento miocárdico, com presença de miofibroblastos. Acredita-se que essas células sejam atraídas para a não evolução do quadro de insuficiência cardíaca congestiva. O objetivo do presente trabalho foi avaliar a função sistólica gerada pela doxorrubicina por meio da ecodopplercadiografia e correlacioná-la com a presença de miofibroblastos no miocárdio. Foram utilizados 25 coelhos da raça Nova Zelândia, alocados em dois grupos (controle e tratados com doxorrubicina). O fármaco foi administrado por seis semanas e a ecodopplercardiografia foi realizada no momento zero e após a última administração da doxorrubicina. A imunodetecção dos miofibroblastos foi realizada por imuno-histoquímica. Houve redução significativa na função sistólica, observada na ecodopplercardiografia e aumento na imundetecção dos miofibroblastos nos animais tratados, na mesma intensidade no ventrículo esquerdo, septo interventricular e ventrículo direito. Houve correlação negativa significativa entre o número de miofibroblastos no septo interventricular e no ventrículo esquerdo com os índices de função sistólica, revelando que quanto mais miofibroblastos presentes, pior é a função sistólica de coelhos tratados com doxorrubicina. O aumento do número de miofibroblastos não foi suficiente para manutenção da função sistólica.

Relationship between the inflammatory infiltrate and the degree of differentiation of the canine cutaneous squamous cell carcinoma.

The inflammatory response may have pro or anti-neoplastic effects in tumors, depending on the histological type and malignancy level of the tumor. This study aimed to evaluate the profile of predominant inflammatory cells by immunohistochemistry in cutaneus squamous cell carcinoma (SCC) of dogs, comparing it with the degree of differentiation of the tumor. Twenty samples of SCC were analyzed. The tumors were histologically classified into two groups, differentiated SCC (SCCd=12) and undifferentiated SCC (SCCu=08). The tumor inflammatory infiltrate was determined by immunohistochemistry, in order to identify macrophages, lymphocytes and plasma cells. The comparison between groups, SCCd and SCCu, was not significant concerning the density of macrophages (P=1.0), T lymphocytes (P=0.335) and plasma cells (P=0.075). However, when comparing the inflammatory infiltrate in each group, the macrophages were the predominant cell type in both groups, a significant difference was found in the SCCd with plasma cells (P<0.0001). In the SCCu the difference occurred with lymphocytes (P<0.05) and plasma cells (P=0.0006). It could be concluded that the presence of inflammation in cases of SCC does not play a role in the differentiation of the neoplasm, since the inflammatory infiltrate was similar in both groups of SCC.

Morphological changes and parasite load of the adrenal from dogs with visceral leishmaniasis.

The objective of this study was to analyze morphological changes and parasite loads in the adrenal gland from 45 dogs with visceral leishmaniasis (VL). The animals were from the Zoonosis Control Center of Araçatuba, state of São Paulo, which is an endemic region for the disease. These animals were euthanized due to positive diagnoses of VL. The dogs were classified into asymptomatic, oligosymptomatic and symptomatic groups. The parasite load was determined by immunohistochemistry, using VL-positive dog hyperimmune serum. Nine dogs showed an inflammatory infiltrate composed, predominantly, of plasma cells and macrophages. However, only eight dogs showed macrophages with amastigote forms of the parasite, immunolabeled in the cytoplasm. The medullary and reticular layers were the most affected areas, possibly due to a favorable microenvironment created by hormones in these regions. The density of parasites in the glandular tissue was not associated with clinical signs of VL (P > 0.05). However, the presence of the parasite was always associated with the presence of a granulomatous inflammatory infiltrate. This gland may not be an ideal place for the parasite's multiplication, but the presence of injuries to the glandular tissue could influence the dog's immune system, thus favoring the parasite's survival in the host's different organs.

Morphological changes in the bone marrow of the dogs with visceral leishmaniasis.

The aim of this study was to evaluate the most frequent lesions in the bone marrow of dogs naturally infected by Leishmania (Leishmania) chagasi. Thirty-three dogs sacrificed at the Zoonosis Control Center of Araçatuba, a municipality endemic for visceral leishmaniasis (VL), were used. The animals were classified as asymptomatic, oligosymptomatic, and symptomatic groups. At the necropsy, bone marrow samples were collected from the femur, fixed, processed, and stained with hematoxylin and eosin. The lesion intensity was classified as mild, moderate, or severe. The parasite load was determined using immunohistochemistry. The most important lesions consisted of multifocal to diffuse granulomas, megakaryocytic dysplasia, and medullary aplasia. There were no statistical differences between the three clinical groups regarding parasite load and lesion intensity. Asymptomatic dogs also presented high parasitism in the bone marrow as dogs with clinical signs of VL. It was concluded that, regardless of clinical group, the bone marrow is a site for multiplication of Leishmania chagasi. Possibly, the bone marrow dysplasia may arise from the presence of many parasitized and activated macrophages in this organ. Consequently, it affects the profile of hematopoietic cells in the bone marrow and systemic circulation.

Morphological changes and parasite load of the adrenal from dogs with visceral leishmaniasis / Alterações morfológicas e carga parasitária da adrenal de cães com leishmaniose visceral

The objective of this study was to analyze morphological changes and parasite loads in the adrenal gland from 45 dogs with visceral leishmaniasis (VL). The animals were from the Zoonosis Control Center of Araçatuba, state of São Paulo, which is an endemic region for the disease. These animals were euthanized due to positive diagnoses of VL. The dogs were classified into asymptomatic, oligosymptomatic and symptomatic groups. The parasite load was determined by immunohistochemistry, using VL-positive dog hyperimmune serum. Nine dogs showed an inflammatory infiltrate composed, predominantly, of plasma cells and macrophages. However, only eight dogs showed macrophages with amastigote forms of the parasite, immunolabeled in the cytoplasm. The medullary and reticular layers were the most affected areas, possibly due to a favorable microenvironment created by hormones in these regions. The density of parasites in the glandular tissue was not associated with clinical signs of VL (P > 0.05). However, the presence of the parasite was always associated with the presence of a granulomatous inflammatory infiltrate. This gland may not be an ideal place for the parasite's multiplication, but the presence of injuries to the glandular tissue could influence the dog's immune system, thus favoring the parasite's survival in the host's different organs.

O objetivo deste estudo foi analisar as alterações morfológicas e a carga parasitária da glândula adrenal de 45 cães com leishmaniose visceral (LV). Os animais eram provenientes do Centro de Controle de Zoonoses (CCZ) de Araçatuba (SP), região endêmica para a doença. Esses animais são submetidos à eutanásia, devido ao diagnóstico positivo para LV. Os cães foram classificados nos grupos assintomático, oligossintomático e sintomático. A determinação da carga parasitária foi feita por imuno-histoquímica, com utilização de soro hiperimune de cão positivo para LV. Em nove cães, verificou-se um infiltrado inflamatório, composto predominantemente por plasmócitos e macrófagos. Entretanto, apenas oito cães apresentaram macrófagos com formas amastigotas do parasito imunomarcadas em seu citoplasma. As camadas medular e reticulada foram as mais afetadas, possivelmente por um microambiente favorável criado pelos hormônios nestas regiões. A densidade de parasitos no tecido glandular não foi relacionada com os sinais clínicos de LV (P > 0,05). No entanto, a presença do parasito sempre esteve associada à presença de infiltrado inflamatório granulomatoso. Possivelmente, essa glândula não é um sítio ideal para a multiplicação do protozoário, mas a presença de injúrias no tecido glandular poderia influenciar o sistema imune do cão, favorecendo a sobrevivência do parasito nos diferentes órgãos do hospedeiro.

Expression of matrix metalloproteases-2 and -9 in horse hoof laminae after intestinal obstruction, with or without Hydrocortisone treatment / Expressão de metaloproteinases 2 e 9 no tecido laminar do casco de equinos após obstrução intestinal e tratamento com hidrocortisona

Twenty horses were used in the experiment, for composed control group, (Cg) instrumented group, (Ig;without intestinal obstruction), treated group (Tg;submitted to intestinal obstruction and hydrocortisone treatment) and non-treated group (Ntg;submitted to intestinal obstruction without treatment). Immunohistochemistry and zymography techniques were used for researches on MMPs 2 and 9 in horse hoof laminae. There was an increase in the expression of MMP-2 in animals of Tg and Ntg. MMP-9 increased on animals from groups Ntg and Ig, however there was no rise of this MMP on the Tg when compared to the other groups in the immunohistochemistry analysis. Based on the results, it was observed that the intestinal injury caused by enterotomy and intestinal obstruction raise the quantities of MMPs in the hoof laminae.

Vinte cavalos foram usados no experimento: para compor o grupo controle (Cg), grupo instrumentado, Ig (sem obstrução intestinal), grupo tratado, Tg (submetidos à obstrução intestinal e tratamento com hidrocortisona) e grupo não tratado, Ntg (submetidos à obstrução intestinal, sem tratamento. Técnicas de zimografia e imunoistoquímica foram utilizadas para pesquisa de MMP-2 e MMP-9 no tecido laminar do casco dos equinos. Houve um aumento na expressão de MMP-2 nos animais dos grupos Tg e Ntg. A MMP-9 aumentou nos animais dos grupos Ig e Ntg. Houve aumento desta MMP no Tg quando comparado aos demais grupos na análise por zimografia. Observou-se que a injúria intestinal, causada pela enterotomia e obstrução intestinal, eleva a quantidade de MMPs no tecido laminar do casco.

Immune response pattern of the popliteal lymph nodes of dogs with visceral leishmaniasis.

The present study aimed to estimate the cell response and parasite load in the popliteal lymph nodes of dogs with visceral leishmaniasis (VL), comparing these findings with the clinical staging of the disease. From the necropsy, 33 dogs were classified as symptomatic (S), asymptomatic (A), or oligosymptomatic (O). Cytology and histopathology were used to determine any presence of microscopic lesions and immunohistochemistry, for parasite load. Dog hyperimmune serum was used as the primary antibody. The inflammatory infiltrate in lymph nodes consisted of macrophages and plasmocytes. The granulomas invaded the trabecular and sinusoid regions and sometimes compressed the lymphocytes of the cortical region (atrophy) and medullary cord cells. Parasite load intensity was unrelated to the density of the macrophages infiltrating the lymph node. Significant differences in parasite load (P < 0.05) were observed between the three groups of infected dogs. Follicular hyperplasia of the cortical region occurred among A and O, while follicular atrophy predominated among S. The parasite load was the greatest among S, followed by O. It can be concluded that, regardless of clinical condition, the most evident cell response consisted of macrophages and plasmocytes. Lymphoid atrophy was observed among animals with intense granulomatous reaction and high parasite load, such as among the symptomatic dogs (P < 0.05). Likewise, the oligosymptomatic dogs also presented high density of parasites in the lymph nodes. Thus, we can confirm that dogs with clinical manifestations of VL have an immune system that is less effective for controlling infection by Leishmania chagasi, thereby favoring parasite multiplication.

Halicephalobus gingivalis (H. deletrix) in the brain of a horse

A 10-year-old Mangalarga gelding with rhabditiform nematode infection in the brain is described. Clinical signs were limited to circling and right side paralysis. Histological examination of the brain revealed marked gliosis and discreet edema. The perivascular mononuclear inflammatory infiltrate was composed of few layers of lymphocytes, plasmocytes and macrophages and rare eosinophils. The presence of rhabditiform nematodes was associated with the infiltrate. Areas of malacia associated with the parasites and parasite tracks with axonal spheroids were also seen close to the vessels and to the etiological agent and were more evident in the white matter. In the meninges there was moderate inflammatory infiltrate associated with perivascular parasites. The identification of the nematode was based on the histological examination of the cerebral fragments.

Um eqüino macho, com 10 anos, Mangalarga, apresentou uma infecção por um nematódeo rabditiforme no cérebro. Os sinais clínicos limitaram-se ao fato de o animal andar em círculos e apresentar paralisia do lado direito. O exame histológico do cérebro revelou acentuada gliose e discreto edema intersticial. O infiltrado inflamatório mononuclear perivascular era composto por poucas camadas de linfócitos, plasmócitos, macrófagos e raros eosinófilos, associados aos nematódeos rabditiformes. Areas de malácia e trajetos com esferóides axonais são vistos ao redor de vasos e do agente etiológico, sendo mais evidentes na substância branca. Nas meninges, o infiltrado inflamatório foi moderado e associado a parasitas perivasculares. A identificação do nematódeo foi baseada no exame histológico do cérebro do cavalo.