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snai-1 and Epithelial-Mesenchymal Transition-Related Protein Immunoexpression in Canine Mammary Carcinomas

Advances in Breast Cancer Research 2014年第4期3卷 111-117页

作者： Breno s.salgado Rafael M.rocha Fernando A.soares Fátima Gartner noeme s.rocha Departamento de Patologia Faculdade de Medicina de BotucatuUniversidade Estadual Paulista(UNESP)BotucatuBrazil Laboratório de Patologia Investigativa e Comparada Departamento de Clínica VeterináriaFaculdade de Medicina Veterinária e ZootecniaUniversidade Estadual Paulista(UNESP)BotucatuBrazil Departamento de Anatomia Patológica Hospital A.C.CamargoFundacao Antonio PrudenteSao PauloBrazil Institute of Pathology and Molecular Immunology(IPATIMUP) OportoPortugal

Epithelial-mesenchymal transition (EMT) is defined as switching of polarized epithelial cells to a migratory fibroblastoid phenotype. EMT is known to be involved in the progression and metastasis of various cancers in humans, but this specific process is still little explored in the veterinary literature. The aim of this research was to evaluate the expression of EMT-related proteins in canine mammary carcinomas (CMCs). The expression of six EMT-related proteins in 94 CMCs of female dogs was evaluated by immunohistochemistry using a tissue array method. Additionally, clinicopathological characteristics were compared with the expression of EMT-related proteins. Loss of epithelial protein and/or acquisition of the expression of mesenchymal proteins were observed in CMCs. Loss of epithelial protein and/or acquisition of the expression of mesenchymal proteins were observed, particularly in tumors with evidence of stromal invasion;however, significance was only observed between the s100A4 and vascular invasion. In addition, snai-1 nuclear immunoexpression was significantly related to E-cadherin loss. In conclusion, loss of epithelial proteins and/or the acquisition of mesenchymal proteins are associated with EMT and may have an important role in the evaluation of CMC patients. The unique immunoexpression pattern of snai-1 could help to distinguish between an adenoma and a non-metastatic carcinoma and seems to be related to conversion of myoepithelial cells to a complete mesenchymal-like phenotype. Loss of E-cadherin and cytokeratin and change of immunoexpression pattern of snai-1, N-cadherin, s100A4 and MMP-2 indicate the occurrence of EMT in canine mammary carcinomas and should result in an en bloc resection or a close follow-up.

关键词： EMT s100A4 Keratin Mammary Tumors Dog

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Tetramethrin Identified in Fat Adjacent to Mammary Carcinoma in a Male Dog—Case Report

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Open Journal of Pathology 2012年第3期2卷 102-105页

作者： Fernanda C.Figueiroa Paulo Ricardo O.Bersano Fabio Henrique E.Andrade noeme s.rocha school of Veterinary Medicine and Animal science Sao Paulo State University(UNESP)BotucatuBrazil Maranhao state University(UEMA) Sao LuisBrazil

Canine mammary tumors (CMT) are very common in female dogs, representing approximately 50% of all tumors in this species, but are highly uncommon in male dogs and are primarily related to hyperestrogenism. There is considerable scientific interest in the possible role of environmental contaminants in the etiology of mammary tumors, specifically in relation to synthetic chemical substances released into the environment to which living beings are either directly or indirectly exposed. In this study, the presence of pyrethroid insecticide was observed in adipose tissue adjacent to canine mammary tumors. High Precision Liquid Chromatography—HPLC was adapted to detect and identify environmental contaminants in adipose tissue adjacent to malignant mammary tumor in one male dog, Akita, 12 years old. After surgery, the mass was carefully examined for malignant neoplastic lesions. Five grams of adipose tissue adjacent to the tumor was collected to detect environmental contaminants. The pyrethroid was identified as tetramethrin, at 0.20 μg/g. This is the first report in which the environmental contaminant level was detectable in adipose tissue of this male dog with a malignant mammary tumor, by HPLC. Results suggest the possible involvement of pyrethroid in the canine mammary tumor carcinogenesis since this animal did not present any other tumor that could cause hyperestrogenism.

关键词： Tetramethrin Mammary Tumor Male Dog Environmental Contaminants

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Phenotypic Characterization Murine sarcoma TG-180 Immunophenotypical Characterization Murine sarcoma TG-180

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Open Journal of Pathology 2015年第2期5卷 59-64页

作者： Thais M.T.Bernardes Hugo E.O.Beserra stephane C.O.R.Vexenat Helio Langoni noeme s.rocha Department of Clinical Veterinary Medicine Laboratory of Investigative and Comparative PathologySchool of Veterinary Medicine and Animal ScienceSao Paulo State UniversityBotucatuBrazil Department of Veterinary Hygiene and Public Health School of Veterinary Medicine and Animal ScienceSao Paulo State UniversityBotucatuBrazil

The sarcoma is the generic nomenclature for neoplasm of mesodermal cells, which express in man and animals. silent growth requires early diagnosis technique for identifying their proteins. The experimental model in vivo murine sarcoma 180-TG (TG-180), is widely used in research to provide the stimuli of infectious and neoplastic antigens. In this case, the technique of immunohisto-chemistry helps identify the expressions of tumor cell variants. The objective of the research was to characterize immunoexpression murine sarcoma TG 180, by immunohistochemistry, antibodies AE1/AE3, vimentin, CD3, CD 45, CD79α and s100A4. For this, murine sarcoma TG-180, was implanted subcutaneously in 20 mice “swiss”, male, 30 days old, 28 g for 10 days. samples were taken and subjected to immunohistochemistry, via use of HistoMouse-MA™? kit. There was specifically labeled s100A4 and vimentin antibodies, indicative of poorly differentiated neoplasms fibroblasts. In fact, the model is established by identifying the origin of the cell, once identified, chemotherapeutic tests can also be performed. Neoplasia like these, when installed in man and animals, depending on the degree of aggressiveness requires treatment protocol varied between surgery and chemotherapy or combination of treatments.

关键词： Immunohistochemistry Neoplasia Mice Tumor Origin

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Immunoexpression of Cathepsin D and s100A4 Protein and Their Molecular subtyptes in Canine Mammary Carcinomas

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Open Journal of Veterinary Medicine 2012年第4期2卷 163-169页

作者： Fernanda C.Figueiroa Breno s.salgado Lidianne N.Monteiro Rafael Malagoli rocha Maria Aparecida C.Domingues Diana Martins Fernando schmitt noeme s.rocha Departamento de Patologia Faculdade de Medicina de BotucatuUniversidade Estadual Paulista“Julio de Mesquita Filho”BotucatuBrazil Departamento de Anatomia Patologica Hospital A.C.CamargoFundacao Antonio PrudenteSao PauloBrazil Instituto de Patologia e Imunologia Molecular da Universidade do Porto OportoPortugal Faculdade de Medicina Universidade do PortoOportoPortugal Departamento de Clínica Veterinaria Servico de Patologia VeterinariaFaculdade de Medicina Veterinaria e ZootecniaUniversidade Estadual Paulista“Julio de Mesquita Filho”BotucatuBrazil

Cathepsin D (CD), a lysosomal protease, and s100A4 protein, a calcium binding motif, are considered to be involved in metastasis in various human cancers. No data regarding such proteins are available for canine mammary carcinomas (CMCs). Accordingly, their expression in association with known factors of prognosis was investigated in this study. For that, 66 surgically resected CMCs were submitted to an immunohistochemical evaluation using anti CD, s100A4 protein, HER2, estrogen receptor α, cytokeratin 5, and p63 antibodies, further characterizing the tumors' molecular subtype. An increase in s100A4 immunoexpression by neoplastic luminal mammary cells was associated with an infiltrative tumor mode of growth, consequently leading us to conclude that s100A4 protein could be related to progression in CMCs. Additionally, the occurrence of the luminal A molecular subtype was associated with the complex histotype in CMCs. Although we have demonstrated that changes in s100A4 protein immunoexpression occurs in CMCs, further studies are needed to determine whether this represents important independent biomarkers for CMCs.

关键词： Cathepsin Mammary Tumors Metastasis-Associated Proteins Molecular subtypes s100 Protein

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Expression of Multidrug Resistance ATP-Binding Cassette(ABC)Transporters in Canine Mammary Tumors

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Advances in Breast Cancer Research 2015年第3期4卷 77-85页

作者： Breno s.salgado suely Nonogaki Luisa M.soares Angela Akamatsu Cristiano R.N.da silva Thiago P.Anacleto Rodolfo Malagó Rafael M.rocha Fátima Gartner noeme s.rocha Departamento de Patologia Faculdade de Medicina de BotucatuUniversidade Estadual PaulistaCampus de BotucatuBotucatuBrazil Instituto de Ciências Biomédicas Abel salazar Universidade do Porto(Intitute of Biomedical Sciences Abel Salazar of University of Porto)OportoPortugal Fundacao Antonio Prudente Hospital A.C.CamargoSao PauloBrazil Departamento de Clínica Veterinária Faculdade de Medicina Veterinária e ZootecniaUniversidade Estadual PaulistaUNESP.BotucatuBrazil Hospital Escola de Medicina Veterinária Fundacao de Ensino e Pesquisa de Itajubá—FEPIItajubáBrazil

Mammary neoplasms are the most common tumors in female dogs. They are usually treated using solely surgical mastectomy—which is recognized as unsatisfactory in many cases. Given this, the benefits of chemotherapy in dogs with mammary cancer need to be further explored. some drugs that can be used for treating canines with mammary tumors may be substrates of uptake and/or efflux transporters such as the ATP-binding cassette (ABC) transporters. Unfortunately, very little is known regarding the pathobiology of such proteins in canine tumors, including mammary cancer. Accordingly, this study was designed to characterize the expression of ABC transporters P-glycoprotein, MRP1, and MRP2 and their relation with clinicopathologic factors in order to allow a better understanding of their influence in canine mammary cancer. P-glycoprotein was expressed in tumors from 55.8% of patients, while MRP1 and MRP2 were expressed in 37.2% and 39.5% of tumors, respectively. P-glycoprotein expression showed to be related with regional lymph node spread (P = 0.0038), as well as with tumor grade (P = 0.0353) and with a shorter survival (P = 0.0245). MRP1 revealed a strong association with a higher histological grade (P survival (P = 0.0002). Additionally, MRP1 was determined as prognostic indicator independent of lymph node status using Cox proportional-hazards regression multivariate analysis (P = 0.0216). No relations between MRP2 and clinicopathologic features were observed. We have found that P-glycoprotein and MRP1 are expressed in highly aggressive canine mammary tumors and are related with poor prognosis. Our results suggest that they may play a significant role in the course of canine mammary cancer progression and be promising candidate markers for a validation study on therapy outcome.

关键词： MDR1 P-Gp MRP1 MRP2 ABCC2 Mammary Neoplasms Prognosis Dog

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Role of selective Cyclo-Oxygenase-2 Inhibitor Celecoxib in Canine Osteosarcoma Cell Culture

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Open Journal of Pathology 2013年第4期3卷 144-149页

作者： Paulo R.O.Bersano Maria T.s.Alves Maria F.M.R.Gartner Adriana Camargo Ferrasi Joao F.Lima-Neto Fernanda C.Landim stephane C.Vexenat Carlos Eduardo Fonseca Alves Glenda Nicioli da silva noeme s.rocha Laboratory of Investigative and Comparative Pathology School of Veterinary Medicine and Animal ScienceUniversidade Estadual PaulistaBotucatuBrazil Department of Pathology Paulista School of MedicineFederal University of Sao PauloSao PauloBrazil Institute of Molecular Pathology and Immunology of University of Porto PortoPortugal Hemocentro Paulista school of Medicine Federal University of Sao PauloSao PauloBrazil Laboratory of Reproduction and Cellular Therapy Department of Animal Reproduction and Veterinary RadiologyUniversidade Estadual PaulistaBotucatuBrazil

Background: Experimental studies have shown that cyclo-oxygenase-2 (Cox2) is related to the development and progression of tumors, since this enzyme is induced and expressed by cells such as macrophages, osteoblasts, “activated” endothelial cells, and tumor cells. The activity in tumors includes proliferation, cell transformation, tumor growth, invasion and metastasis and may play an important role in carcinogenesis of the canine osteosarcoma, since it has high expression in tissue fragments. The combination of selective Cox2 inhibitors and other treatment modalities is the basis for a new anti-cancer therapy strategy. This in vitro study exposed primary cells of five different canine osteosarcoma cultures to selective Cox2 inhibitor at increasing concentrations and times. Results: For Cox2 negative cultures, despite the absence of differences, greater sensitivity of cells to treatment was observed. For Cox2 positive cultures, a higher number of necrotic cells were observed (P ≤ 0.05), when compared with negative cultures. For exposure times with Celecoxib doses, no difference (P > 0.05) was found between the three times analyzed for living, apoptotic and apoptotic/necrotic cells. There are similarities in the values of 24 h and 48 h, with slight reduction of living cells, increasing those undergoing apoptosis and apoptosis/necrosis. There was significance for necrosis (P ≤ 0.05). In 72 hours, a significant difference was observed between the other two previous values (P ≤ 0.05). It was found for the group of 100 μML-1, that there was a numerically greater signaling for apoptosis and lower (P = 0.08) for necrosis, and this point was the onset of the pharmacodynamic phenomenon, with drop in the values for living cells and increased number of necrotic cells, with a tendency (P = 0.08) for reducing the percentage of necrotic cells for the group of 100 μML-1 when compared to that of 10 μML-1. Conclusions: For Cox2 positive and negative cultures, there was difference for

关键词： Canine Osteosarcoma Cyclo-Oxygenase-2 Celecoxib

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Fine Needle Aspiration Cytology in the Diagnosis of Canine Cutaneous Transmissible Venereal Tumor—Case Report

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Open Journal of Veterinary Medicine 2014年第9期4卷 204-209页

作者： noeme sousa rocha Talia Missen Tremori Joao Alexandre Matos Carneiro Departament of Veterinary Clinic Faculty of Veterinary Medicine and Animal Science-UNESPBotucatuBrazil Department of Animal Reproduction and Veterinary Radiology Faculty of Veterinary Medicine and Animal Science-UNESPBotucatuBrazil

Fine needle aspiration cytology (FNAC) has been widely used in the diagnosis of lesions from various origins, especially neoplastic. The technique is simple, fast, safe, minimally invasive and inexpensive, which allows through the evaluation of cell morphology to establish prognosis, delineate surgical margins, monitor lesion growth, validate indication euthanasia during surgery and monitor chemotherapy protocols. Diagnosis of canine transmissible venereal tumor (TVT) can be accomplished with ease and precision, even to be rated, according to the degree of aggressiveness. The study objective was to demonstrate the effectiveness of the examination in the diagnosis of TVT plasmacytoid type. An eight-month dog presented to the veterinary hospital (HV), faculty of veterinary medicine and animal science, FMVZ, UNEsP-Univ Estadual Paulista with clinical suspicion of cutaneous lymphoma. By presenting multiple nodular lesions, FNAC was performed to cytological diagnosis. The tissue showed cells consistent with TVT. The animal was treated, and a total cure was achieved. According to the literature, TVT mainly affects external genitalia of sexually active animals and its transmission is more frequent during intercourse. In addition, animals sexually immature and without contact to the street dogs, hardly have injuries by TVT. In this case, verrucous and ulcerated lesions on the vulva of its mother during pregnancy and childbirth infected the animal. Diffuse and predominant dorsal injuries occurred due to both exfoliation of breast tumor during delivery and immunosuppression of pup at birth, thus favoring an atypical transmission.

关键词： Cytological Diagnosis TVT Atypical Transmission

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Analisys of Immunohistochemical Prognostic Markers in Canine Mammary Cancer and Its Relation to Postsurgical survival

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Open Journal of Pathology 2015年第3期5卷 90-99页

作者： Francisco Pedraza-Ordonez Luis Mauricio Montoya-Florez Camilo Bulla noeme sousa rocha Department of Veterinary Clinics Laboratory of Investigative and Comparative PathologySchool of Veterinary Medicine and Animal ScienceUniversidade Estadual Paulista FMVZ-UNESPSao PauloBrazil Animal Pathology Research Group Animal Health DepartmentUniversidad de CaldasManizalesColombia Department of Pathology/Population Medicine-Mississippi College of Veterinary Medicine Mississippi State UniversityStarkvilleUSA Department of Pathology Botucatu Medical SchoolUniversidade Estadual Paulista–UNESPDistrito de Rubiao Junior s/nBotucatuBrazil

Background: several animal models, including dogs, have been useful to compare the pathogenesis of mammary neoplasm in humans, showing biological parallelism in the growth and development of breast cancer. The causes of cancer could be attributed to change in several tumor suppressor genes. The relationship between molecule associated to senescence and clinical prognosis of patients affected by mammary cancer is little known. Beyond a collection of data, the major interest of the present study was to carry out a clinical follow-up of patients affected by these tumors, through association with new molecular markers by immunohistochemical technic. Upon completing the study, 15 patients survived, while 45 died. In the case of malignant neoplasms, 40 patients died because of the illness. The type of surgery most used by veterinarian surgeons was the simple lumpectomy, followed by the regional mastectomy. sentinel node was removed by surgery only when clearly affected. Result: Markings against steroid hormones were positive. Regarding the markings against HER2 and Ki-67, they were negative in all cases. The markings against P53 and CD31 were all positives. Markings against molecules associated with cellular senescence were all positives. No statistical differences were found in immunomarcation for the different antigens used as clinical prognosis factors in mammary neoplasms. Conclusions: According to the study conditions, the survival of patients affected by breast tumors is directly related to diagnosis and malignancy histological grade, but not to animal breed, number of affected glands or patient reproductive status. On the other hand, immunohistochemical markings were not related to the patient prognosis. For this reason, it is important to highlight the persistance of a high percentage of mammary neoplasm cases clinically diagnosed with poor results on patient survival. Thus, educating owners and veterinarians for using diagnostic available tools to improve the pro

关键词： Celular senescence Prognostic Markers Immunohistochemical Mammary Cancer Animal Model

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Quantitative Analysis of Photodynamic Therapy Effects in Rat Mammary Tumor Vascular Density Using Image-Pro plus software

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Open Journal of Veterinary Medicine 2013年第5期3卷 259-262页

作者： Isabelle Ferreira Camilo Bulla Wes Baumgartner Vanderlei salvador Bagnato noeme sousa rocha Veterinary Clinics Department School of Veterinary Medicine and Animal Science Universidade Estadual Paulista Botucatu Brazil

Photodynamic therapy (PDT) is a treatment modality that has advanced rapidly in recent years. It causes tissue and vascular damage with the interaction of a photosensitizing agent (Ps), light of a proper wavelength, and molecular oxygen. Evaluation of vessel damage usually relies on histopathology evaluation. Results are often qualitative or at best semi-quantitative based on a subjective system. The aim of this study was to evaluate, using CD31 immunohistochemistry and image analysis software, the vascular damage after PDT in a well-established rodent model of chemically induced mammary tumor. Fourteen sprague-Dawley rats received a single dose of 7,12-dimethylbenz(a)anthraxcene (80 mg/kg by gavage), treatment efficacy was evaluated by comparing the vascular density of tumors after treatment with Photogem®?as a Ps, intraperitoneally, followed by interstitial fiber optic lighting, from a diode laser, at 200 mW/cm and light dose of 100 J/cm directed against his tumor (7 animals), with a control group (6 animals, no PDT). The animals were euthanized 30 hours after the lighting and mammary tumors were removed and samples from each lesion were formalin-fixed. Immunostained blood vessels were quantified by Image Pro-Plus version 7.0. The control group had an average of 3368.6 ± 4027.1 pixels per picture and the treated group had an average of 779 ± 1242.6 pixels per area (P 0.01), indicating that PDT caused a significant decrease in vascular density of mammary tumors. The combined immunohistochemistry using CD31, with selection of representative areas by a trained pathology, followed by quantification of staining using Image Pro-Plus version 7.0 system was a practical and robust methodology for vessel damage evaluation, which probably could be used to assess other antiangiogenic treatments.

关键词： Mammary Tumors Photodynamic Therapy Vascular Damage

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ROsE: Alternative for Cancelled and Inconclusive Cytopathologic Examinations, as Well as Professional Training at the UNEsP-Botucatu Veterinary Hospital

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Open Journal of Veterinary Medicine 2019年第9期9卷 121-135页

作者： Fabiane Andrade Correia Neiva Eduardo Eburnio Paula de sanctis Nayara Maria Gil Mazzante noeme sousa rocha Investigative and Comparative Pathology Laboratory S&Atilde o Paulo state University (UNEsP) Botucatu Brazil school of Medicine from Botucatu S&Atilde o Paulo state University (UNEsP) Botucatu Brazil

ROsE is a method for fast sample evaluation that does not compromise sensitivity and specificity in the hospital environment when establishing the diagnosis and the therapeutic protocol. The technique is already well-established in human medicine as it operates within the standard operational procedure, reducing the rates of inconclusive results and cancelled cytopathology assays in the hospital environment. However, its application is scarce in Veterinary Medicine, including in hospitals where intensive care is available. This study aims at conducting a case-by-case assessment of inconclusive and cancelled examinations in the Cytopathology Outpatient Clinic at UNEsP (HV), Botucatu, Brazil, from 2012 to 2016 and ascertains the causes. For this purpose, a retrospective study was conducted for 9587 examinations in canines, of which 4.1% and 10.44% were inconclusive and cancelled, respectively. These results are not in line with ROsE, which foresees a total rate of 5% for these occurrences. The reasons for these high rates in the outpatient clinic were the lack of a standard Operating Procedure (sOP) and the relative inexperience of the residents in a university hospital. Therefore, with the adoption of the ROsE protocol for veterinary cytopathological examinations, together with adequate training for the outpatient professionals responsible for the examinations, the goals set forth by the ROsE protocol would be met and the rates of inconclusive and cancelled examinations would decrease considerably. Lastly, this study hopes to contribute towards the diagnosis and therapeutic protocols of the main diseases affecting dogs.

关键词： PAAF Inconclusive or Canceled Cytology Canine ROsE Rapid On-site Evaluation

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