Phosphoproteomics analysis of serum from dogs affected with pulmonary hypertension secondary to degenerative mitral valve disease.

Pulmonary hypertension (PH), a common complication in dogs affected by degenerative mitral valve disease (DMVD), is a progressive disorder characterized by increased pulmonary arterial pressure (PAP) and pulmonary vascular remodeling. Phosphorylation of proteins, impacting vascular function and cell proliferation, might play a role in the development and progression of PH. Unlike gene or protein studies, phosphoproteomic focuses on active proteins that function as end-target proteins within signaling cascades. Studying phosphorylated proteins can reveal active contributors to PH development. Early diagnosis of PH is crucial for effective management and improved clinical outcomes. This study aimed to identify potential serum biomarkers for diagnosing PH in dogs affected with DMVD using a phosphoproteomic approach. Serum samples were collected from healthy control dogs (n = 28), dogs with DMVD (n = 24), and dogs with DMVD and PH (n = 29). Phosphoproteins were enriched from the serum samples and analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Data analysis was performed to identify uniquely expressed phosphoproteins in each group and differentially expressed phosphoproteins among groups. Phosphoproteomic analysis revealed nine uniquely expressed phosphoproteins in the serum of dogs in the DMVD+PH group and 15 differentially upregulated phosphoproteins in the DMVD+PH group compared to the DMVD group. The phosphoproteins previously implicated in PH and associated with pulmonary arterial remodeling, including small nuclear ribonucleoprotein G (SNRPG), alpha-2-macroglobulin (A2M), zinc finger and BTB domain containing 42 (ZBTB42), hemopexin (HPX), serotransferrin (TRF) and complement C3 (C3), were focused on. Their unique expression and differential upregulation in the serum of DMVD dogs with PH suggest their potential as biomarkers for PH diagnosis. In conclusion, this phosphoproteomic study identified uniquely expressed and differentially upregulated phosphoproteins in the serum of DMVD dogs with PH. Further studies are warranted to validate the diagnostic utility of these phosphoproteins.

Hematological and blood biochemistry parameters as prognostic indicators of survival in canine multicentric lymphoma treated with COP and L-COP protocols.

Background and Aim: Hematological and blood chemistry parameters are crucial for evaluating and monitoring canine multicentric lymphoma during chemotherapy. Pre-treatment hematological and blood chemistry parameters can be used as prognostic survival outcomes for this disease. Therefore, this study aimed to investigate the effect of hematological and blood chemistry parameters pre-treatment and 4 weeks post-treatment on the survival outcomes of dogs treated with either a combination of cyclophosphamide, vincristine, and prednisolone (COP) or a combination of COP with L-asparaginase (L-COP) protocols. Materials and Methods: We conducted a retrospective study. Medical records and hematological and blood chemistry parameters of 41 dogs with multicentric lymphoma treated with L-COP (n = 26) and the COP protocols (n = 15) were obtained from the hospital information system. Most cases were classified as high-grade lymphoma based on the Kiel cytological classification. The effects of hematological and blood chemistry parameters on survival outcomes were investigated using the Cox proportional hazard regression model. The median survival time (MST) for each hematological and blood chemistry parameter affecting survival outcome was established and compared using the Kaplan-Meier product limit method with the log-rank test. Results: Dogs with high-grade multicentric lymphoma that were treated with the COP protocol and had monocytosis at pre-treatment had a significantly shorter MST than dogs with normal monocyte counts (p = 0.033). In addition, dogs with azotemia, both pre-treatment and 4 weeks post-treatment, had a significantly shorter MST than dogs with normal serum creatinine levels (p = 0.012). Dogs with high-grade multicentric lymphoma treated with the L-COP protocol who had hypoalbuminemia (serum albumin concentration <2.5 mg/dL) at both pre-treatment and 4 weeks post-treatment had a significantly shorter MST than dogs with normal serum albumin levels (p < 0.001). Furthermore, dogs with leukocytosis at 4 weeks post-treatment had a significantly shorter MST than those with a normal total white blood cell count (p = 0.024). Conclusion: Serum albumin level can serve as a simple negative prognostic indicator of survival outcomes in dogs with high-grade multicentric lymphoma treated with the L-COP protocol. Dogs with hypoalbuminemia pre-treatment and 4 weeks post-treatment tended to have a shorter MST than those with normal serum albumin concentrations.

Proteomic analysis of the serum in dogs with pulmonary hypertension secondary to myxomatous mitral valve disease: the preliminary study.

Background: Pulmonary hypertension (PH) is a common complication in dogs with myxomatous mitral valve disease (MMVD), characterized by elevated blood pressure in pulmonary artery. Echocardiography is a reliable technique for PH diagnosis in veterinary medicine. However, it is limited to use as an early detection method. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) has found extensive application in the discovery of serum protein biomarkers for various diseases. The objective of this study was to identify serum proteins in healthy control dogs and MMVD dogs both with and without PH using LC-MS/MS. Materials and methods: In this research, a total of 81 small-breed dogs participated, and they were categorized into three groups: the control (n = 28), MMVD (n = 24) and MMVD+PH (n = 29) groups. Serum samples were collected and analyzed by LC-MS/MS. Results: Differentially expressed proteins were identified, and the upregulated and downregulated proteins in MMVD+PH group including Myomesin 1 (MYOM1) and Histone deacetylase 7 (HDAC7), Pleckstrin homology domain containing M3 (PLEKHM3), Diacylglycerol lipase alpha (DAGLA) and Tubulin tyrosine ligase like 6 (TTLL6) were selected as proteins of interest in MMVD dogs with PH. Conclusion: Different types of proteins have been identified in healthy dogs and MMVD dogs with and without PH. Additional studies are needed to investigate the potential of these proteins as biomarkers for PH in dogs with MMVD.

Anti-proliferative and apoptotic effect of cannabinoids on human pancreatic ductal adenocarcinoma xenograft in BALB/c nude mice model.

Human pancreatic ductal adenocarcinoma (PDAC) is a highly malignant and lethal tumor of the exocrine pancreas. Cannabinoids extracted from the hemp plant Cannabis sativa have been suggested as a potential therapeutic agent in several human tumors. However, the anti-tumor effect of cannabinoids on human PDAC is not entirely clarified. In this study, the anti-proliferative and apoptotic effect of cannabinoid solution (THC:CBD at 1:6) at a dose of 1, 5, and 10 mg/kg body weight compared to the negative control (sesame oil) and positive control (5-fluorouracil) was investigated in human PDAC xenograft nude mice model. The findings showed that cannabinoids significantly decreased the mitotic cells and mitotic/apoptotic ratio, meanwhile dramatically increased the apoptotic cells. Parallelly, cannabinoids significantly downregulated Ki-67 and PCNA expression levels. Interestingly, cannabinoids upregulated BAX, BAX/BCL-2 ratio, and Caspase-3, meanwhile, downregulated BCL-2 expression level and could not change Caspase-8 expression level. These findings suggest that cannabinoid solution (THC:CBD at 1:6) could inhibit proliferation and induce apoptosis in human PDAC xenograft models. Cannabinoids, including THC:CBD, should be further studied for use as the potent PDCA therapeutic agent in humans.

Domestic Cat Hepadnavirus Antigens in Lymphoma Tissues. Comment on Beatty et al. Domestic Cat Hepadnavirus and Lymphoma. Viruses 2023, 15, 2294.

We are addressing the comments made by Beatty et al [...].

Proteomic analysis of pulmonary arteries and lung tissues from dogs affected with pulmonary hypertension secondary to degenerative mitral valve disease.

In dogs with degenerative mitral valve disease (DMVD), pulmonary hypertension (PH) is a common complication characterized by abnormally elevated pulmonary arterial pressure (PAP). Pulmonary arterial remodeling is the histopathological changes of pulmonary artery that has been recognized in PH. The underlying mechanisms that cause this arterial remodeling are poorly understood. This study aimed to perform shotgun proteomics to investigate changes in protein expression in pulmonary arteries and lung tissues of DMVD dogs with PH compared to normal control dogs and DMVD dogs without PH. Tissue samples were collected from the carcasses of 22 small-sized breed dogs and divided into three groups: control (n = 7), DMVD (n = 7) and DMVD+PH groups (n = 8). Differentially expressed proteins were identified, and top three upregulated and downregulated proteins in the pulmonary arteries of DMVD dogs with PH including SIK family kinase 3 (SIK3), Collagen type I alpha 1 chain (COL1A1), Transforming growth factor alpha (TGF-α), Apoptosis associated tyrosine kinase (AATYK), Hepatocyte growth factor activator (HGFA) and Tyrosine-protein phosphatase non-receptor type 13 (PTPN13) were chosen. Results showed that some of the identified proteins may play a role in the pathogenesis of pulmonary arterial remodeling. This study concluded shotgun proteomics has potential as a tool for exploring candidate proteins associated with the pathogenesis of PH secondary to DMVD in dogs.

Comparative E-cadherin and syndecan-1 protein expression in human and canine oral squamous cell carcinoma.

Oral squamous cell carcinoma (OSCC) is a prevalent form of oral cancer in humans and dogs. The altered expression of cell adhesion molecules, including E-cadherin (CDH1) and syndecan-1 (SDC1), is involved in cancer progression. This study aimed to investigate the protein expression of CDH1 and SDC1 in early and late clinical stages of human and canine OSCC (hOSCC and cOSCC, respectively), using immunohistochemistry. Formalin-fixed and paraffin embedded tissue blocks were obtained from 21 hOSCC, 8 human normal gingiva, 26 cOSCC, and 13 canine normal gingiva. Clinical stages and histological subtypes of samples were evaluated. The results indicated that both human and canine OSCC exhibited reduced levels of CDH1 and SDC1 expression at the cell membrane regardless of clinical stage or histological subtype. Additionally, decreased levels of total SDC1 expression were observed in hOSCC compared with normal controls. In conclusion, this study demonstrates a similarity in the immunohistochemical expression of CDH1 and SDC1 between humans and dogs with OSCC, lending support to the potential use of dogs as a model for studying human head and neck squamous cell carcinoma.

High-resolution melting analysis for simultaneous detection and discrimination between wild-type and vaccine strains of feline calicivirus.

High-resolution melting (HRM) analysis, a post-polymerase chain reaction (PCR) application in a single closed tube, is the straightforward method for simultaneous detection, genotyping, and mutation scanning, enabling more significant dynamic detection and sequencing-free turnaround time. This study aimed to establish a combined reverse-transcription quantitative PCR and HRM (RT-qPCR-HRM) assay for diagnosing and genotyping feline calicivirus (FCV). This developed method was validated with constructed FCV plasmids, clinical swab samples from living cats, fresh-frozen lung tissues from necropsied cats, and four available FCV vaccines. We performed RT-qPCR to amplify a 99-base pair sequence, targeting a segment between open reading frame (ORF) 1 and ORF2. Subsequently, the HRM assay was promptly applied using Rotor-Gene Q® Software. The results significantly revealed simultaneous detection and genetic discrimination between commercially available FCV vaccine strains, wild-type Thai FCV strains, and VS-FCV strains within a single PCR reaction. There was no cross-reactivity with other feline common viruses, including feline herpesvirus-1, feline coronavirus, feline leukemia virus, feline immunodeficiency virus, and feline morbillivirus. The detection limit of the assay was 6.18 × 101 copies/µl. This study, therefore, is the first demonstration of the uses and benefits of the RT-qPCR-HRM assay for FCV detection and strain differentiation in naturally infected cats.

Domestic cat hepadnavirus detection in blood and tissue samples of cats with lymphoma.

Domestic cat hepadnavirus (DCH), a relative hepatitis B virus (HBV) in human, has been recently identified in cats; however, association of DCH infection with lymphoma in cats is not investigated. To determine the association between DCH infection and feline lymphoma, seven hundred and seventeen cats included 131 cats with lymphoma (68 blood and 63 tumor samples) and 586 (526 blood and 60 lymph node samples) cats without lymphoma. DCH DNA was investigated in blood and formalin-fixed paraffin-embedded (FFPE) tissues by quantitative polymerase chain reaction (qPCR). The FFPE lymphoma tissues were immunohistochemically subtyped, and the localization of DCH in lymphoma sections was investigated using in situ hybridization (ISH). Feline retroviral infection was investigated in the DCH-positive cases. DCH DNA was detected in 16.18% (11/68) (p = 0.002; odds ratio [OR], 5.15; 95% confidence interval [CI], 2.33-11.36) of blood and 9.52% (6/63) (p = 0.028; OR, 13.68; 95% CI, 0.75-248.36) of neoplastic samples obtained from lymphoma cats, whereas only 3.61% (19/526) of blood obtained from non-lymphoma cats was positive for DCH detection. Within the DCH-positive lymphoma, in 3/6 cats, feline leukemia virus was co-detected, and in 6/6 were B-cell lymphoma (p > 0.9; OR, 1.93; 95% CI, 0.09-37.89) and were multicentric form (p = 0.008; OR, 1.327; 95% CI, 0.06-31.18). DCH was found in the CD79-positive pleomorphic cells. Cats with lymphoma were more likely to be positive for DCH than cats without lymphoma, and infection associated with lymphoma development needs further investigations.

Development and characterization of mouse anti-canine PD-L1 monoclonal antibodies and their expression in canine tumors by immunohistochemistry in vitro.

Immune escape is the hallmark of carcinogenesis. This widely known mechanism is the overexpression of immune checkpoint ligands, such as programmed cell death protein 1 and programmed death-ligand 1 (PD-1/PD-L1), leading to T cell anergy. Therefore, cancer immunotherapy with specific binding to these receptors has been developed to treat human cancers. Due to the lack of cross-reactivity of these antibodies in dogs, a specific canine PD-1/PD-L1 antibody is required. The aim of this study is to develop mouse anti-canine PD-L1 (cPD-L1) monoclonal antibodies and characterize their in vitro properties. Six mice were immunized with recombinant cPD-L1 with a fusion of human Fc tag. The hybridoma clones that successfully generated anti-cPD-L1 antibodies and had neutralizing activity were selected for monoclonal antibody production. Antibody properties were tested by immunosorbent assay, surface plasmon resonance, and immunohistochemistry. Four hybridomas were effectively bound and blocked to recombinant cPD-L1 and cPD-1-His-protein, respectively. Candidate mouse monoclonal antibodies worked efficiently on formalin-fixed paraffin-embedded tissues of canine cancers, including cutaneous T-cell lymphomas, mammary carcinomas, soft tissue sarcomas, squamous cell carcinomas, and malignant melanomas. However, functional assays of these anti-cPD-L1 antibodies need further investigation to prove their abilities as therapeutic drugs in dogs as well as their applications as prognostic markers.

Isolation of Oral Bacteria, Measurement of the C-Reactive Protein, and Blood Clinical Parameters in Dogs with Oral Tumor.

Canine oral cancers have a poor prognosis and are related to chronic inflammation. This may pose a risk of secondary bacterial infection. This study aimed to compare the bacteria isolated from oral swab samples, values of C-reactive proteins (CRPs), and clinical blood profiles of dogs with and without oral mass. A total of 36 dogs were divided in three groups: no oral mass (n = 21), oral mass (n = 8), and metastasis groups (n = 7). Significantly, both the clinical groups (the oral mass group and metastasis group) showed anemia, a decrease in the albumin-to-globulin ratio (AGR), and an increase in the neutrophil-to-lymphocyte ratio (NLR), globulin-to-albumin ratio (GAR), CRP, and CRP-to-albumin ratio (CAR) compared to the normal group. CAR showed an increasing trend in the oral mass and metastasis groups (10 times and 100 times, respectively) compared to the no oral mass group (P < 0.001). Neisseria spp. (20.78%) was the main isolated bacteria in all groups. The main genera in the no oral mass group were Neisseria spp. (28.26%), Pasteurella spp. (19.57%), and Staphylococcus spp. (19.57%). Neisseria spp., Staphylococcus spp., Klebsiella spp., and Escherichia spp. were found equally (12.5%) in the oral mass group. Escherichia spp. (26.67%), Pseudomonas spp. (13.33%), and Staphylococcus spp. (13.33%) were the main genera in the metastasis group. Interestingly, Neisseria spp. decreased in the clinical groups (Fisher's exact = 6.39, P=0.048), and Escherichia spp. increased in the metastasis group (Fisher's exact = 14.00, P=0.002). The difference of oral bacteria in clinical dogs compared to healthy dogs may be related to microbiome alterations, and both the clinical groups showed the increment of inflammatory biomarkers. This suggested that further studies should be conducted on the correlation between the specific bacteria, CRP, blood clinical parameters, and type of canine oral mass.

Perivascular inflammatory cells and their association with pulmonary arterial remodelling in dogs with pulmonary hypertension due to myxomatous mitral valve disease.

Pulmonary hypertension (PH), an increase in pulmonary arterial pressure (PAP), may occur in dogs affected with myxomatous mitral valve disease (MMVD). Recent studies suggest that an accumulation of perivascular inflammatory cells may be involved with medial thickening which is a sign of the pulmonary artery remodelling in PH. The aim of this study was to characterise perivascular inflammatory cells in the surrounding pulmonary arteries of dogs with PH due to MMVD compared to MMVD dogs and healthy control dogs. Nineteen lung samples were collected from cadavers of small-breed dogs (control n = 5; MMVD n = 7; MMVD + PH n = 7). Toluidine blue stain and multiple IHC targeting α-SMA, vWF, CD20, CD68 and CD3 was performed to examine intimal and medial thickening, assess muscularisation of the small pulmonary arteries and characterise perivascular leucocytes. Medial thickening without intimal thickening of pulmonary arteries and muscularisation of normally non-muscularised small pulmonary arteries was observed in the MMVD and MMVD + PH groups compared with the control group. The perivascular numbers of B lymphocytes, T lymphocytes and macrophages was significantly increased in the MMVD + PH group compared with the MMVD and control groups. In contrast, the perivascular number of mast cells was significantly higher in the MMVD group compared with the MMVD + PH and control groups. This study suggested that pulmonary artery remodelling as medial thickening and muscularisation of the normally non-muscular small pulmonary arteries is accompanied by the accumulation of perivascular inflammatory cells.

Case report: Mature extragonadal teratoma at the proximal part of the tail in a kitten.

An 8-month-old, intact male, domestic shorthair cat was referred for a mass on the proximal ventral part of the tail which had been found since the animal was born, and due to the presence of a linear fissure with rows of ectopic teeth, the veterinarian suspected that the mass had recently ruptured. Tail amputation was elected and the entire mass was successfully surgically excised. From the gross examination, this mass had an open cyst-like structure with a prominent area composed of hair, teeth, and bone. Histopathology revealed two components of germinal layers including hair follicles, adnexal tissue, neural tissue, teeth, muscle, fat, bone, and lymphatic vessels. The histopathological diagnosis was consistent to mature teratoma. Although, complete excision could not be definitively confirmed histologically, this kitten is currently well and has not developed any recurrent mass at the surgical site after 2 years of post-operation.

Diagnostic utility of LDH measurement for determining the etiology of modified transudate pleural effusion in cats.

Fluid analysis is an initial approach for determining the underlying causes of body cavity effusions. Modified transudate is commonly diagnosed in pleural effusion in cats, however, it provides limited diagnostic information. Aims of this study were to investigate common etiologies causing different pleural fluid types and to evaluate the usefulness of lactate dehydrogenase (LDH) for differentiating the etiology in modified transudates in cats. Pleural effusion samples from 122 cats were analyzed and classified into three types: transudate, modified transudate, and exudate. Causes of pleural effusion were classified into four conditions: cardiac disease, neoplasia, feline infectious peritonitis (FIP), and pyothorax. The relationship of underlying etiology and fluid types was described. The LDH levels in pleural fluid and plasma were compared between the causes in the samples classified as modified transudate. The fluid analysis of pleural effusion showed that modified transudate was the most common fluid type (44.2%). Neoplasia was predominantly diagnosed (38.5%) as the etiology of pleural effusion. There was no significant correlation between pleural fluid and plasma LDH level in any type of pleural fluid, suggesting that pleural fluid LDH does not appear to be affected by plasma LDH. The occurrence of modified transudate was not associated to its etiologies, however, the LDH level in modified transudates showed significant differences between etiologic groups. The LDH level in modified transudate was excellent in separating cardiac from non-cardiac diseases with a cut-off value of <535 U/L and separating FIP from non-FIP diseases with a cut-off value of >641 U/L. Based on the current findings, pleural fluid LDH can be a useful adjunctive marker for differentiating some causes of modified transudate pleural effusion and should be added in the routine diagnostic work-up of feline patients with pleural effusions.

Case report: BCL-2 and CD31 immunoexpression related to clinical and histopathological evaluation of renal dysplasia in a Welsh Corgi Puppy.

A case of renal dysplasia (RD) in the Welsh Corgi dog has been reported. Clinically, the affected 3-month-old, female, Welsh Corgi dog showed unclear symptoms of chronic kidney disease. Grossly, both left and right kidneys revealed cystic hypoplasia. Histologically, the primary lesions included immature or fetal glomeruli/tubules, proliferative arterioles, persistent metanephric ducts, persistent mesenchyme, and atypical tubular epithelium were presented. A group of degenerative and inflammatory lesions consisting of interstitial nephritis, interstitial fibrosis, and mineralization of tubules were found. Immunohistochemically, the epithelial cells of immature (fetal) tubules had BCL-2 labeling whereas CD31 (PECAM-1) was labeled in the endothelial cells of the proliferative arterioles. The immunohistochemical findings were confirmed and consolidated with the routine histopathological findings. This study was the first demonstration of the clinical, histopathological, and immunohistochemical features of RD disease in a Welsh Corgi puppy.

Investigation of Genes and Proteins Expression Associating Serotonin Signaling Pathway in Lung and Pulmonary Artery Tissues of Dogs with Pulmonary Hypertension Secondary to Degenerative Mitral Valve Disease: The Preliminary Study.

Pulmonary hypertension (PH) is defined as an increase in pulmonary vascular pressure. It is one of the most common complications that occur as a result of degenerative mitral valve disease (DMVD) in dogs. Serotonin (5-HT) can trigger the development of PH. Accordingly, this study investigated the changes in the expression of genes and proteins associated with local 5-HT signaling in the lungs and pulmonary arteries (PA) of dogs with PH secondary to DMVD. Lung and PA tissue samples were collected from the cadavers of fourteen small-breed dogs and divided into normal (n = 4), DMVD (n = 5) and DMVD with PH (n = 5) groups. Gene expression (tph1, slc6a4 and htr2a) was analyzed by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The expression of proteins (TPH-1, SERT, 5-HTR2A, ERK1/2 and pERK1/2) was examined by Western blot analysis and immunohistochemical staining. The results showed that the expression of genes and proteins evaluated by qRT-PCR and Western blot analysis in lung and PA tissues did not differ among groups. However, the expression of proteins related to 5-HT signaling tended to be upregulated in PA tissues from DMVD dogs with and without PH. Immunohistochemical examination revealed the overexpression of these proteins in the DMVD and DMVD with PH groups in lung tissue. These findings suggest a local effect of 5-HT signaling in DMVD dogs with and without PH.

Activating Mutation in the Receptor Tyrosine Kinase FLT3 with Clinicopathological Relevance in Canine Mast Cell Tumors.

Recent research has focused on the receptor tyrosine kinase (RTK) KIT which is involved in the pathogenesis of canine mast cell tumors (MCT). However, the role of other RTKs in this neoplasm remains unclear. The present study aimed to determine the frequency of FLT3 mutations and to evaluate the mutational status and clinicopathological relevance of canine MCT patients. There were a total of 20 cases that were cytologically and histopathological diagnosed as canine MCTs; genomic polymerase chain reaction (PCR) and Sanger sequencing were used to identify mutations. For the juxtamembrane (JM) domain, the FLT3 14/15 primer pair was used to investigate exon 14/15 loci. Based on genomic PCR amplification of exon 14/15 and 20 of the FLT3 gene and Sanger sequencing of 20 cases of canine MCTs, the overall frequency of FLT3 mutation in canine MCTs was 75%. The majority of FLT3 mutations (70%) were internal tandem duplications (ITD) of the JM domain, while one case arose from deletion mutations of the tyrosine kinase domain (TKD). However, double mutations were not observed in this study. Furthermore, there is also clinicopathological relevance to MCT dogs carrying FLT3-ITD mutations, showing a tendency toward leukocytosis due to neutrophilia, and resembling human acute myeloid leukemia (AML) with FLT3-ITD mutations. A subset of MCTs with FLT3-ITD mutations, showing an enhanced signal of phosphorylated ERK1/2 identified by immunoblotting, suggests that an activating mutation may be driven by a distinct signal of the ERK pathway. Our results indicate that FLT3-ITD mutation is an oncogenic driver of canine MCTs, and that it shares some clinicopathologic features with human AML. These findings may offer new opportunities for further studies on canine mast cell tumorigenesis and a novel therapeutic target for canine MCT cases harboring FLT3-ITD mutations.

Investigation of proteomic profiles in canine lymphoma using tandem mass tag-based quantitative proteomics approach.

Background and Aim: Specific tumor biomarkers are useful for the early diagnosis of cancer or can predict the recurrence of neoplastic disease in humans and animals. Lymphoma in dogs could be classified into B-, T-, and NK-cell origins. T-cell lymphoma has the worst prognosis with a shorter survival time and disease-free interval. This study aimed to identify the differential serum protein expressions of canine B- and T-cell lymphomas compared with healthy dogs using a tandem mass tag (TMT)-based quantitative proteomics. Materials and Methods: Serum samples were collected from 20 untreated canine lymphomas (14 B-cells and 6 T-cells) and four healthy control dogs. Sera peptides from each sample were processed for TMT 10-plex tagging and analyzed using liquid chromatography-mass spectrometry (MS). Differential proteome profiling was then compared between lymphoma and control. Results: We discovered 20 elevated and 14 decreased serum proteins in the lymphoma group relative to the healthy group. Six candidate increased proteins in canine lymphomas were beta-actin cytoplasmic 1 (ACTB, p=0.04), haptoglobin (p=0.002), beta-2 microglobulin (aaaaaaaa2M, p=0.007), beta-2 glycoprotein 1 (APOH, p=0.03), metalloproteinase inhibitor 1 (TIMP-1, p=0.03), and CD44 antigen (p=0.02). When compared between B- and T-cell lymphomas, B-cell phenotypes had upregulated immunoglobulin (Ig) heavy chain V region GOM (p=0.02), clusterin (p=0.01), apolipoprotein C1 (APOC1, p=0.05), and plasminogen (p=0.02). Conclusion: These findings were investigated quantitative serum proteomes between B- and T-cell lymphomas using TMT-based MS. ACTB, aaaaaaaa2M, APOH, TIMP-1, CD44 antigen, Ig heavy chain V region GOM, and APOC1 are novel candidate proteins and might serve as a lymphoma biomarker in dogs. However, evaluation with an increased sample size is needed to confirm their diagnostic and prognostic ability.

Comparative compositional and functional venomic profiles among venom specimens from juvenile, subadult and adult Russell's viper ( Daboia siamensis ): correlation with renal pathophysiology in experimental rabbits.

Background: Eastern Russell's viper (Daboia siamensis) is one of the most medically significant snakes responsible for the development of acute renal failure. However, variation of the clinical picture and renal pathophysiology following bites by young and adult D. siamensis have not been elucidated. Methods: In this study, we analyzed the venomic profiles of D. siamensis at different maturation stages of juvenile, subadult and adult groups. The same pooled venom from each group was subjected to enzymatic, electrophoretic and proteomic analysis, including sublethal toxicity (0.1 mg/kg iv.) examined on bodily functions by comparing the venom compositional and functional profiles among venom specimens from juvenile, subadult and adult D. siamensis by correlating them with the renal pathophysiology in experimental rabbits. Results: The comparative studies revealed that juvenile venom possessed higher phospholipase A2, metalloproteinase and serine proteinase levels, while subadult and adult venoms contained more L-amino acid oxidase, phosphodiesterase, the Kunitz-type serine protease inhibitor, disintegrin families and endothelial growth factor. An in vivo study revealed that the adult and subadult venoms caused persistent hypotension and bradycardia, while thrombocytopenia was a more characteristic effect of juvenile venom. All venom age groups showed significant reductions in renal hemodynamics and electrolyte excretions. The juvenile venom caused a higher tubulonephrosis lesion score than adult and subadult venoms. Conclusions: The D. siamensis venom shows an ontogenetic shift in its compositions and activities. Renal function alterations after envenomation depend on either the synergistic actions of different venom components or the disproportionate expression between the concentrations of enzymatic and non-enzymatic proteins in each age venom group. The high proportion of enzymatic toxin proteins in the juvenile venom results in greater nephrotoxicity.

Expression of apoptotic proteins in the pulmonary artery of dogs with pulmonary hypertension secondary to degenerative mitral valve disease.

Degenerative mitral valve disease (DMVD) is a common cause of pulmonary hypertension (PH) in dogs. Medial thickening of the pulmonary artery is a common histopathological change in PH. The cause of medial thickening could be a decrease in apoptosis of pulmonary arterial smooth muscle cells (SMCs). This study aimed to detect the expression of apoptosis-related proteins in the pulmonary artery of dogs with PH secondary to DMVD (DMVD+PH) compared with DMVD without PH (DMVD) and healthy dogs (control). Lung samples were collected from three groups, including the control (n = 5), DMVD (n = 7) and DMVD+PH (n = 7) groups. Masson trichrome, α-SMA and apoptotic proteins including Bax, Bcl2 and caspase-3 and -8 were stained. The results showed that the percentage of medial thickness (%MT) and the average number of pulmonary arterial SMCs were greater in the DMVD and DMVD+PH groups than in the control group, and they were greatest in the DMVD+PH group. Bax, Bcl2, and caspase-3 and -8 were mainly expressed in the medial layer of the pulmonary artery. The percentage of Bax and caspase-3 and -8 positive cells was higher in the DMVD group than in the DMVD+PH group, whereas the percentage of Bcl2 positive cells was increased in the DMVD+PH group. These results suggested that the pro-apoptotic activity of pulmonary arterial SMCs occurred mainly in the DMVD group and decreased dramatically in the DMVD+PH group. In conclusion, an increase in medial thickness in dogs with PH secondary to DMVD may relate to a decrease in pro-apoptotic activity of pulmonary arterial SMCs.