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Infections of the genital tract can perturb the fertility in humans and animals. Pathogen recognition and activation of innate immunity onset through the pattern recognition receptor activation, such as Toll-like receptor 4 (TLR4), leading to the production of proinflammatory cytokines and mediators. TLR4 is expressed both on leukocytes and nonimmune cells. Rabbit TLR4 shows great similarity to its human counterpart. Moreover, the TLR4 signalling pathway could be modulated by long-chain polyunsaturated fatty acids (LC-PUFA). The objectives of this study were (i) to determine the expression levels of TLR4 and proinflammatory cytokines in the reproductive hypothalamic-gonadal axis of the male rabbit and (ii) to evaluate if the n-3 PUFA-enriched diets can modify their expression levels in the tissues and LC-PUFA profiles in seminal plasma. Fifteen rabbit bucks (n=5/experimental group) were fed with different diets: commercial standard (group C), rich in extruded linseed (10%, group L), and in fish oil (3%, group FO) for 110 days. TLR4, TNF-α, and IL-1βmRNA were ubiquitously expressed throughout the hypothalamic-gonadal axis. However, TLR4 mRNA expression was lower in the hypothalamus than the epididymis (P<0.01), seminal vesicles (P<0.01), and pituitary gland (P<0.05). Dietary enrichment in PUFA did not modify the gene expression profile nor the histological characteristics of the tissues. Conversely in seminal plasma, rabbits fed with L and FO had lower n-6 (P<0.05), LC-PUFA n-6 (P<0.05), and n-6/n-3 ratio (P<0.05) but higher n-3 (P<0.001) and LC-PUFA n-3 (P<0.01) compared to the control group. Our study builds a map of the gene expression of TRL4 and proinflammatory cytokines in the reproductive hypothalamic-gonadal axis of the male rabbit, fundamental step for understanding the immune defence mechanisms. Diets enriched in LC-PUFA did not affect basal gene expression but modulated sperm fatty acid composition. Finally, rabbit may be an excellent animal model to study the relationship between inflammation and infertility, and the nutritional modulation of immune functions.

HER2 is overexpressed, amplified, and mutated in a subset of human lung cancer. The aim of this study was to investigate HER2 protein overexpression and gene amplification in feline pulmonary carcinomas. Thirteen pulmonary carcinomas were selected and TTF-1 and HER2 expression was evaluated by immunohistochemistry. Fluorescence in situ hybridization (FISH) was performed with a HER2 probe and a BAC probe for the feline chromosome E1p1.12-p1.11 region. Twelve adenocarcinomas and 1 squamous cell carcinoma were diagnosed. TTF-1 was positive in 7 carcinomas (58%). HER2 was overexpressed in 2 (15%), equivocal in 5 (38%), and negative in 6 cases (46%). FISH analysis of HER2 was indeterminate in 2 cases. Three pulmonary carcinomas (27%) had HER2 amplification and 8 cases were not amplified (73%). The significant correlation between HER2 protein overexpression and gene amplification are promising preliminary data, but study of additional cases is needed to confirm HER2 as a target for possible innovative treatments.

The presence and the role of tumor-infiltrating lymphocytes (TILs) in different types of tumors, but particularly in melanoma, has become more and more investigated during the last decade, both in human and veterinary medicine. Melanocytic tumors are quite rare in cats, with diffuse iris melanoma being the most commonly diagnosed in this species. The aim of this study was to characterize the lymphocytic infiltration in feline melanocytic tumors and to analyze their association with the histological features of malignancy recognized in these tumors, as well as with the expression of the most commonly used immunohistochemical markers. Thirty-eight feline melanocytic tumors were retrospectively selected; histological and immunohistochemical characterization of the tumors (histologic criteria of malignancy; S100, Melan A, and PNL2 expression) and of TILs (presence/absence, density, distribution, and grade; CD3, CD20 expression) were performed and associations between them tested. Results showed that TILs grade increased with cellular pleomorphism (P This preliminary study suggests that TILs in feline melanoma may be associated with histologic features of malignancy and loss of melanocytic-specific markers, such as Melan A and PNL2. Further studies, with a larger cohort and follow-up information, are recommended.

Despite promising immunotherapy strategies in human melanoma, there are few studies on the immune environment of canine melanocytic tumors. In humans, the activation of immunosuppressive cell subpopulations, such as regulatory T cells (Tregs) that express forkhead box protein P3 (FoxP3), the engagement of immunosuppressive surface receptors like cytotoxic T lymphocyte antigen (CTLA-4), and the secretion of molecules inhibiting lymphocyte activation, such as indoleamine-pyrrole 2,3-dioxygenase (IDO), are recognized as immunoescape mechanisms that allow tumor growth and progression. The aim of our study was to investigate the expression of these immunosuppression markers in canine melanocytic tumors and to postulate their possible role in melanoma biology and progression. Fifty-five formalin-fixed, paraffin-embedded canine melanocytic tumors (25 oral melanomas; 20 cutaneous melanomas; 10 cutaneous melanocytomas) were selected to investigate the expression of FoxP3, CTLA-4, and IDO by immunohistochemistry and RT-qPCR (real-time quantitative polymerase chain reaction). All of the tested markers showed high gene and protein expression in oral melanomas and were differently expressed in cutaneous melanomas when compared to their benign counterpart. IDO expression was associated with an increased hazard of death both in univariable and multivariable analyses ( P < .05). FoxP3 protein expression >6.9 cells/HPF (high-power field) was an independent predictor of death ( P < .05). CTLA-4 gene and protein expressions were associated with a worse prognosis, but only in the univariable analysis ( P < .05). FoxP3, CTLA-4, and IDO likely play a role in canine melanoma immunoescape. Their expression, if supported by future studies, could represent a prognostic tool in canine melanoma and pave the way to future immunotherapeutic approaches in dogs.

Interactions between tumor cells and tumor microenvironment are considered critical in carcinogenesis, tumor invasion and metastasis. To examine transcriptome changes and to explore the relationship with tumor microenvironment in canine cutaneous melanocytoma and melanoma, we extracted RNA from formalin-fixed, paraffin-embedded (FFPE) specimens and analyzed them by means of RNA-seq for transcriptional analysis. Melanocytoma and melanoma samples were compared to detect differential gene expressions and significant enriched pathways were explored to reveal functional relations between differentially expressed genes. The study demonstrated a differential expression of 60 genes in melanomas compared to melanocytomas. The differentially expressed genes cluster in the extracellular matrix-receptor interaction, protein digestion and absorption, focal adhesion and PI3K-Akt (phosphoinositide 3-kinase/protein kinase B) signaling pathways. Genes encoding for several collagen proteins were more commonly differentially expressed. Results of the RNA-seq were validated by qRT-PCR and protein expression of some target molecules was investigated by means of immunohistochemistry. We hypothesize that the developing melanoma actively promotes collagen metabolism and extracellular matrix remodeling as well as enhancing cell proliferation and survival contributing to disease progression and metastasis. In this study, we also detected unidentified genes in human melanoma expression studies and uncover new candidate drug targets for further testing in canine melanoma.

Osteochondrosis is a developmental orthopedic disease affecting growing cartilage in young horses. In this study we compared the proteomes of equine chondrocytes obtained from healthy and osteochondrotic cartilage using a label-free mass spectrometry approach. Quantitative changes of some proteins selected for their involvement in different functional pathways highlighted by the bioinformatics analysis, were validated by western blotting, while biochemical alterations of extracellular matrix were confirmed via Raman spectroscopy analysis. In total 1637 proteins were identified, of which 59 were differentially abundant. Overall, the results highlighted differentially represented proteins involved in metabolic and functional pathways that may be related to the failure of the endochondral ossification process occurring in osteochondrosis. In particular, we identified proteins involved in extracellular matrix degradation and organization, vitamin metabolism, osteoblast differentiation, apoptosis, protein folding and localization, signalling and gene expression modulation and lysosomal activities. These results provide valuable new insights to elucidate the underlying molecular mechanisms associated with the development and progression of osteochondrosis.Significance: Osteochondrosis is a common articular disorder in young horses mainly due to defects in endochondral ossification. The pathogenesis of osteochondrosis is still poorly understood and only a limited number of proteomic studies have been conducted. This study provides a comprehensive characterization of proteomic alterations occurring in equine osteochondrotic chondrocytes, the only resident cell type that modulates differentiation and maturation of articular cartilage. The results evidenced alterations in abundance of proteins involved in functional and metabolic pathways and in extracellular matrix remodelling. These findings could help clarify some molecular aspects of osteochondrosis and open new fields of research for elucidating the pathogenesis of this disease.

Canine testicular tumors account for about 90 % of tumors affecting the male genitalia. Seminomas (SEM), Sertoli cell tumors (SCT), and interstitial cell tumors (ICT) are the most common histological diagnoses, but their incidence shows high variability among studies. Our aim is to report the results on the analysis of testicular tumors recorded by the population-based Umbria Canine Cancer Registry (CCR) for a 5-year time period and to assess the value of tumor major diameter, measured during trimming, in discriminating neoplastic from non-neoplastic lesions. The study was conducted on 388 testicular tumors (on 1969 total male tumors) diagnosed in 355 dogs from 2014 to 2018. The median incidence was 35 cases/100,000 dogs, with a proportion equal to 19,7 % of all tumors. The incidence on neutered male dogs was 352/100,000. Most tumors were ICTs (50 %), with fewer SEMs and SCTs (29 % and 17 %, respectively). Only 3 % of tumors were mixed germ cell-sex cord-stromal tumors (MGC-SCST). Ten percent of cases had multiple tumors in the same testicle, with SEM-ICT being prevalent (69.2 %). Tumors in cryptorchid testes were 5.9 % of the total, mostly SCT (60.9 %). Mean age at diagnosis was 10.7 ± 2.7, with similar values for different tumor types. Significant incidence ratios (IRR) were found in Golden retriever (IRR 7.18, CI95 % 4.72-10.92) and in English cocker spaniel (IRR 2.30, CI95 % 1.28-4.13) when compared with mixed breed dogs. A value of 0.3 cm (major diameter) of lesions at the moment of trimming was helpful in discriminating a final tumor histological diagnosis from a non-tumor lesion. Since the number of tumors included in this evaluation was limited, further studies to confirm the diagnostic value of this measure are recommended. Our results provided information on the incidence of canine testicular tumors in Umbria region that can be compared with future results and incidence from other geographical areas if provided with reliable data on the total population, can offer solid information on the incidence and proportion of different tumor types in specific territories, contributing also to the supervision of its inhabitants' health. Moreover, pathological data such as the major diameter of tumors can be obtained and contribute to diagnostic routine and standardization.

Osteochondrosis is a failure of the endochondral ossification that affects developing joints in humans and several animal species. It is a localized idiopathic joint disorder characterized by focal chondronecrosis and growing cartilage retention, which can lead to the formation of fissures, subchondral bone cysts, or intra-articular fragments. Osteochondrosis is a complex multifactorial disease associated with extracellular matrix alterations and failure in chondrocyte differentiation, mainly due to genetic, biochemical, and nutritional factors, as well as traumas. This study describes the main proteomic alterations occurring in chondrocytes isolated from osteochondrotic cartilage fragments. A comparative analysis performed on equine osteochondrotic and healthy chondrocytes showed 26 protein species as differentially represented. In particular, quantitative changes in the extracellular matrix, cytoskeletal and chaperone proteins, and in cell adhesion and signaling molecules were observed in osteochondrotic cells, compared to healthy controls. Functional group analysis annotated most of these proteins in &ldquo, growth plate and cartilage development&rdquo, while others were included in &ldquo, glycolysis and gluconeogenesis&rdquo, &ldquo, positive regulation of protein import&rdquo, cell&ndash, cell adhesion mediator activity&rdquo, and &ldquo, mitochondrion nucleoid&rdquo, These results may help to clarify some chondrocyte functional alterations that may play a significant role in determining the onset and progression of equine osteochondrosis and, being related, of human juvenile osteochondrosis.

The study of the immune response in several types of tumours has been rapidly increasing in recent years with the dual aim of understanding the interactions between neoplastic and immune cells and their importance in cancer pathogenesis and progression, as well as identifying targets for cancer immunotherapy. Despite being considered one of the most immunogenic tumour types, melanoma can progress in the presence of abundant lymphocytic infiltration, therefore suggesting that the immune response is not able to efficiently control tumour growth. The purpose of this study was to investigate whether the density, distribution and grade of tumour-infiltrating lymphocytes (TILs) in 97 canine melanocytic tumours is associated with histologic indicators of malignancy and can be considered a prognostic factor in the dog. As a further step in the characterization of the immune response in melanocytic tumours, an immunohistochemical investigation was performed to evaluate the two main populations of TILs, T-lymphocytes (CD3

Human melanoma is one of the deadliest forms of cancer, with poor prognosis and high resistance to chemotherapy and radiotherapy. The discovery of immunosuppressive mechanisms in the human melanoma microenvironment led to the use of new prognostic markers and to the development of immunotherapies targeting immune checkpoint molecules. Immunoescape mechanisms in canine melanoma have not yet been investigated, and no such immunotherapy has been tested. The aim of this study was to provide preliminary data on the expression of transcription factor forkhead box protein P3 (FoxP3) and indoleamine 2,3-dioxygenase (IDO) in primary canine melanocytic tumors and to investigate their prognostic role. Formalin-fixed, paraffin-embedded samples from 74 canine melanocytic tumors (26 oral melanomas, 23 cutaneous melanomas, and 25 cutaneous melanocytomas) were retrospectively evaluated by immunohistochemistry to explore the expression of FoxP3 and IDO. An increased risk of death due to melanoma was associated with a higher number of FoxP3+ cells per high-power field (FoxP3+/HPF), a higher percentage of CD3+ cells that were also FoxP3+ infiltrating and surrounding the tumor (%FoxP3), and a higher number of IDO+ cells/HPF (IDO+/HPF). A prognostic value for FoxP3 and IDO is suggested by our study, with optimal cutoffs of 14.7 FoxP3+ cells/HPF, 6.1 IDO+ cells/HPF, and 12.5% FoxP3+ cells. Both markers were also associated with tumor type. Multivariable analysis identified IDO+/HPF ( P < .001) as an independent prognostic marker. Even though stratification by diagnosis caused a loss of significance, results from the present study suggest a prognostic role for IDO and FoxP3, possibly related to the establishment of an immunosuppressive microenvironment.

A 14-year-old female spayed Dachshund was presented with generalized scaling, erythema, pruritus, poor quality of hair coat, and progressive weight loss. Cutaneous epitheliotropic T-cell lymphoma (CETCL) was suspected. Skin biopsies were suggestive of CETCL. However, immunohistochemistry revealed the presence of numerous CD20+ and CD3+ cells. Clonality assay demonstrated a clonal T-cell receptor gamma rearrangement and a polyclonal IgH gene rearrangement. Double-label immunofluorescence confirmed coexpression of CD3 and CD20 by neoplastic cells. By double immunohistochemistry, neoplastic cells were CD3+ and PAX5–. The results are compatible with a CD3+, CD20+ CETCL. Coexpression of CD20 and CD3 has been recognized in peripheral T-cell lymphomas. Although documented in human CETCL, it has not been reported in canine CETCL. The pathogenetic basis of CD20 expression in mycosis fungoides is explored.

Background Feline herpesvirus ulcerative dermatitis is an uncommon skin disease in cats, with a predominantly facial distribution characterized by massive infiltration of eosinophils and, occasionally, predominant neutrophils. Objective To describe the clinical and histopathological features of a putative atypical case of feline herpesvirus dermatitis. Animal A 10-month-old, intact male, European cat was presented with chronic monolateral ulcerative dermatitis with adherent crusts on the left pinna. The lesion had been present for six months and worsened after the administration of corticosteroids. Methods Clinical and histopathological examination, immunohistochemistry, nested PCR and transmission electron microscopy (TEM). Results Histological examination of skin biopsies showed multifocal ulcerative and necrotic lesions, involving the superficial and deep dermis covered by thick haemorrhagic and serocellular crusts. The superficial, medium and deep dermis was heavily infiltrated with mast cells and plasma cells, with a lower number of neutrophils and eosinophils. In the nuclei of some cells in the deep dermis, whose histotype was unrecognizable with routine haematoxylin and eosin stain, intranuclear eosinophilic inclusion bodies were noticed. Nested PCR and TEM supported the hypothesis of FeHV-1-induced dermatitis. Conclusions and clinical importance This case is noteworthy for the infrequent location on the pinna and the atypical histopathological features of the lesion, with a predominant infiltration of mast cells and plasma cells. Our findings suggest that herpesvirus dermatitis should be listed as a differential diagnosis in case of ulcerative dermatitis when the location and histological features are atypical.

Testudinid herpesviruses (TeHVs) have a worldwide distribution among tortoises. However, information such as risk factors promoting the occurrence or the recrudescence of the associated disease and the mid-term sequelae of an outbreak comprising the extent and dynamic of the viral shedding have been only minimally investigated. Critical management information is also lacking or anecdotal. Furthermore, major aspects of the viral pathogenesis including the likelihood of vertical transmission of the virus are virtually unknown. The present study describes the occurrence and the management of an outbreak of Testudinid herpesvirus genotype 3 (TeHV3) in a large, private collection of chelonians. Clinical, pathological, molecular and serological characterization of the outbreak were carried out. Seventy-five percent of the infected tortoises died. Complementation of molecular and serological testing was a critical point for successful management implementations. A case-control study was performed to analyze possible risk factors associated with the infection. Furthermore, a subgroup of six asymptomatic infected tortoises was monitored for two consecutive seasons after the outbreak: all the infected tortoises were determined to be intermittent shedders, except one, which was a persistent shedder. Post-hibernation was associated with the highest number of shedders. Finally, evidence of the most likely vertical transmission of the virus was obtained for the first time.

A cholangiocarcinoma is reported in an 18-yr-old, female African lion (Panthera leo). The primary tumor consisted of multifocal to coalescing, hepatic, white–yellow masses distributed throughout the liver lobes. Metastases were present in regional lymph nodes, peritoneal surface, and lungs. Histologically, the tumor was characterized by a tubular pattern with alcian- and periodic acid–Schiff–positive secretory material in cystic spaces. The neoplastic cells were positive to broad-spectrum cytokeratins. Histochemical and immunohistochemical stains were consistent with bile duct carcinoma. Biliary tumors arising from the gallbladder have been reported in lions. However, to the authors' knowledge, this is the first case of intrahepatic bile duct carcinoma reported in an African lion.

Feline injection-site sarcoma (FISS) is an aggressive tumor believed to arise from the proliferation of fibroblasts and myofibroblasts in areas of chronic inflammation, particularly at sites of injection. Local recurrence is frequent after surgical excision. Gelatinases (MMP-2 and MMP-9) and their inhibitor (TIMP-2) are endopeptidases pivotal in extracellular matrix remodeling and therefore in tumor invasiveness. The aim of this study was to investigate the immunohistochemical expression of MMP-2, MMP-9, and TIMP-2 in FISS to assess their usefulness as prognostic factors. Size, soft tissue sarcoma (STS) grading system, depth of infiltration, surgical margins, and Ki-67 index were evaluated as additional prognostic markers. Twenty-four cases of primary FISS were classified according to clinical follow-up as nonrecurrent (NR, n = 14; 58.3%) and recurrent (R, n = 10; 41.7%). MMP-2, MMP-9, and TIMP-2 were variably expressed in the FISS examined, confirming their role in tumor invasiveness, yet they did not show significant differences between the R and NR groups. These results could be due to different tumor stages or to the multiple activities of these enzymes, not limited to ECM remodeling. The immunohistochemical expression of these enzymes considered alone does not seem to be useful as a prognostic marker. STS grading system, depth of infiltration, surgical margins, and Ki-67 index did not relate to recurrence. Instead, the size of the tumor, measured after formalin fixation, with an optimal cutoff of 3.75 cm (accuracy = 86%; P < .05), and the mitotic count, with an optimal cutoff of 20 mitoses/10 HPF (accuracy = 80%; P < .05), could be evaluated as useful prognostic markers.

In recent years, there has been increasing evidence of the potential pathogenic significance of equine gammaherpesviruses in the horse. In humans, cattle and mice, gammaherpesviruses have already been associated with uterine infection. The aim of the present study was to investigate the presence of gammaherpesviruses in uterine flushings of mares with reproductive problems and to evaluate if there was a possible statistical association with clinical and laboratory findings in these cases. A total of 80 uterine flushings were collected from 61 mares with different reproductive problems and these were tested for equine herpesviruses (EHV) 1–5 by PCR. In the case of each mare in the study, the age, history of infertility, presence of anatomical defects in the reproductive tract, presence of systemic or local disease at time of sampling, phase in the oestrous cycle, post-partum interval, nature of uterine lavage performed (low versus large volume lavage), cytological and bacteriological examination results from the uterine flushing, and PCR herpesvirus results were recorded. Univariate analysis and multivariable logistic regression models were used to identify possible statistical associations and risk factors. Nine out of 61 mares (14.7%) had EHV-5 DNA in their uterine flushings. Co-infections with EHV-1 and EHV-2 were present in two cases. Of all the variables analyzed, only the cytological examination findings were associated with EHV-5 PCR positive results, both on univariate and multivariable analysis, especially in cases with an inflammation score of 3. It is postulated that presence of EHV-5 infection in the non-pregnant uterus may have a role to play in reproductive dysfunction and have a negative consequence on the pregnant uterus. Additional studies involving both healthy mares and mares with reproductive problems need to be performed, however, to elucidate whatever role equine gammaherpesviruses may play in the reproductive tract. This would be very worthwhile, since reproductive problems can have a significant impact on the equine breeding industry. Gaining a greater understanding of its causes could lead to new approaches for prevention and treatment.

Background Feline Progressive Histiocytosis (FPH) is a cutaneous dendritic cell neoplasm characterized by slow progression and spread to internal organs in the terminal stage. FPH is often misdiagnosed as an inflammatory reaction and has not been fully characterized from a cytologic diagnostic perspective. Objectives The purpose of the study was to characterize the cytologic and immunocytochemical aspects useful for FPH diagnosis. Methods Fine-needle aspiration cytologic samples of 5 cases of FPH confirmed by skin biopsy and necropsy were evaluated. Immunocytochemistry with antibodies recognizing CD1a, CD1c, CD3, CD11b, CD18, CD21, and MHCII was performed on air-dried, acetone-fixed smears. E-cadherin expression was assessed on paraffin-embedded skin biopsies. Transmission electron microscopy (TEM) was performed in one case. Results Main cytologic findings on variably cellular samples were characterized by single to cohesive large, round to polygonal cells with intermediate to low N/C ratio, abundant clear homogeneous cytoplasm, and round to oval nuclei with rare bi- to multinucleated atypical cells, associated with low numbers of small lymphocytes and/or neutrophils. Neoplastic cells expressed CD1a, CD1c, CD11b, CD18, and MHCII. Anti-CD3 antibodies identified reactive T cells admixed with the neoplastic cells. E-cadherin expression was observed in all but one case. TEM failed to identify Birbeck granules in one case. Conclusions FPH is a distinctive neoplastic lesion composed of nonphagocytizing histiocytes variably admixed with neutrophils and small mature lymphocytes. Immunocytochemical analysis with CD1 is mandatory to confirm a dendritic cell origin. Immunocytochemistry and cytomorphology allowed the specific and rapid diagnosis of FPH on cytologic samples.

CPV1 (also called COPV) is a papillomavirus responsible for oral papillomatosis in young dogs. The involvement of this viral type in oral oncogenesis has been hypothesized in oral squamous cell carcinomas (SCCs), but has never been investigated in other neoplastic and hyperplastic oral lesions of dogs. Aim of this study was to investigate the presence of CPV1 in different neoplastic and hyperplastic lesions in order to assess its role in canine oral oncogenesis; according to the results obtained, a second aim of the study was to define if the dog can be considered a valid animal model for oral high risk HPV-induced tumors. Eighty-eight formalin-fixed, paraffin-embedded (FFPE) canine oral lesions including 78 oral tumors (papillomas, SCCs, melanomas, ameloblastomas, oral adenocarcinomas) and 10 hyperplastic lesions (gingival hyperplasia) were investigated with immunohistochemistry for the presence of papillomavirus L1 protein and with Real-Time PCR for CPV1 DNA. RT-PCR for RNA was performed on selected samples. All viral papillomas tested were positive for immunohistochemistry and Real-time PCR. In 3/33 (10%) SCCs, viral DNA was demonstrated but no viral RNA could be found. No positivity was observed both with immunohistochemistry and Real-Time PCR in the other hyperplastic and neoplastic lesions of the oral cavity of dogs. Even though the finding of CPV1 DNA in few SCCs in face of a negative immunohistochemistry could support the hypothesis of an abortive infection in the development of these lesions, the absence of viral RNA points out that CPV1 more likely represents an innocent bystander in SCC oncogenesis. The study demonstrates a strong association between CPV1 and oral viral papillomas whereas viral contribution to the pathogenesis of other oral lesions seems unlikely. Moreover, it suggests that a canine model of CPV1 infection for HPV-induced oncogenesis could be inappropriate.

Epithelial-to-mesenchymal transition (EMT) is a multistep process, important in tumour invasion and metastasis, characterized by loss of epithelial markers, redistribution of β-catenin and gain of mesenchymal markers. HYPOSTHESIS/OBJECTIVES: Our aim was to investigate the immunohistochemical aberrant expression of cytokeratin, vimentin, survivin and heat shock protein 72 (Hsp72) in canine cutaneous epithelial tumours, to understand the association of expression of these molecules with features of malignancy and their role in the EMT phenotype.Ten canine squamous cell carcinomas (SCCs; one with lymph node metastasis), 30 canine hair follicle tumours (six pilomatricomas, eight infundibular keratinizing acanthomas, six trichoepitheliomas and 10 trichoblastomas) and five normal skin samples were investigated by immunohistochemistry using specific anti-vimentin, -cytokeratin, -survivin and -Hsp72 antibodies. A semi-quantitative method was used to analyse the results, as follows: 0 to5%; ≥ 5 to10%; ≥ 10 to25%; and ≥ 25% of positive cells. Immunofluorescence was performed to investigate survivin-vimentin and survivin-Hsp72 colocalization in selected SCCs. Results - In malignant hair follicle tumours and SCCs, a reduced intensity of cytokeratin and increased survivin and Hsp72 expression were observed. In SCCs, loss of cytokeratin expression and vimentin immunolabelling, suggestive of the EMT phenotype, were evident in5% of neoplastic cells in the front of tumour invasion. In the same areas, strong nuclear survivin and cytoplasmic Hsp72 staining was evident, often colocalizing. Only a few neoplastic cells in the front of tumour invasion showed vimentin-survivin colocalization.A possible simultaneous involvement of survivin and Hsp72 in tumour invasion and the multistep process of EMT of cutaneous epithelial tumours of dogs is suggested.

Background - Although cutaneous stem cells have been implicated in skin tumourigenesis in humans, no studies have been conducted to elucidate the presence and the possible role of stem cells in hair follicle tumours in the dog. Hypothesis - Stem cell markers are expressed in canine epidermal and follicular tumours and can be used to better understand the biology and origin of these tumours. Animals and Methods - In the present study, normal skin sections and 44 follicular tumours were retrospectively investigated for the immunohistochemical expression of keratin15 (K15) and nestin. In addition, 30 squamous cell carcinomas were evaluated for K15 expression. Results - In normal skin, K15 and nestin were expressed in the outer root sheath cells of the isthmic portion of the hair follicle (bulge region), and K15 expression was also scattered in the basal cell layer of the epidermis. Infundibular keratinizing acanthomas, pilomatricomas and squamous cell carcinomas were mostly negative for K15, trichoblastomas were moderately to strongly positive, tricholemmomas were either negative or strongly positive, and trichoepitheliomas had heterogeneous staining. Nestin expression was generally faint in all follicular tumours. Conclusions and clinical importance - Our results show that K15 can be a reliable marker for investigating the role of stem cells in hair follicle tumours of the dog, while nestin was judged to be a nonoptimal marker. Furthermore, our study suggests that hair follicle stem cells are present in the bulge region of hair follicles and could possibly play a role in tumourigenesis of canine tumours originating from this portion of the follicle, namely trichoblastomas, tricholemmomas and trichoepitheliomas. The loss of K15 expression in squamous cell carcinomas compared with normal skin suggests that this event could be important in the malignant transformation.

Feline Rhodococcus equi infection is rare, despite the bacteria is widespread in the environment. R equi infection is typically observed in equine species but the infection has also been reported in dogs, cats and other domestic animals. There are a few reports regarding pulmonary R equi infection in cats and the disease appears to be limited to the skin and the subcutaneous tissue. This report describes the pathological, microbiological and the virulence features associated with an acute necrosuppurative pneumonia in a cat. To the best of our knowledge, this is the first report of feline pulmonary R equi infection in Italy.

Background – Epithelial-to-mesenchymal transition (EMT) is a multistep process, important in tumour invasion and metastasis, characterized by loss of epithelial markers, redistribution of β-catenin and gain of mesenchymal markers. Hyposthesis/Objectives – Our aim was to investigate the immunohistochemical aberrant expression of cytokeratin, vimentin, survivin and heat shock protein 72 (Hsp72) in canine cutaneous epithelial tumours, to understand the association of expression of these molecules with features of malignancy and their role in the EMT phenotype. Methods – Ten canine squamous cell carcinomas (SCCs; one with lymph node metastasis), 30 canine hair follicle tumours (six pilomatricomas, eight infundibular keratinizing acanthomas, six trichoepitheliomas and 10 trichoblastomas) and five normal skin samples were investigated by immunohistochemistry using specific anti-vimentin, -cytokeratin, -survivin and -Hsp72 antibodies. A semi-quantitative method was used to analyse the results, as follows: 0 to

Background – Although cutaneous stem cells have been implicated in skin tumourigenesis in humans, no studies have been conducted to elucidate the presence and the possible role of stem cells in hair follicle tumours in the dog. Hypothesis – Stem cell markers are expressed in canine epidermal and follicular tumours and can be used to better understand the biology and origin of these tumours. Animals and Methods – In the present study, normal skin sections and 44 follicular tumours were retrospectively investigated for the immunohistochemical expression of keratin 15 (K15) and nestin. In addition, 30 squamous cell carcinomas were evaluated for K15 expression. Results – In normal skin, K15 and nestin were expressed in the outer root sheath cells of the isthmic portion of the hair follicle (bulge region), and K15 expression was also scattered in the basal cell layer of the epidermis. Infundibular keratinizing acanthomas, pilomatricomas and squamous cell carcinomas were mostly negative for K15, trichoblastomas were moderately to strongly positive, tricholemmomas were either negative or strongly positive, and trichoepitheliomas had heterogeneous staining. Nestin expression was generally faint in all follicular tumours. Conclusions and clinical importance – Our results show that K15 can be a reliable marker for investigating the role of stem cells in hair follicle tumours of the dog, while nestin was judged to be a nonoptimal marker. Furthermore, our study suggests that hair follicle stem cells are present in the bulge region of hair follicles and could possibly play a role in tumourigenesis of canine tumours originating from this portion of the follicle, namely trichoblastomas, tricholemmomas and trichoepitheliomas. The loss of K15 expression in squamous cell carcinomas compared with normal skin suggests that this event could be important in the malignant transformation. Resume Contexte – Bien que les cellules souches cutanees aient ete impliquees dans la genese des tumeurs cutanees chez l’homme, aucune etude n’a ete menee pour determiner la presence et le role potentiel des cellules souches dans les tumeurs des follicules pileux chez le chien. Hypothese – Les marqueurs des cellules souches sont exprimes par les tumeurs canines epidermiques et folliculaires et peuvent etre utilises pour une meilleure comprehension de la biologie et de l’origine de ces tumeurs. Sujets et Methodes – Dans cette etude, l’expression immunohistochimiques de keratine 15 (K15) et de nestine a ete retrospectivement etudiee sur des sections cutanees normales et 44 tumeurs folliculaires. De plus, l’expression de K15 a eteevaluee pour 30 carcinomes epidermoides. Resultats – Dans la peau normale, K15 et nestine sont exprimees par les cellules de la gaine epitheliale externe de la portion isthmique du follicule pileux (region du bulge); K15 s’exprime aussi de facon disseminee dans la couche basale de l’epiderme. Les acanthomes keratinises infundibulaires, les pilomatrichomes et les carcinomes epidermoides etaient principalement negatifs pour K15, les trichoblastomes etaient moderement a fortement positifs, les tricholemmomes etaient soit negatifs soit fortement positifs enfin, les trichoepitheliomes avaient une coloration heterogene. L’expression de nestine etait generalement faible pour toutes les tumeurs folliculaires. Conclusion et importance Clinique – Nos resultats montrent que K15 peut etre un marqueur fiable pour l’etude du role des cellules souches dans les tumeurs du follicule pileux du chien tandis que la nestine se revele pas etre un marqueur adapte. En outre, notre etude suggere que les cellules souches des follicules pileux sont presentes dans la region du bulge des follicules pileux et pourraient jouer un role dans la genese des tumeurs canines provenant de cette portion du follicule, a savoir les trichoblastomes, les tricholemmomes et les trichoepitheliomes. La perte de l’expression de K15 par les carcinomes epidermoides comparea la peau normale suggere que cet evenement pourrait etre important dans la transformation maligne. Resumen Introduccion – aunque las celulas madre cutaneas han sido implicadas en tumorigenesis de la piel en seres humanos, no se ha conducido ningunos estudios para aclarar la presencia y el papel posible de las celulas madre en tumores del foliculo piloso en el perro. Hipotesis – los marcadores de la celula madre se expresan en tumores epidermicos y foliculares caninos y se pueden utilizar para entender mejor la biologia y el origen de estos tumores. Animales y metodos – en este estudio, se investigaron secciones de piel normal y de 44 tumores de forma retrospectiva para la expresion inmunohistoquimica de queratina 15 (K15) y de nestina. Ademas, se evaluaron 30 carcinomas de celulas escamosas para la expresion de K15. Resultados – en piel normal, la K15 y nestina se expresaron en celulas de la vaina externa de la raiz del pelo en la porcion istmica del foliculo del pelo (region de la protuberancia), y tambien hubo expresion dispersa de K15 en la capa basal de la epidermis. Los acantomas queratinizantes del infundibulo, los pilomatricomas y los carcinomas de celulas escamosas fueron todos negativos para K15, los tricoblastomas fueron de moderado a intensamente positivos, los tricolemmomas fueron negativos o intensamente positivos y los tricoepiteliomas presentaron tincion heterogenea. La expresion de nestina fue generalmente debil en todos los tumores foliculares. Conclusiones e importancia clinica – nuestros resultados demuestran que la K15 puede ser un marcador fiable para investigar el papel de las celulas madre en los tumores del foliculo del pelo del perro, mientras que la nestina fue considerada un marcador no optimo. Ademas, nuestro estudio sugiere que las celulas madre del foliculo piloso estan presentes en la region del la protuberancia de los foliculos del pelo y podrian desempenar posiblemente un papel en la tumorigenesis de los tumores caninos que se originan de esta porcion del foliculo, a saber tricoblastomas, tricolemmomas y tricoepiteliomas. La perdida de la expresion K15 en los carcinomas de celulas escamosas comparada con la piel normal sugiere que este proceso podria ser importante en la transformacion maligna. Zusammenfassung Hintergrund – Obwohl kutane Stammzellen mit der Entstehung von Tumoren beim Menschen in Verbindung gebracht werden, sind noch keine Studien durchgefuhrt worden, um ihr Vorkommen und ihre mogliche Rolle bei der Entstehung von Haarfollikeltumoren beim Hund zu beleuchten. Hypothese – Stammzellmarker werden in caninen epidermalen und follikularen Tumoren exprimiert und konnen dazu verwendet werden, die Biologie und den Ursprung dieser Tumoren besser zu verstehen. Tiere und Methoden – In der vorliegenden Studie wurden normale Hautschnitte und 44 Haarfollikel Tumoren retrospektiv auf ihre immunhistochemische Exprimierung von Keratin 15 (K15) und Nestin untersucht. Zusatzlich wurden 30 Plattenepithelkarzinome auf K15 Exprimierung untersucht. Ergebnisse – In normaler Haut wurden K15 und Nestin in den Zellen der auseren Wurzelschicht des Isthmus-Abschnitts des Haarfollikels (“Bulge Region”) exprimiert und eine K15 Exprimierung konnte unregelmasig verteilt in der Basalschicht der Epidermis gefunden werden. Keratinisierende Akanthome im Infundibulum, Pilomatrikome und Plattenepithelkarzinome waren weitgehend negativ fur K15, Trichoblastome waren moderat bis stark positiv, Tricholemmome waren entweder negativ oder stark positiv und Trichoepitheliome zeigten eine heterogene Farbung. Nestin Exprimierung war in allen Tumoren der Haarfollikel generell schwach. Schlussfolgerungen und klinische Bedeutung – Unsere Ergebnisse zeigen, dass K15 ein verlasslicher Marker bei der Untersuchung der Rolle der Stammzellen der Haarfollikeltumoren des Hundes sein kann, wahrend Nestin als nicht optimaler Marker beurteilt wurde. Weiters weist unsere Studie darauf hin, dass Haarfollikelstammzellen in der Bulge Region der Haarfollikel vorkommen und moglicherweise bei der Entstehung der Tumoren des Hundes aus diesem Abschnitt des Haarfollikels eine Rolle spielen, und zwar bei Trichoblastomen, Tricholemmomen und Trichoepitheliomen. Der Verlust der K15 Exprimierung in den Plattenepithelkarzinomen im Vergleich zu normaler Haut weist darauf hin, dass dieser Vorgang bei der malignen Umwandlung wichtig sein konnte.