Angélica Cavalheiro Bertagnolli

Cox-2 Expression in Canine Mammary Carcinomas: Correlation with Angiogenesis and Overall Survival

Mammary tumors are among the most common neoplastic processes in female dogs. Prostaglandin E2, the catalytic product of Cox-2, may promote tumor development and angiogenesis. It has been investigated in several human cancers and also correlated with the evolution of the disease. However, the clinical implications of tumor pathology require more investigation in veterinary medicine. Angiogenesis is essential for the growth and metastasis of major solid tumors and has been correlated with prognosis in human and canine breast cancer. The aim of this study was to evaluate Cox-2 expression and microvessel density in canine mammary carcinomas and to correlate them with overall survival of the animal. Cox-2 and angiogenesis were assessed by immunohistochemistry in 46 mammary carcinomas (19 ductal and 27 metaplastic) and in healthy mammary glands. To assess tumor angiogenesis, microvessel density (MVD) was determined by CD31 staining. Immunostaining revealed that 46/46 (100%) of the tumors were positive for Cox-2 and CD31, and there was no statistical difference among tumor types. Cox-2 protein expression correlated positively with CD31 staining (r = 0.3742, P = .0104) but did not correlate significantly with tumor type. Longer overall survival was observed in metaplastic carcinomas (P = .028), in tumors with low microvessel density (P = .0002) and with low Cox-2 score (P = .01). Our results demonstrate that increased microvessel density and increased Cox-2 expression were linearly related in the canine mammary tumors studied and were also related to worse prognosis and shorter overall survival. This suggests that Cox-2 inhibitors could be an alternative for the treatment and control of advanced neoplastic mammary disease in female dogs.

The relationship between tumour size and expression of prognostic markers in benign and malignant canine mammary tumours

Tumour size is considered one of the most important determinants of clinical staging in cancer patients. The aim of this study was to assess the value of tumour size as an indicator of the differentiation of mammary neoplasias in female dogs. The tumour, nodes metastates (TNM) system, based on primary lesion size, the extent of its dissemination to regional lymph nodes and the presence or absence of distant metastases, was applied to 120 female dogs diagnosed with mammary neoplasias. Paraffin blocks from 38 cases were selected and studied by immunohistochemical staining for prognostic and predictive markers of breast cancer. The Kaplan-Meier survival curve was estimated for 110 female dogs. Larger tumours (T3) were mostly malignant and showed lower expression of progesterone receptor and higher expression of cellular proliferation markers. Global survival time was shorter in female dogs with large tumour masses. This study highlights the importance of tumour size as a prognostic indicator of mammary neoplasias in female dogs.

Effects of dantrolene on apoptosis and immunohistochemical expression of NeuN in the spinal cord after traumatic injury in rats

Dantrolene has been shown to be neuroprotective by reducing neuronal apoptosis after brain injury in several animal models of neurological disorders. In this study, we investigated the effects of dantrolene on experimental spinal cord injury (SCI). Forty‐six male Wistar rats were laminectomized at T13 and divided in six groups: GI (n = 7) underwent SCI with placebo and was euthanized after 32 h; GII (n = 7) underwent laminectomy alone with placebo and was euthanized after 32 h; GIII (n = 8) underwent SCI with dantrolene and was euthanized after 32 h; GIV (n = 8) underwent SCI with placebo and was euthanized after 8 days; GV (n = 8) underwent laminectomy alone with placebo and was euthanized after 8 days; and GVI (n = 8) underwent SCI with dantrolene and was euthanized after 8 days. A compressive trauma was performed to induce SCI. After euthanasia, the spinal cord was evaluated using light microscopy, TUNEL staining and immunochemistry with anti‐Caspase‐3 and anti‐NeuN. Animals treated with dantrolene showed a smaller number of TUNEL‐positive and caspase‐3‐positive cells and a larger number of NeuN‐positive neurons, both at 32 h and 8 days (P ≤ 0.05). These results showed that dantrolene protects spinal cord tissue after traumatic SCI by decreasing apoptotic cell death.

Canine Mammary Mixed Tumours: A Review

Mammary mixed tumours are the most frequent neoplasias in female dogs. In humans, mixed tumours are frequently found in the salivary glands and are known as pleomorphic adenomas. In addition to their histomorphologic similarities, mixed tumours and pleomorphic adenomas have the potential to become malignant and give rise to carcinomas in mixed tumours and carcinomas ex-pleomorphic adenoma, respectively. The factors associated with malignant transformation are still poorly known in the case of canine mixed tumours. However, this form of neoplasia tends to be associated with a better prognosis than other malignant histological types. This paper discusses the main features associated with female canine mammary mixed tumours.

Evaluation of adverse effects in tamoxifen exposed healthy female dogs

BackgroundMammary tumors are among the most frequent neoplasms in female dogs, but the strategies employed in animal treatment are limited. In human medicine, hormone manipulation is used in cancer therapy. Tamoxifen citrate is a selective inhibitor of oestrogen receptors and exerts a potent anti-oestrogen effect on the mammary gland. The aim of this study was to evaluate the adverse effects when exposing healthy female dogs to tamoxifen.MethodsTamoxifen was administered for 120 days at a dose of 0.5 or 0.8 mg/kg/day to either intact or spayed female dogs. The effects were assessed through clinical examination, haematology, serum biochemistry, ophthalmology and bone marrow aspirate examination. Ovariohysterectomy was performed and the uterus examined by histopathology.ResultsVulva oedema and purulent vaginal discharge developed with 10 days of tamoxifen exposure in all groups. Pyometra was diagnosed after around 90 days of exposure in intact females with frequencies increasing during the following 30 days of exposure. Up to 50% of dogs within the groups developed retinitis but none of the dogs had signs of reduced visual acuity. The prevalence of retinitis in each group was similar after 120 days of exposure. Haematological, biochemical and bone marrow changes were not observed. Due to the high risk of developing pyometra after prolonged exposure to tamoxifen, only spayed animals should be given this medication.ConclusionsA dose of 0.8 mg tamoxifen/kg body weight/day is recommended when treating tamoxifen-responsive canine mammary tumors. Due to the high risk of developing pyometra, ovariohysterectomy is recommended.

An assessment of the clonality of the components of canine mixed mammary tumours by mitochondrial DNA analysis

The aim of this study was to investigate if mutations in the mitochondrial DNA (mtDNA) D-loop fragment control region of canine mammary mixed tumours could be used as clonal markers that identified the cell population of origin. Ten benign mixed mammary tumours and nine carcinomas arising from benign mixed tumours were microdissected and DNA from epithelial and mesenchymal tumour cells and from normal mammary tissue was examined for sequence variations in a fragment of the hypervariable control region. Identical sequence variants in both the epithelial and mesenchymal components (as well as in the corresponding normal tissue) were found in 80% of the benign mixed tumours and in 89% of the carcinomas arising from benign mixed tumours suggesting a shared clonal origin. The distinctive sequence alterations identified in the epithelial and mesenchymal components of 15.8% of all 19 tumours examined, suggests the possibility that a minority of mammary tumours are polyclonal in origin or that early clonal divergence occurs. Increased mutation within the mtDNA D-loop fragment of mixed tumour components was not observed.

Morphological aspects and immunophenotypic profiles of mammary carcinomas in benign-mixed tumors of female dogs

Carcinoma in benign-mixed tumor (CBMT) is common in the female canine mammary gland and comprises malignant epithelial between benign mesenchymal elements. This study investigated the morphological aspects of 29 CBMT and their immunophenotypical profiles, by using an immunohistochemistry panel based on five molecular markers—estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2), cytokeratin 5 (CK5), and human epidermal growth factor receptor 1 (EGFR). From these, CBMT was classified into four subtypes: luminal A, luminal B, HER2-like, basal-like, and normal. “In situ” and invasive carcinomatous components were analyzed and compared. Histological grade I carcinoma was observed in 16 cases (55.2%) of the tumors analyzed, grade II in 10 cases (34.5%), and grade III in three cases (10.3%). The invasive carcinomatous component has shown, more frequently, luminal A (12/29 cases, 41.4%), followed by basal-like phenotype (8/29 cases, 27.6%). There was high concordance between immunophenotypical profiles of the in situ and invasive carcinomatous components (kappa coefficient = 0.816, P < 0.001). We concluded that CBMT predominantly has features of low-grade neoplasms of malignancy. The various immunophenotypic profiles suggest the origin of these lesions in more than one cell type (luminal and myoepithelial).

Versican expression in canine carcinomas in benign mixed tumours: is there an association with clinical pathological factors, invasion and overall survival?

BackgroundComponents of the extracellular matrix have been studied in an attempt to elucidate the mechanisms involved in the biological behaviour of tumours. The presence of the proteoglycan versican has been strongly associated with cancer development and progression. However, relationship between versican expression and clinical pathological factors and overall survival has not been previously studied in veterinary medicine. Carcinomas in benign mixed tumours (CBMTs) are one of the most common malignant tumours in female canines and can serve as models for studies of tumour progression. The aim of this study was to evaluate the expression of versican in in situ and invasive carcinomatous areas of canine CBMTs and to evaluate possible associations of versican expression with other classic prognostic factors and overall survival.ResultsClinical staging; histological grade determination; immunohistochemical staining for versican, E-cadherin and Ki-67; and confirmation of invasion areas by staining for p63 and smooth muscle α-actin (α-SMA) were performed on 49 canine cases of CBMT. Tumour invasion was considered when suspicious Haematoxylin-Eosin (HE)-stained areas showed a total loss of α-SMA and p63 immunoreactivity. Versican immunoreactivity was less intense in the areas adjacent to the in situ carcinomatous regions, compared to invasive regions, which showed extensive and strong staining.ConclusionsOur data reveal that in canine CBMTs, versican expression differs significantly between invasive and in situ areas, suggesting a role for this molecule in tumour progression. Although a direct relationship exists between versican and invasiveness, our results indicate that the isolated evaluation of this proteoglycan does not represent an independent prognostic factor in canine CBMTs.

Mast cells and angiogenesis in canine mammary tumor

Estimou-se a correlacao entre a densidade de microvasos e a densidade de mastocitos em tumores mamarios caninos. Sessenta e cinco amostras de tumores mamarios caninos - 24 benignos e 41 malignos - foram analisadas, pela tecnica rotineira de coloracao com Azul de Toluidina para avaliacao da densidade de mastocitos. Para a avaliacao da angiogenese, foi utilizada a tecnica de imunoistoquimica para expressao de CD31. Nao foram observadas diferencas significativas de mastocitos (P=0.44) ou densidade microvascular (P=0.77) entre tumores malignos e benignos. A correlacao entre densidade microvascular e densidade de mastocitos foi positiva (r=0,39; P=0,011) em tumores malignos. Estes resultados sugerem que os mastocitos podem exercer um importante papel no desenvolvimento de tumores mamarios malignos caninos mediante promocao da angiogenese, similarmente a alguns tumores descritos na especie humana

A simple ductal mammary papilloma in a male maned wolf (Chrysocyon brachyurus)

A 1-cm-diameter nodule was identified in the left inguinal mammary gland of a 9-year-old male maned wolf (Chrysocyon brachyurus). The mass was surgically excised and examined histologically. Microscopically, the neoplasm consisted of papillary proliferations of epithelial cells on well-defined fibrovascular stalks. A myoepithelial layer was located between the single layer of epithelial cells and the fibrovascular stalk. This histologic appearance was compatible with a diagnosis of simple ductal mammary papilloma. Immunohistochemical staining was positive for p63, cytokeratins AE1/AE3, and estrogen receptors. The clinical and histologic observations in the present case indicate that male maned wolves may develop mammary tumors that are similar to those observed in domestic dogs and humans.