Serum Extracellular Vesicles Cargo Approach in Bitches with Mammary Tumors.

This study investigated serum extracellular vesicles (EVs) in bitches with mammary neoplasms, in order to understand their size, shape, and concentration, as well as their association with tumor malignancy. Thirty bitches were categorized into control (n = 10), mammary tumor grades I and II (GI, n = 13), and grade III (GII, n = 7). Serum was separated from blood collected during mastectomy, and EVs were isolated using size exclusion chromatography. The analysis revealed no significant differences in EV concentrations among groups, with similar concentrations for control, GI, and GII. Ninety-one proteins were identified in EV-enriched samples, with six showing varied abundance across groups. Notably, keratin 18 was highly abundant in GI, while sushi domain-containing protein, EvC ciliary subunit 2, and the joining chain of multimeric IgM and IgA were increased in GII. Additionally, protocadherin 17 and albumin were upregulated in both GI and GII. ROC curves identified potential biomarkers for differentiating tumor grades. Enrichment pathway analysis revealed AFP gene upregulation in the GI. Mass spectrometry proteomics data were deposited in Mendeley Data. The study provides valuable insights into serum EV characterization in bitches, suggesting keratin 18 and protocadherin 17 as potential biomarkers for canine mammary neoplasia, with implications for future diagnostic and therapeutic strategies.

Simultaneous occurrence of inflammatory bowel disease and trichomonosis in a Maine coon cat.

A 2-year-old, 4.2 kg, spayed female, Maine coon cat was referred to the veterinary hospital for evaluation of hyporexia, slow growth, and chronic, intermittent, mucoid, bloody, voluminous, and fetid diarrhea. The diarrhea had been observed since the cat was acquired from a cattery at 4 months of age; with acute worsening in the 5 d before presentation. Abdominal palpation revealed moderate pain. Ultrasonographic examination showed thickening of the jejunal wall and ileal loops, increased echogenicity of the jejunal mucosa, and enlargement of the jejunal and ileocolic lymph nodes. Histopathology of full-thickness intestinal biopsies showed moderate, diffuse, lymphoplasmacytic, erosive enteritis with hemorrhage and edema. Diffuse, lymphoplasmacytic, erosive colitis with mild, interstitial fibrosis and hemorrhage was also noted. The ileocecal lymph node biopsy showed eosinophilic lymphadenitis. Based on the immunohistochemical evaluation of intestinal samples with CD3 and CD79a antibodies, a diagnosis of lymphoma was ruled out. Fecal polymerase chain reaction testing was positive for Tritrichomonas foetus. Based on these results, inflammatory bowel disease and trichomonosis were diagnosed. Treatment for the cat included a hypoallergenic diet and an oral omega-3 fatty acid supplement, in conjunction with prednisolone, to manage the inflammatory bowel disease. Ronidazole was administered to target the Tritrichomonas foetus. The cat was clinically normal during a follow-up examination after 6 months of treatment.

Apparition simultanée d'une maladie inflammatoire de l'intestin et d'une trichomonose chez un chat Maine coon. Une chatte Maine coon de 2 ans, pesant 4,2 kg, stérilisée, a été référée à l'hôpital vétérinaire pour une évaluation d'hyporexie, de croissance lente et de diarrhée chronique, intermittente, mucoïde, sanglante, volumineuse et fétide. La diarrhée avait été observée depuis que le chat avait été acquis en chatterie à l'âge de 4 mois; avec une aggravation aiguë dans les 5 jours avant la présentation. La palpation abdominale a révélé une douleur modérée. L'examen échographique a montré un épaississement de la paroi jéjunale et des anses iléales, une augmentation de l'échogénicité de la muqueuse jéjunale et une hypertrophie des ganglions lymphatiques jéjunaux et iléocoliques. L'histopathologie des biopsies intestinales de pleine épaisseur a montré une entérite modérée, diffuse, lymphoplasmocytaire, érosive avec hémorragie et oedème. Une colite érosive diffuse, lymphoplasmocytaire avec fibrose interstitielle légère et hémorragie a également été notée. La biopsie ganglionnaire iléo-caecale montrait une lymphadénite à éosinophiles. Sur la base de l'évaluation immunohistochimique d'échantillons intestinaux avec des anticorps CD3 et CD79a, un diagnostic de lymphome a été écarté. Le test de réaction en chaîne par la polymérase sur les matières fécales était positif pour Tritrichomonas foetus. Sur la base de ces résultats, une maladie inflammatoire de l'intestin et une trichomonose ont été diagnostiquées. Le traitement du chat comprenait un régime hypoallergénique et un supplément oral d'acides gras oméga-3, en association avec de la prednisolone, pour gérer la maladie inflammatoire de l'intestin. Le ronidazole a été administré pour cibler Tritrichomonas foetus. Le chat était cliniquement normal lors d'un examen de suivi après 6 mois de traitement.(Traduit par Dr Serge Messier).

Proteomics Approach of Rapamycin Anti-Tumoral Effect on Primary and Metastatic Canine Mammary Tumor Cells In Vitro.

Rapamycin is an antifungal drug with antitumor activity and acts inhibiting the mTOR complex. Due to drug antitumor potential, the aim of this study was to evaluate its effect on a preclinical model of primary mammary gland tumors and their metastases from female dogs. Four cell lines from our cell bank, two from primary canine mammary tumors (UNESP-CM1, UNESP-CM60) and two metastases (UNESP-MM1, and UNESP-MM4) were cultured in vitro and investigated for rapamycin IC50. Then, cell lines were treated with rapamycin IC50 dose and mRNA and protein were extracted in treated and non-treated cells to perform AKT, mTOR, PTEN and 4EBP1 gene expression and global proteomics by mass spectrometry. MTT assay demonstrated rapamycin IC50 dose for all different tumor cells between 2 and 10 µM. RT-qPCR from cultured cells, control versus treated group and primary tumor cells versus metastatic tumor cells, did not shown statistical differences. In proteomics were found 273 proteins in all groups, and after data normalization 49 and 92 proteins were used for statistical analysis for comparisons between control versus rapamycin treatment groups, and metastasis versus primary tumor versus metastasis rapamycin versus primary tumor rapamycin, respectively. Considering the two statistical analysis, four proteins, phosphoglycerate mutase, malate dehydrogenase, l-lactate dehydrogenase and nucleolin were found in decreased abundance in the rapamycin group and they are related with cellular metabolic processes and enhanced tumor malignant behavior. Two proteins, dihydrolipoamide dehydrogenase and superoxide dismutase, also related with metabolic processes, were found in higher abundance in rapamycin group and are associated with apoptosis. The results suggested that rapamycin was able to inhibit cell growth of mammary gland tumor and metastatic tumors cells in vitro, however, concentrations needed to reach the IC50 were higher when compared to other studies.

Masquelet-Induced Membrane Characteristics in Chicken Radii Bone Defects.

This study evaluated the formation of a Masquelet-induced membrane created through the formation of segmental bone defects in the radii of 15 healthy domestic chickens. When the chickens were in a surgical plane of anesthesia, a 1.5-cm segmental bone defect was produced in the left radius, which was subsequently filled with a bone cement spacer during its pasty polymerization phase. The bone defects were evaluated through radiographic imaging immediately after surgery and at 7, 15, 21, and 30 days after the creation of the bone defect. Five birds were euthanatized at 15, 21, and 30 days postoperatively for histological evaluation of the bone defect site. Immediate postoperative radiographic examination of the radii showed the presence of bone cement, which occupied the segmental bone defect. Thirty days after the surgical procedure, the presence of new bone formation at the fractured extremities was evident in the 5 remaining chickens. Histologically, the induced-membrane had 3 distinct zones at 15 days postoperatively, including 1 cell layer in contact with the bone cement spacer, 1 layer with collagen fibers, and 1 layer in contact with muscle, which was composed of disorganized connective tissue, active fibroblasts, and blood vessels. Twenty-one days after surgery, the zones were less defined, and there were metaplastic areas comprising cartilage and bone. Postoperative, diffuse mineralization of the membrane was observed 30 days after the surgical procedure. Formation of the induced membrane was observed during all periods of evaluation. The best histological characteristics for the Masquelet-induced membrane were detected 15 days after the formation of the bone defect, suggesting this would be the optimal time for second-stage surgery for bone reconstruction.

Vitrification of cat ovarian tissue: Does fragment size matters?

The aim of the present study was to investigate whether or not the size of the ovarian fragment influences its resistance to cryostorage. For that purpose, ovaries were collected from 34 queens (various breeds, age 1-5 year) by routine ovariectomy, transported to the laboratory and then sectioned in different sizes (3 mm × 3 mm × 3 mm, 5 mm × 3 mm × 3 mm and 7 mm × 3 mm × 3 mm) and randomly assigned to a control (GC3, GC5 and GC7, respectively) or vitrified (GV3, GV5 and GV7, respectively) groups. Vitrified-warmed fragments were evaluated by histomorphology and immunohistochemistry (for apoptotic rates by using cleaved caspase-3). Histological examination reveals that 72.97% of the follicles in GV3 and 72.58% in GV5 were normal while only 42.86% of the follicles in GV7. The main morphological alteration presented in all groups was a detachment of the epithelial cells. Similarly, immunohistochemistry evaluation using caspase 3 revealed a small proportion of apoptotic cells in GV3 (8.43%) while in GV7 30.43% of the cells expressed cleaved caspase-3. These findings indicate that fragments sectioned in 3 mm × 3 mm × 3 mm (27 mm3 ) seem more adequate for perfusion of the cryoprotectant, causing less damage to the cell after vitrification-warming.

Intratesticular transplantation of allogenic mesenchymal stem cells mitigates testicular destruction after induced heat stress in Miniature-horse stallions.

Testicular degeneration (TD) is the most frequent cause of sub or infertility in stallions. Currently, mesenchymal stem cells (MSC) have been studied as a therapeutic option for several diseases including induced-TD in laboratory animals. Therefore, this study aimed to evaluate the effect of intratesticular MSC therapy on the testicular histology of stallions submitted to scrotal heat stress. Ten healthy Miniature-horse stallions were submitted to testicular heat stress induced by a heating wrap device (42-45°C). Afterward, the stallions were divided into two groups and treated seven days later. MSCs-treated stallions were treated with an intratesticular injection of 10 × 106 of MSCs diluted in 5 mL of PBS, whereas placebo-treated stallions had 5 mL of PBS intratesticular injected. All stallions had testicular biopsies collected seven days before and one- and 14-days post-heat stress and were castrated 30 days after testicular insult. Tissue sections were stained with H&E and evaluated for the tubular and luminal diameter, epithelial thickness, seminiferous tubules (STs) integrity, the number of spermatozoa in the STs, and the percent of abnormal STs. Significance was set at P≤0.05. In both groups, testicular heat stress damaged the STs (P<0.05). However, STs' parameters were improved in MSCs-treated stallions compared to placebo-treated stallions 30 days after the testicular insult (P<0.05). In conclusion, the results of the present study suggest that intratesticular MSC therapy provided a therapeutic advantage in rescuing acute TD in stallions. However, further studies are essential to evaluate the benefits of this therapy on semen parameters and stallions with idiopathic TD.

Review of the comparative pathological and immunohistochemical features of human and canine cutaneous melanocytic neoplasms.

Melanocytic neoplasms originate from melanocytes and melanoma, the malignant form, is a common canine neoplasm and the most aggressive human skin cancer. Despite many similarities between these neoplasms in both species, only a limited number of studies have approached these entities in a comparative manner. Therefore, this review compares benign and malignant melanocytic neoplasms in dogs and humans, exclusively those arising in the haired skin, with regard to their clinicopathological, immunohistochemical and molecular aspects. Shared features include spontaneous occurrence, macroscopic features and microscopic findings when comparing human skin melanoma in the advanced/invasive stage and canine cutaneous melanoma, immunohistochemical markers and several histopathological prognostic factors. Differences include the apparent absence of active mutations in the BRAF gene in canine cutaneous melanoma and less aggressive clinical behaviour in dogs than in humans. Further studies are required to elucidate the aetiology and genetic development pathways of canine cutaneous melanocytic neoplasms. Evaluation of the applicability of histopathological prognostic parameters commonly used in humans for dogs are also needed. The similarities between the species and the recent findings regarding genetic mutations in canine cutaneous melanomas suggest the potential utility of dogs as a natural model for human melanomas that are not related to ultraviolet radiation.

Canine primary ureteral leiomyosarcoma treated with unilateral left ureteronephrectomy.

Background: Primary ureteral neoplasms are extremely rare in dogs, and ureteral involvement usually occurs owing to the invasion of renal and bladder tumors. Case Description: This case report describes a 12-year-old intact male mixed-breed dog referred to a private clinic with a six-month history of abdominal distention. A physical examination revealed mild abdominal pain. Hematological tests detected normocytic-normochromic anemia (hematocrit 33.6% [reference interval-RI: 37%-55%], red blood cells 4.93 M/µl [RI: 5.5-8.5 M/µl], and hemoglobin 12.4 g/dl [RI: 12-18.0 g/dl]). The results from the leukogram, thrombogram, renal, and hepatic panels were within the reference intervals for dogs. Abdominal ultrasonography revealed a cavitary mass measuring approximately 12 cm in diameter as the largest tumor in the left abdominal region over the left hepatic lobe or mesenteric site. Chest radiography did not reveal any metastasis. Therefore, the patient underwent exploratory laparotomy, during which the left ureter was found to be affected by a 12-cm mass that adhered to the left kidney. A unilateral left ureteronephrectomy was performed, and histology and immunohistochemistry (IHC) confirmed well-differentiated primary ureteral leiomyosarcoma. The patient survived for 130 days but died of lung metastasis. Conclusion: Ureteral leiomyosarcoma should be investigated and included in the list of differential diagnoses for primary ureteral neoplasms. Regardless of the therapeutic modality, the prognosis of ureteral leiomyosarcoma may be unfavorable, as shown in this report.

Histological, Immunohistochemical, Biomechanical, and Wettability Evaluations of the Leukocyte- and Platelet-Rich Fibrin Membranes Derived from Canine Blood.

This study aimed to perform histological, immunohistochemical, biomechanical, and wettability assessments of leukocyte- and platelet-rich fibrin (L-PRF) membranes obtained from the blood of healthy dogs. Ten client-owned Labrador Retriever dogs were enrolled. Blood samples were obtained from the external jugular vein using a vacuum tube without anticoagulant, which was immediately centrifuged at 400g for 12 min in a dedicated centrifuge. The L-PRF clot was removed from the tube, and the red clot was released from the buffy coat using a spatula. The membrane was produced using a PRF box. Histological examination identified the three portions of the L-PRF membranes. The first portion was composed mainly of red blood cells with the presence of a low number of leukocytes among them. The second portion was composed of white blood cells, mainly neutrophils. The third portion was composed of the fibrin network which was characterized by acidophilic staining. The immunohistochemical analysis showed that vascular endothelial growth factor and platelet-derived growth factor were expressed in all samples at different intensities, both in cellular components and fibrin mesh. The tensile test and wettability assessments were measured in membranes 30 min and 3 h after production. The 30 min L-PRF membranes supported twice the ultimate tensile strength compared to 3 h L-PRF membranes. The wettability of the 30 min sample membranes was statistically higher than the 3 h sample membranes. In conclusion, the centrifugation protocol allowed production of the L-PRF membrane using canine blood and this was confirmed by histological and immunohistochemical analysis. The mechanical resistance and wettability of the L-PRF membrane were significantly reduced over time.

In Vivo Models to Evaluate Antitumor Drugs Effect on Vasculogenic Mimicry.

The vasculogenic mimicry (VM) in vivo evaluation is challenging, and new models have been proposed to evaluate antitumor effect of different compounds using in vivo models. However, there is no gold standard in vivo models established for VM evaluation. As occurs for other in vivo tumor analysis, the use of immunodeficient mouse model and cell line with in vivo tumorigenicity and ability to induce vasculogenic mimicry is the most used model.

Pathways Involved in the Development of Vasculogenic Mimicry in Canine Mammary Carcinoma Cell Cultures.

Vasculogenic mimicry (VM) is the ability of highly aggressive cancer cells to form fluid-conducting channels that facilitate the nutrition and metastasis of cancer cells. Considering the importance of VM in the prognosis of canine mammary gland tumours, this study aimed to investigate global gene expression in two canine mammary carcinoma cell cultures associated with the capacity for VM in vitro. The cell lines were subjected to an in-vitro assay to form VM channels (3D culture). Each cell line was then used in 2D conditions as controls and we compared the global gene expression with that of the 3D cultures. A total of 1,217 differentially expressed genes (DEGs) (P <0.05, fold change >2.0 or <2.0) were observed in 3D conditions compared with 2D culture in the UNESP-CM9 cell line, of which 677 were upregulated genes and 540 were downregulated. In contrast, the UNESP-CM60 cell line had only one upregulated and two downregulated genes. Overall, we identified several genes and pathways involved in the development of VM and these molecular data will be useful for future studies aimed at identifying diagnostic and therapeutic targets for VM in canine mammary carcinoma.

Structural and Ultrastructural Morphological Evaluation of Giant Anteater (Myrmecophaga tridactyla) Prostate Gland.

The giant anteater (Myrmecophaga tridactyla) is a vulnerable species from Central and South America, and is considered possibly extinct in Belize, Guatemala, El Salvador, and Uruguay. Due to the species' conservation and reproductive importance, this research aimed to characterize the morphology, histochemical, immunohistochemical, and ultrastructural feature of the giant anteater prostate gland. For this, we collected 11 giant anteater prostate glands and performed macroscopic, morphological, histochemical, immunohistochemical, and ultrastructural analysis. Nine prostate glands from an adult subject and two from young subjects were studied. Grossly, the adult giant anteater prostate gland is divided in two distinct zones; the central zones (composed mainly of ducts) and the peripheral zones (of acini formed by secretory cells). The secretory cells showed positive periodic acid-Schiff staining. Furthermore, the immunohistochemical characterization revealed a similar human prostate pattern, with p63 staining basal cells, uroplakin III (UPIII) superficial cells of prostatic urethra, androgen receptor (AR) expressing nucleus of secretory and stromal cells, and prostatic specific antigen (PSA) staining prostatic epithelial cells. Overall, our research provided an in-depth morphological description of the giant anteater's prostate gland, providing valuable information for futures studies focused on giant anteater conservation.

Primary Uterine Hemangiosarcoma in a Thoroughbred: A Case Report.

A 1.8-year-old maiden Thoroughbred filly, without previous history of mating or reproductive management, was referred for clinical inspection due to the presence of sanguineous vaginal discharge and severe abdominal pain. Transrectal palpation indicated uterine asymmetry, and transrectal ultrasonography revealed a mass near the cervix measuring 8.3 cm in diameter, with heterogeneous echogenicity, a trabeculated center, and a well-defined hyperechoic border. Smaller masses surrounded the larger uterine mass. During the examination, the mare expelled a uterine mass through the vulva. Histological and immunohistochemical (CD31 and Factor VIII) examinations of the expelled mass suggested a diagnosis of hemangiosarcoma. Therefore, a therapeutic hysterectomy was performed, and examinations of the uterine tissue confirmed the diagnosis. However, the mare was euthanized 2 weeks later due to postoperative complications. The animal was subjected to necropsy, and intestinal adhesions in the surgical incision were diagnosed as postoperative complications. No other neoplasms were found during necropsy, establishing the primary origin of the tumor. This case study presents the first known report of uterine hemangiosarcoma in an equine species.

Comparison of two different doses of bleomycin in electrochemotherapy protocols for feline cutaneous squamous cell carcinoma nonsegregated from ultraviolet light exposure.

Cutaneous squamous cell carcinoma (cSCC) is one of the most common skin tumors in cats due to chronic exposure to ultraviolet light. Local treatments such as electrochemotherapy (ECT) promote disease control or even complete remission. We hypothesize that cats could benefit from treatments using bleomycin at reduced dosages. A prospective nonrandomized single-blind study evaluated the clinical parameters, site lesion, staging, disease-free interval (DFI) and survival time by comparing the standard dose of bleomycin (15,000 UI/m2) (n = 22) with a reduced dose (10,000 UI/m2) (n = 34) in cats with cSCC that underwent ECT as the sole treatment modality. No statistically significant difference in DFI or overall survival was observed between the 2 groups. A higher DFI was found in cats with a small tumor size (less than 0.33 cm3) compared with that for cats with a large tumor size (P = 0.045). Furthermore, a reduced overall survival time for cats with a higher stage in the standard group SG (T3 and T4) (P = 0.004) was observed when compared to that for cats with a lower stage (T1 and T2). In conclusion, ECT using both doses of bleomycin may achieve the same response rate in terms of the overall response, DFI, and overall survival.

Transcriptome of Two Canine Prostate Cancer Cells Treated With Toceranib Phosphate Reveals Distinct Antitumor Profiles Associated With the PDGFR Pathway.

Canine prostate cancer (PC) presents a poor antitumor response, usually late diagnosis and prognosis. Toceranib phosphate (TP) is a nonspecific inhibitor of receptor tyrosine kinases (RTKs), including vascular endothelial growth factor receptor (VEGFR), platelet-derived growth factor receptor (PDGFR), and c-KIT. This study aimed to evaluate VEGFR2, PDGFR-ß, and c-KIT protein expression in two established canine PC cell lines (PC1 and PC2) and the transcriptome profile of the cells after treatment with TP. Immunofluorescence (IF) analysis revealed VEGFR2 and PDGFR-ß protein expression and the absence of c-KIT protein expression in both cell lines. After TP treatment, only the viability of PC1 cells decreased in a dose-dependent manner. Transcriptome and enrichment analyses of treated PC1 cells revealed 181 upregulated genes, which were related to decreased angiogenesis and cell proliferation. In addition, we found upregulated PDGFR-A, PDGFR-ß, and PDGF-D expression in PC1 cells, and the upregulation of PDGFR-ß was also observed in treated PC1 cells by qPCR. PC2 cells had fewer protein-protein interactions (PPIs), with 18 upregulated and 22 downregulated genes; the upregulated genes were involved in the regulation of parallel pathways and mechanisms related to proliferation, which could be associated with the resistance observed after treatment. The canine PC1 cell line but not the PC2 cell line showed decreased viability after treatment with TP, although both cell lines expressed PDGFR and VEGFR receptors. Further studies could explain the mechanism of resistance in PC2 cells and provide a basis for personalized treatment for dogs with PC.

Inflammatory Breast Cancer: Clinical Implications of Genomic Alterations and Mutational Profiling.

Inflammatory breast cancer (IBC) is a rare and aggressive type of breast cancer whose molecular basis is poorly understood. We performed a comprehensive molecular analysis of 24 IBC biopsies naïve of treatment, using a high-resolution microarray platform and targeted next-generation sequencing (105 cancer-related genes). The genes more frequently affected by gains were MYC (75%) and MDM4 (71%), while frequent losses encompassed TP53 (71%) and RB1 (58%). Increased MYC and MDM4 protein expression levels were detected in 18 cases. These genes have been related to IBC aggressiveness, and MDM4 is a potential therapeutic target in IBC. Functional enrichment analysis revealed genes associated with inflammatory regulation and immune response. High homologous recombination (HR) deficiency scores were detected in triple-negative and metastatic IBC cases. A high telomeric allelic imbalance score was found in patients having worse overall survival (OS). The mutational profiling was compared with non-IBC (TCGA, n = 250) and IBC (n = 118) from four datasets, validating our findings. Higher frequency of TP53 and BRCA2 variants were detected compared to non-IBC, while PIKC3A showed similar frequency. Variants in mismatch repair and HR genes were associated with worse OS. Our study provided a framework for improved diagnosis and therapeutic alternatives for this aggressive tumor type.

p-mTOR, p-4EBP-1 and eIF4E expression in canine prostatic carcinoma.

The mTOR/4E-BP1/eIF4E pathway plays important roles in the neoplastic transformation process and in tumour growth. In men, the mTOR/4E-BP1/eIF4E pathway was described as altered in different tumours, including prostate cancer (PC). Apart from humans, the dog is the only species that develops PC with high frequency and is considered a good model for comparative oncology initiatives. Due to limited information on this pathway in canine tumours, this study aimed to investigate mTOR, 4E-BP1 and eIF4E gene and protein expression in canine PC, as well as in metastatic and normal prostatic tissues, and to evaluate the correlations between gene/protein expression and Gleason score (GS) in PC. A total of 35 formalin-fixed paraffin-embedded (FFPE) samples, including 13 of normal prostatic tissue, 17 PC samples and 5 metastasis samples, were evaluated by immunohistochemistry and qPCR. mTOR gene mutation in the kinase domain was also investigated. We identified higher p-mTOR and eIF4E protein levels in canine PC with higher GS values (≥ 8) and a significant positive correlation in expression between these proteins. eIF4E overexpression was observed in metastasis relative to expression in normal samples. Our data suggest that p-mTOR and eIF4E expression is positively correlated with GS in canine PC, similar to the pattern in humans. More studies of the mTOR/4EBP1/eIF4E pathway should be performed to identify possible correlations of the proteins involved with clinical and pathologic findings in canine PC and the roles of these proteins as therapeutic targets for the treatment of canine PC.

Electrochemotherapy induces tumor regression and decreases the proliferative index in canine cutaneous squamous cell carcinoma.

Canine cutaneous squamous cell carcinoma (cSCC) is the most common skin cancer in dogs, and, due to its low metastatic rate, local treatments, such as electrochemotherapy (ECT), promote disease control or even complete remission (CR). This study aimed to evaluate the gene and protein expression of Bcl-2 and Bcl-2 associated X protein (BAX), the proliferative index and clinical parameters in dogs with cSCC subjected to ECT. A prospective nonrandomized clinical study was performed using dogs with naturally occurring cSCC that was treated with ECT. Eighteen lesions from 11 dogs were selected. The tumor size at day 0 (D0) had no impact on survival or prognosis (P > 0.05). Tumor samples had a lower proliferative index after ECT (D21) than before ECT (P = 0.031). The survival of subjects with Ki67 values lower and higher than the Ki67 median value were not significantly different (P > 0.05). Regarding apoptotic markers, there were no significant differences in the gene and protein expression levels of BAX or Bcl-2 at D0 and D21 (P > 0.05) or in the overall survival of subjects with different levels of apoptotic markers. In conclusion, there was no change in BAX or Bcl-2 gene and protein expression in response to ECT at the time points evaluated, but ECT was able to reduce tumor volume and cellular proliferation in cSCC.

Deregulation of E-cadherin, ß-catenin, APC and Caveolin-1 expression occurs in canine prostate cancer and metastatic processes.

Prostate cancer is a heterogeneous disease with high levels of clinical and gene heterogeneity, consequently offering several targets for therapy. Dogs with naturally occurring prostate cancer are useful models for molecular investigations and studying new treatment efficacy. Three genes and proteins associated with the WNT pathway (ß-catenin, APC and E-cadherin) and Caveolin-1 (CAV-1) were evaluated in canine pre-neoplastic proliferative inflammatory atrophy (PIA), prostate cancer and metastatic disease. The APC gene methylation status was also investigated. As in human prostate cancer, cytoplasmic and nuclear ß-catenin, which are fundamental for activating the canonical WNT pathway, were found in canine prostate cancer and metastasis. Membranous E-cadherin was also lost in these lesions, allowing cellular migration to the stroma and nuclear localization of ß-catenin. In contrast to human prostate tumours, no APC downregulation or hypermethylation was found in canine prostate cancer. The CAV-1 gene and protein overexpression were found in canine prostate cancer, and as in humans, the highest levels were found in Gleason scores ≥8. In conclusion, as with human prostate cancer, ß-catenin and E-cadherin in the WNT pathway, as well as Caveolin-1, are molecular drivers in canine prostate cancer. These findings provide additional evidence that dogs are useful models for studying new therapeutic targets in prostate cancer.

Pilot assessment of vascular endothelial growth factor receptors and trafficking pathways in recurrent and metastatic canine subcutaneous mast cell tumours.

Canine subcutaneous mast cell tumour (scMCT) shows less aggressive biological behaviour than cutaneous MCT. Vascular endothelial growth factor receptor 2 (VEGFR2) is expressed by neoplastic cells in canine scMCT, but the relevance of this signalling pathway for disease pathobiology is not clear. The objective of this study was to quantify VEGF-A, VEGFR2, pVEGFR2, the VEGF co-receptor Neuropilin 1 (NRP-1) and the E3 ubiquitin protein ligase c-Cbl in canine scMCT, and to evaluate their association with disease outcome. Immunohistochemical staining for biomarkers was quantified from 14 cases of canine scMCT using manual and computer-assisted methods. Kaplan-Meier curves were generated for disease-free survival (DFS) and compared using Mantel-Cox log-rank analysis. Cases with high levels of neoplastic cell VEGFR2, pVEGFR2 or c-CBL immunoreactivity had significantly reduced DFS. All cases displayed neoplastic cells positive for VEGF-A, which was significantly associated with pVEGFR2 immunoreactivity. There were also significant positive correlations between VEGFR2 and pVEGFR2, and between c-CBL and pVEGFR2 levels. This pilot study demonstrates the potential utility of these markers in a subset of scMCT in dogs.