Canine and feline in situ mammary carcinoma: A comparative review.

Carcinoma in situ of the breast is a well-known entity in humans. In veterinary medicine, particularly in canine and feline mammary literature, there is no agreement whether the term in situ should be used to indicate a specific carcinoma histotype or the noninvasive status of a carcinoma of any histotype. Moreover, in the most recent histologic classification of mammary tumors published by the Davis-Thompson Foundation, it is suggested to abandon the term carcinoma in situ given the lack of standardized criteria defining this entity, replacing it with epitheliosis or ductal/lobular hyperplasia with severe atypia. This publication presents a critical review of the term in situ in human and veterinary medicine considering the evolution of the term over the years and its heterogeneous use by different authors, including variations in immunohistochemical markers for classification. This review aims to point out the lack of uniformity in the nomenclature and classification issues in veterinary medicine regarding the use of the term in situ, laying the ground for a process of standardization in future publications.

Canine Mammary Carcinoma With Vacuolated Cytoplasm: Glycogen-Rich Carcinoma, a Histological Type Distinct From Lipid-Rich Carcinoma.

Lipid-rich carcinoma is a rare histotype of canine mammary tumors with cytoplasmic vacuolation. In humans, glycogen-rich carcinoma, secretory carcinoma, and myoepithelial neoplasms are included in the differential diagnosis for lipid-rich carcinoma. The aim of the study was to investigate the existence of histotypes other than lipid-rich in canine mammary carcinomas with vacuolated cytoplasm using a diagnostic algorithm based on histopathology, histochemistry, immunohistochemistry, and ultrastructure and to evaluate the molecular phenotype of these neoplasms. Ten mammary carcinomas were collected, histologically reviewed, and subjected to histochemistry (PAS, PAS with diastase, Alcian blue, Sudan III [1 case], and Congo red [1 case]); immunohistochemistry for CK19, CK5/6, CK14, p63, calponin, vimentin, ER, PR, and HER2; and transmission electron microscopy (TEM). Cytokeratin immunolabeling demonstrated the epithelial origin of all tumors. Sudan III and TEM confirmed the diagnosis of lipid-rich carcinoma in 8 tumors (one amyloid-producing). One tumor was reclassified as a glycogen-rich carcinoma based on PAS reactivity that was diastase-labile, and a second tumor was reclassified as a carcinoma-and-malignant myoepithelioma based on the differentiation markers. Lipid-rich carcinomas were basal-like (5/8), null-type (2/8), and luminal A phenotype (1/8). The glycogen-rich carcinoma was basal-like, while the carcinoma-and-malignant myoepithelioma was luminal A. Vacuolated morphology of neoplastic cells in canine mammary carcinoma can indicate either a neoplasm of luminal epithelial origin with cytoplasmic lipid or glycogen, or vacuolated neoplastic suprabasal myoepithelial cells. Glycogen-rich carcinoma is a novel histological type that should be considered in the differential diagnosis for canine mammary carcinomas with vacuolated cytoplasm.

Effects of Electrochemotherapy with Cisplatin and Peritumoral IL-12 Gene Electrotransfer on Canine Mast Cell Tumors: a Histopathologic and Immunohistochemical Study.

BACKGROUND: The study was aimed to characterize tumor response after combined treatment employing electrochemotherapy with IL-12 gene electrotransfer in dogs with spontaneous mast cell tumors (MCT). MATERIALS AND METHODS: Eleven dogs with eleven MCTs were included in the study. Histological changes were investigated in biopsy specimens collected before the treatment (T0), and 4 (T1) and 8 weeks (T2) later. Cellular infiltrates were characterized immunohistochemically by using anti CD3, CD20, Foxp3 (Treg), CD68 and anti MHC-class II antibodies. Proliferation and anti-apoptotic activity of neoplastic cells were assessed using anti Ki-67 and Bcl-2 antibodies. Angiogenetic processes were investigated immunohistochemically by using anti Factor VIII and anti CD31 antibodies and micro vessel density quantification. RESULTS: Histopathological examination of samples at T0 confirmed the diagnosis and the presence of scanty infiltrates consisted mainly of T-lymphocytes and macrophages. At T1 and T2 neoplastic cells were drastically reduced in 7/11 cases, small clusters of neoplastic cells were detected in 3/11 cases and 1/11 cases neoplastic cells were still evident. Proliferation activity of neoplastic cells was significantly reduced at T1 and T2 and expression of anti-apoptotic protein at T1. Microvessel density was drastically reduced in all samples after treatment. The number of T-lymphocytes increased at T1, although not significant, while Treg were significant higher at T1 and macrophages at T2. CONCLUSIONS: The combined electrochemotherapy and IL-12 gene electrotransfer effectively induced a cellular response against neoplastic cells characterized mainly by the recruitment of T-lymphocytes and macrophages and a fibrotic proliferation with reduction of microvessels.

A Preliminary Evaluation of the Prognostic Role of HER-2 and HER-3 Immunohistochemical Expression in Canine Melanomas.

Canine melanoma is a malignant and aggressive neoplasm showing clinical, histological, and molecular features similar to the human counterpart. In human medicine, epidermal growth factor receptors (EGFRs) have already been suggested as prognostic markers and potential therapeutic targets in cutaneous melanoma. The aim of this study was to evaluate the expression of HER-2 and HER-3 in canine melanomas by immunohistochemistry and correlate their expression to the clinicopathological parameters of the examined tumors. Thirty-seven canine melanoma samples were recruited. Data regarding signalment and clinical parameters were also collected. The population was composed of 18 cutaneous, 16 oral/mucosal, and three digital/foot pad melanomas. Histopathological investigations were carried out to analyze histological type, ulceration, and mitotic count. On each sample, immunohistochemistry was performed using an anti-Melan-A or anti-Melanoma antigen, i.e., anti-HER-2 and anti-HER-3 antibodies. HER-2 and HER-3 positivity were classified using already established scoring criteria and a statistical analysis was carried out. The results highlighted that HER-2 expression was observed in 48.6% of the samples and HER-3 expression in 18.9%. The highest HER 2 score (3+) was recorded in 16.2% of the samples, while the coexpression of the two receptors was detected in 13.5% of the samples. A statistically significant association (p < 0.05) was observed between the expression of HER-2 and HER-3 and the presence of ulceration in oromucosal tumors. This work confirms the expression of HER-2 and HER-3 in canine melanomas and suggests a putative association with negative prognostic parameters. Further studies are necessary to strengthen these data by increasing the samples size and combining pathological examinations with molecular biology in the investigation of EGFR family receptors.

Age at Tumor Diagnosis in 14,636 Canine Cases from the Pathology-Based UNIPI Animal Cancer Registry, Italy: One Size Doesn't Fit All.

Cancer is the most common cause of death in adult dogs. All dogs would benefit from early diagnosis, but there are no specific guidelines regarding the schedule of cancer screening in companion animals. The aim of this study was to retrospectively evaluate the age at diagnosis in Italian oncological canine patients. A total of 14,636 canine histologically confirmed neoplastic cases were coded according to the Vet-ICD-O-canine-1 and stratified by malignancy, sex, neutering status, breed, cephalic index, body size, and tumor type. Differences in age distribution were analyzed and the influence of these variables on the time of first malignancy diagnosis was assessed using an event history analysis model. The median age at diagnosis for benign and malignant tumors was 9 and 10 years, respectively. Intact and purebred dogs were diagnosed earlier, but the median age differed significantly by breed. The earliest age at diagnosis was recorded for lymphomas and mast cell tumors. The model showed an accelerating effect of large size, brachy- and dolichocephaly, and sexual integrity in female dogs on the time of malignancy diagnosis. Our results confirm that a "one-size-fits-all" approach to cancer screening is not accurate in dogs and provide relevant data that may lead to the establishment of breed-based screening schedules.

Protein Expression, Amplification, and Mutation of HER2 Gene in Canine Primary Pulmonary Adenocarcinomas: Preliminary Results.

Recently, human epidermal growth factor receptor 2 (HER2) has emerged as a therapeutic target of interest for non-small-cell lung cancer in humans. The role of HER2 in canine pulmonary adenocarcinomas is poorly documented. To address this gap, this study employed three methodologies: immunohistochemistry (IHC), fluorescence in situ hybridization (FISH), and next-generation sequencing (NGS) to investigate the protein expression, gene amplification, and mutation of HER2 in 19 canine primary pulmonary adenocarcinomas. By IHC, 3 out of 19 cases were overexpressed 3+, 6 were 2+, and 10 were negative. With FISH, 2 cases were amplified (12.5%), 3 were inadequate for the analyses, and the others were non-amplified. With NGS, seven cases were inadequate. All other cases were wild-type, except for one IHC 3+ case, which was amplified with FISH and with a specific mutation already described in human pulmonary adenocarcinoma, V659E. This mutation is probably sensitive to tyrosine kinase inhibitory drugs. These results are similar to those in human medicine and to the few data in the literature on canine lung carcinomas; the presence of 12.5% of amplified cases in dogs lays the foundation for future targeted drugs against HER2 alterations.

Clinical outcome and cyclo-oxygenase-2 expression in five dogs with solar dermatitis/actinic keratosis treated with firocoxib.

BACKGROUND: The conversion of arachidonic acid into prostaglandin is catalysed by the cyclo-oxygenases (COX-1/COX-2). Several studies indicate that COX-2 is overexpressed in actinic keratosis in humans and dogs. Firocoxib is a COX-2-selective inhibitor that blocks the biochemical activity of COX-2. HYPOTHESIS/OBJECTIVES: To evaluate the efficacy of firocoxib (5 mg/kg orally once daily) for the treatment of dogs with solar dermatitis/actinic keratosis. METHODS: Firocoxib 5 mg/kg was given orally once daily for 180 days to five dogs with clinical signs and histopathological lesions consistent with solar dermatitis/actinic keratosis. On days 0, 50 and 180, the severity of erythema, skin shine, induration and the number of comedones were evaluated by a clinical scoring system. On the same days, samples were collected for histopathology from 'target lesions' and COX-2 expression was evaluated by immunohistochemistry. RESULTS: The clinical follow-up showed that four of five dogs improved with the treatment; improvement in terms of histological findings was correlated with the regularization of the epidermal proliferation rather than the recovery of dermal changes. CONCLUSIONS AND CLINICAL IMPORTANCE: A role for COX-2 might thus be hypothesized in the pathogenesis of canine solar dermatitis.

Micronodular ultrasound lesions in the colonic submucosa of 42 dogs and 14 cats.

Micronodular ultrasound lesions have been detected in the colonic submucosa of dogs and cats at our hospital. The lesions had rounded/oval shapes, measured 1-3 mm in size, and exhibited a hypo/anechoic ultrasonographic pattern. To our knowledge, these lesions have not been previously reported in human or veterinary patients. The purpose of this retrospective study was to determine whether micronodular lesions were associated with other abdominal ultrasound abnormalities or clinical findings. Medical records of dogs and cats with sonographic reports describing micronodular lesions within the colonic submucosa were reviewed. Concurrent ultrasonographic abnormalities were recorded and compared with clinical sidgns and follow-up data. A total of 42 dogs and 14 cats met inclusion criteria. Concurrent sonographic abnormalities included the following: increased colon wall thickness (12.5%); small bowel wall thickening, altered layering, and/or hyperechoic mucosa (45%); abdominal effusion (29%); caudal mesenteric lymphadenopathy (46%); mesenteric lymphadenopathy (27%); and pericolic peritoneal fat reactivity (9%). Fifty of 56 animals presented with diarrhea. Twenty-seven cases had clinical signs of colitis and ultrasonographic lesions were limited to the colonic submucosa. In nine cases, follow-up examination at 6-8 weeks showed resolution of clinical and ultrasonographic signs. Ultrasonographic and clinical examinations in 17 patients at 12-18 months and in 20 patients at 18-30 months from initial diagnosis showed resolution of submucosal lesions and clinical signs of enteropathy. The authors propose that micronodular submucosal ultrasound lesions may represent reactive intraparietal lymphoid follicles and may be indicators of colonic inflammatory diseases in dogs and cats.

Confirmation of the Prognostic Value of Foxp3+ Cells in Canine Mammary Tumors.

Foxp3+ cell counts were evaluated by immunohistochemistry in 59 canine mammary tumors, 20 adenomas, and 39 carcinomas in three different compartments: intratumoral, within the adjacent stroma, and in the distant stroma. Foxp3+ lymphocyte counts were compared with histotype, grading, presence of lymphatic invasion, immunohistochemical expression of estrogen and progesterone receptors, expression of c-erbB-2, and the overall survival (OS). Our findings confirmed that Foxp3+ cells were significantly higher in canine mammary carcinomas compared to adenomas. A significantly higher number of Foxp3+ cells were detected in grade III carcinomas compared to grade II carcinomas, as well as in tumors with lymphatic invasion and loss of ER-expression. Finally, a high number of Foxp3+ cells was associated with poor prognosis. In conclusion, our findings highlighted the association of Foxp3+ lymphocytes with negative clinicopathological features and shorter overall survival (OS), thus confirming the role of Tregs as a negative prognostic marker in canine mammary carcinomas.

Exploring the link between viruses and cancer in companion animals: a comprehensive and comparative analysis.

Currently, it is estimated that 15% of human neoplasms globally are caused by infectious agents, with new evidence emerging continuously. Multiple agents have been implicated in various forms of neoplasia, with viruses as the most frequent. In recent years, investigation on viral mechanisms underlying tumoral transformation in cancer development and progression are in the spotlight, both in human and veterinary oncology. Oncogenic viruses in veterinary medicine are of primary importance not only as original pathogens of pets, but also in the view of pets as models of human malignancies. Hence, this work will provide an overview of the main oncogenic viruses of companion animals, with brief notes of comparative medicine.

Preliminary Assessment of Tumor-Associated Tissue Eosinophilia (TATE) in Canine Mast Cell Tumors: Prevalence and Prognostic Relevance and Its Association with Neoangiogenesis.

Mast cell tumor (MCT) is the most common malignant skin tumor in dogs. In order to gain more information on the prognostic markers in MCT, the role of the eosinophil granulocytes infiltrates was investigated and assessed by the evaluation of tumor-associated tissue eosinophilia (TATE) in 87 canine cutaneous MCTs. In human medicine, high TATE are often described in highly angiogenic tumors: we therefore assessed the vascular endothelial growth factor (VEGF) expression in neoplastic mast cells. TATE and VEGF expression were compared between themselves, with histological grading, immunohistochemical expression of KIT and Ki-67, and with the recurrence. We found a statistically significant correlation between TATE and Patnaik grading (p = 0.041), Kiupel grading (p = 0.022), immunohistochemical KIT expression (p = 0.015), and tumor recurrence (p = 0.000). No associations were observed with Ki-67 and VEGF expression. This is the first evaluation of TATE and its prognostic value in canine MCTs in veterinary oncology. This study suggest that this investigation could be an important source of information for this tumor and for other neoplasms.

Cross-species oncogenomics offers insight into human muscle-invasive bladder cancer.

BACKGROUND: In humans, muscle-invasive bladder cancer (MIBC) is highly aggressive and associated with a poor prognosis. With a high mutation load and large number of altered genes, strategies to delineate key driver events are necessary. Dogs and cats develop urothelial carcinoma (UC) with histological and clinical similarities to human MIBC. Cattle that graze on bracken fern also develop UC, associated with exposure to the carcinogen ptaquiloside. These species may represent relevant animal models of spontaneous and carcinogen-induced UC that can provide insight into human MIBC. RESULTS: Whole-exome sequencing of domestic canine (n = 87) and feline (n = 23) UC, and comparative analysis with human MIBC reveals a lower mutation rate in animal cases and the absence of APOBEC mutational signatures. A convergence of driver genes (ARID1A, KDM6A, TP53, FAT1, and NRAS) is discovered, along with common focally amplified and deleted genes involved in regulation of the cell cycle and chromatin remodelling. We identify mismatch repair deficiency in a subset of canine and feline UCs with biallelic inactivation of MSH2. Bovine UC (n = 8) is distinctly different; we identify novel mutational signatures which are recapitulated in vitro in human urinary bladder UC cells treated with bracken fern extracts or purified ptaquiloside. CONCLUSION: Canine and feline urinary bladder UC represent relevant models of MIBC in humans, and cross-species analysis can identify evolutionarily conserved driver genes. We characterize mutational signatures in bovine UC associated with bracken fern and ptaquiloside exposure, a human-linked cancer exposure. Our work demonstrates the relevance of cross-species comparative analysis in understanding both human and animal UC.

Molecular portrait-based correlation between primary canine mammary tumor and its lymph node metastasis: possible prognostic-predictive models and/or stronghold for specific treatments?

BACKGROUND: This study aimed to evaluate the relationship between the molecular phenotype of the primary mammary tumor and its related lymph node metastasis in the dog to develop prognostic-predictive models and targeted therapeutic options. RESULTS: Twenty mammary tumor samples and their lymph node metastases were selected and stained by immunohistochemistry with anti-estrogen receptor (ER), -progesterone receptor (PR), -human epidermal growth factor receptor 2 (c-erbB-2), -cytokeratin 5/6 (CK 5/6), -cytokeratin 14 (CK14), -cytokeratin 19 (CK 19) and -protein 63 (p63) antibodies. Four phenotypes (luminal A, luminal B, c-erbB2 overexpressing and basal-like) were diagnosed in primary tumors and five (luminal A, luminal B, c-erbB-2 overexpressing, basal-like and normal-like) in the lymph node metastases. Phenotypic concordance was found in 13 of the 20 cases (65%), and seven cases (35%) showed discordance with different lymph node phenotypic profile from the primary tumor. CONCLUSIONS: The phenotype of the primary tumor assumes a predictive-therapeutic role only in concordant cases, meaning that both the primary tumor and its lymph node metastasis should be evaluated at the same time. A treatment plan based only on the primary tumor phenotype could lead to therapeutic failures if the phenotype of the lymph node metastasis differs from that of the primary tumor.

Skin Photodamage Lesions in a Bilateral Feline Auricular Primary Fibrosarcoma.

As with human species, recent studies also suggest a photoinduced etiopathology for non-epithelial cutaneous tumors in feline species. We report a recent case of a ten-year-old male cat with a white-hair coat and mesenchymal neoplasms of both auricles. Cytology, complete blood count (CBC), serum biochemistry and imaging examinations were performed. After surgery, the samples underwent routinary histopathology and were additionally stained with orcein. A routine analysis yielded values within a normal range and the imaging examination showed no abnormalities, suggesting that the bilateral presentation of neoplasms was primary rather than metastatic. The cytology was inconclusive, but, through histopathology, two well-differentiated fibrosarcomas were diagnosed and histopathological changes related to chronic UV exposure (such as epidermal hyperplasia, stratification disorders, keratinocyte dysplasia and an accumulation of elastotic material) were documented in the skin adjacent to the lesions. An orcein stain succeeded in highlighting elastosis. The elastic fibers lost their regular structure and orientation and appeared to be fragmented, wavy to branched and knotted. A morphometric analysis showed that the amount of elastotic material in the dermis close to the tumors was more than double compared with the more distant areas. Elastosis is considered to be a hallmark of photodamage; thus, an involvement of UV rays in the carcinogenic process of the tumors may be suspected.

Auricular Non-Epithelial Tumors with Solar Elastosis in Cats: A Possible UV-Induced Pathogenesis.

The photoinduced etiopathology of actinic keratosis and squamous cell carcinoma in feline species is well known. This etiology has also been reported for non-epithelial cutaneous tumors in other species. To date, no cases of auricular non-epithelial cutaneous neoplasms erased in a contest of actinic keratosis in cats have been reported. The aim of this study was to describe feline auricular non-epithelial cutaneous neoplasms associated with typical UV-induced cutaneous lesions and solar elastosis. The study was conducted on five feline cases diagnosed with auricular non-epithelial cutaneous tumors (two fibrosarcomas, one mixosarcoma, one epithelioid melanoma and one hemangiosarcoma), selected from the Tumor Registry of the Department of Veterinary Sciences of the University of Pisa (1998-2018). Ten and six feline auricular biopsies of normal skin and skin with actinic keratosis, respectively, were used as controls. Orcein stain was used to investigate solar elastosis. Histological changes related to chronic solar irradiation were documented in the skin adjacent to the neoplastic lesions in the five cats. Considering the anatomical localization and the results of histopathology, this study suggests that non-epithelial cutaneous neoplasms may have a UV-induced etiopathogenesis in the feline species.

Immunohistochemical Characterization of Immune System Cells in Lymphoid Organs from Roe and Fallow Deer.

Roe and Fallow deer are common wild ruminants widely distributed in Italy. Infectious diseases of these species can potentially pose health risks to domestic animals and humans. However, few studies have been conducted in which immune system cells in these species were phenotyped. The aims of this study were to determine the cross-reactivity of a wide anti-human panel of commercial antibodies on formalin-fixed and paraffin-embedded (FFPE) samples and to describe the distribution of roe and fallow deer main immune cell subsets in the lymph nodes and spleen. Twenty retromandibular lymph nodes (RLNs) and spleen samples were collected from 10 roe deer and 10 fallow deer and were tested by a panel of 12 commercial anti-human antibodies. The CD79a, CD20, CD3, Iba-1, MAC387, and AM-3K antibodies were successfully labeled cells in cervine tissue, while the Foxp3 and the CD68 did not show suitable immunostaining. This study supplies the first immunohistochemical description of immune cell subpopulations in non-pathological spleen and RLNs from roe and fallow deer and provides an easily repeatable manual IHC protocol to immunolocalize cervine B-, T-cells, and macrophages subsets in FFPE tissue samples.

Mammary Fibroadenoma in Cats: A Matter of Classification.

Benign mammary lesions are infrequent in cats. Among these, the most common is feline fibroadenomatous change, a hyperplastic/dysplastic change associated with hormonal imbalances. Although never thoroughly described in scientific literature, feline fibroadenomas, which share some morphological features with fibroadenomatous change, have been variably included in classification systems. The aim of this study was to characterise feline mammary fibroadenomas from a histological and immunophenotypical point of view in order to allow the standardisation of classification. Nine cases were retrospectively collected from eight female and one male cat with no history of hormonal stimulation. Diagnostic inclusion criteria were defined and immunohistochemistry was performed. Histologically, nodules were composed of neoplastic epithelial cells arranged in arborizing lobular-like structures surrounded by abundant proliferating stroma. In all analysed cases, epithelial elements showed immunolabelling for pancytokeratin, cytokeratin19, and ß-catenin. Interestingly, five cases showed multifocal epithelial vimentin positivity. Epithelial nuclear oestrogen receptor positivity was observed in three of the nine samples. In all cases, myoepithelial cells did not extend into the interstitium. Stromal cells expressed vimentin, calponin, and mild ß-catenin. The median Ki67 scores were 18% and 8.3% in the epithelial and stromal components, respectively. This study describes, for the first time, the morphological and immunophenotypical features of feline mammary fibroadenoma, highlighting its existence as a separate entity from fibroadenomatous change.

Presence of a mouse mammary tumour virus-like in feline lymphomas: a preliminary study.

The mouse mammary tumour virus (MMTV) is implicated in the aetiology of murine mammary carcinomas and a variant of it, the type B leukemogenic virus, can cause murine thymic lymphomas. Interestingly, a MMTV-like virus is suspected to be involved in human breast cancer and feline mammary carcinomas. However, to date, no cases of MMTV-like sequence amplifications have been described in lymphoid neoplasms in veterinary literature. The aim of this study was to investigate the presence of env nucleotide sequences and protein 14 (p14) of a MMTV-like virus in fifty-three feline lymphoma samples. Our results show that MMTV-like sequences were detected in 5/53 tumours (9.4%): three gastrointestinal lymphomas (one B-type diffuse large, one B-type small non-cleaved, and one T-type diffuse mixed lymphoma); and two nasal lymphomas (one B-type diffuse small cleaved lymphoma and one B-type diffuse mixed lymphoma). P14 expression was detected in the cytoplasm, and rarely in nuclei, exclusively of neoplastic cells from PCR-positive tumours. The correlation between the presence of the MMTV-env like sequences (MMTVels) and p14 antigen was statistically significant in nasal lymphomas. All cats with MMTVels-positive lymphoma had a history of contact with the outdoor environment and/or catteries, and two deceased subjects shared their environment with cats that also died of lymphoma. In conclusion, this study succeeds in demonstrating the presence of MMTVels and p14 in feline lymphomas. The characterization of the immunophenotype of MMTVels-positive lymphomas could contribute to the understanding of a possible role of a MMTV-like virus in feline tumour aetiology. The significant association between the presence of the viral sequences in lymphoid tumours and their nasal localization, together with the data collected through supplementary anamnesis, should be further analysed in order to understand the epidemiology of the virus.

Reproducibility and Feasibility of Classification and National Guidelines for Histological Diagnosis of Canine Mammary Gland Tumours: A Multi-Institutional Ring Study.

Histological diagnosis of Canine Mammary Tumours (CMTs) provides the basis for proper treatment and follow-up. Nowadays, its accuracy is poorly understood and variable interpretation of histological criteria leads to a lack of standardisation and impossibility to compare studies. This study aimed to quantify the reproducibility of histological diagnosis and grading in CMTs. A blinded ring test on 36 CMTs was performed by 15 veterinary pathologists with different levels of education, after discussion of critical points on the Davis-Thompson Foundation Classification and providing consensus guidelines. Kappa statistics were used to compare the interobserver variability. The overall concordance rate of diagnostic interpretations of WP on identification of hyperplasia-dysplasia/benign/malignant lesions showed a substantial agreement (average k ranging from 0.66 to 0.82, with a k-combined of 0.76). Instead, outcomes on ICD-O-3.2 morphological code /diagnosis of histotype had only a moderate agreement (average k ranging from 0.44 and 0.64, with a k-combined of 0.54). The results demonstrated that standardised classification and consensus guidelines can produce moderate to substantial agreement; however, further efforts are needed to increase this agreement in distinguishing benign versus malignant lesions and in histological grading.

M1 and M2 tumour-associated macrophages subsets in canine malignant mammary tumours: An immunohistochemical study.

Among the innate and adaptative immune cells recruited to the tumour site, tumour associated macrophages (TAMs) are particularly abundant and by simplified classification can be classified into (M1) and (M2) TAMs. In the present study, we quantified by immunohistochemistry ionized calcium binding adaptor molecule 1 (Iba1)-positive total and CD204-positive M2-polarized TAMs in 60 canine malignant mammary tumours (CMMTs) to analyze the relationship between M1 or M2 response and the histopathologic features of examined CMMTs, the dogs' body condition score (BCS) and the progression of the neoplastic disease. The mean number of total and CD204+ TAMS were significantly higher in solid and in grade III than in grades I and II carcinomas. Moreover, the mean number of CD204-positive TAMs was significantly higher in CMMTs with lymphatic invasion and necrosis rather than CMMTs without. The presence of higher number of CD204-positive M2-polarized TAMs was associated with a worst outcome of the neoplastic disease: bitches bearing CMMTs with a prevalent M2-polarized TAM response had a median cancer-specific survival time of 449 days, while in animals with a M1-polarized TAM response the median cancer-specific survival time was 1209 days. The results of our study confirm that in CMMTs the presence of a M2-polarized TAMs response might affect the tumour development and behaviour. Finally, it strongly suggests the potential of CD204 expression as a prognostic factor.