Case Report: Aortic Regurgitation of Postocclusion and Long-Term Outcome Following PDA Correction in an Adult Dog.

A 9-year-old intact female Maltese dog was admitted for further evaluation of previously diagnosed patent ductus arteriosus (PDA). The dog showed severe coughing and exercise intolerance. On physical examination, a grade VI/VI continuous heart murmur was auscultated. Thoracic radiography demonstrated cardiomegaly, pulmonary overcirculation, and moderate bronchointerstitial pattern. Echocardiography revealed severe dilation of the left ventricle and atrium, decreased left ventricular contractility, and left-to-right PDA. On electrocardiography (ECG), R amplitude was increased. Computed tomographic angiography revealed type IIA PDA. The serum N-terminal pro-B-type natriuretic peptide (NT-proBNP) concentration was >10,000 pmol/L. Transarterial occlusion was performed and the Amplatz® Canine Duct Occluder was successfully deployed. On echocardiography 48 h after the procedure, aortic regurgitation (AR) and residual ductal flow were noted. Long-term follow-up on clinical signs, physical examination, radiography, echocardiography, ECG, and serum NT-proBNP were evaluated until 30 months after correction of PDA. The clinical indices of physical examination, thoracic radiography, echocardiography, ECG, and serum NT-proBNP concentration were improved, although the postocclusion AR and residual ductal flow persisted. The dog followed up without clinical signs for 41 months following the correction. To our knowledge, this is the first case report to demonstrate quite a long time of follow-up (41 months) in an older dog with transarterial occlusion of PDA with postocclusion AR and residual flow.

Circulating Galectin-3 Evaluation in Dogs With Cardiac and Non-cardiac Diseases.

Galectin-3 is involved in important biological functions such as fibrogenesis and inflammation. Notably, it is associated with various diseases and plays a major role in cardiac inflammation and fibrosis. Although heart diseases are relatively common in dogs, a few studies have analyzed the circulating galectin-3 concentration in dogs with various heart diseases, including myxomatous mitral valve disease, patent ductus arteriosus, and pulmonic stenosis. The aims of the present study were to evaluate the effect of heart disease on circulating galectin-3 levels in dogs, and also to evaluate the correlation between galectin-3 concentration and conventional echocardiographic indices along with N-terminal pro-B-type natriuretic peptide (NT-proBNP) concentration in dogs with heart diseases. The medical records and archived serum samples of 107 dogs were evaluated retrospectively. In total, 107 dogs were classified into healthy dogs (n = 8), cardiac disease (n = 26), and non-cardiac disease groups (n = 73). The circulatory galectin-3 levels were analyzed using a commercially available canine-specific galectin-3 enzyme-linked immunosorbent assay kit. This study demonstrated that dogs with heart, endocrine, and dermatologic diseases had significantly higher galectin-3 levels than healthy dogs (p = 0.009, p = 0.007, and p = 0.026, respectively). Among dogs with heart diseases, dogs with concentric cardiomyopathy had significantly increased circulatory galectin-3 levels compared with healthy dogs (p = 0.028). E'/A' had a positive association with galectin-3 levels among conventional echocardiographic indices. Moreover, the galectin-3 concentration could predict diastolic dysfunction. In dogs with myxomatous mitral valve disease, a significantly positive correlation was revealed between galectin-3 levels and NT-proBNP levels (p = 0.007). Overall, this study demonstrates that circulatory galectin-3 levels increase in dogs with heart, endocrine, and dermatologic diseases. Moreover, this study demonstrates that galectin-3 concentration could be helpful to evaluate cardiac remodeling and diastolic function. Further large-scale research is required to evaluate the role of circulating galectin-3 in dogs with heart diseases.

Evaluation of circulating IGF-I and IGFBP-3 as biomarkers for tumors in dogs.

BACKGROUND: Serum-based parameters are considered non-invasive biomarkers for cancer detection. In human studies, insulin-like growth factor-I and II (IGF-I and IGF-II) and insulin-like growth factor binding protein-3 (IGFBP-3) are useful as diagnostic or prognostic markers and potential therapeutic targets. OBJECTIVES: This study examined the diagnostic utility of circulating IGF-I, IGF-II, and IGFBP-3 levels in healthy dogs and dogs with tumors. METHODS: The serum concentrations of these biomarkers in 86 dogs with tumors were compared with those in 30 healthy dogs using an enzyme-linked immunosorbent assay (ELISA). RESULTS: The ELISA results showed no difference between healthy dogs and dogs with tumors in the serum IGF-II concentrations. On the other hand, there was a significant difference in the circulating IGF-I and IGFBP-3 levels between healthy dogs and dogs with tumors. The concentrations of serum IGF-I (median [interquartile range], 103.4 [59.5-175] ng/mL) in dogs with epithelial tumors were higher than those (58.4 ng/mL [43.5-79.9]) in healthy dogs. Thus, the concentrations of serum IGFBP-3 (43.4 ng/mL [33.2-57.2]) in dogs with malignant mesenchymal tumors were lower than those (60.8 ng/mL [47.6-70.5]) in healthy dogs. CONCLUSIONS: The serum IGF-I and IGFBP-3 levels can be used as diagnostic biomarkers in dogs with tumors.

Identification and Characterization of Circulating MicroRNAs as Novel Biomarkers in Dogs With Heart Diseases.

Background: Previous studies in humans have confirmed dysregulations of circulating microRNAs (miRNAs) in patients with various cardiovascular diseases. However, studies on circulating miRNAs in dogs with various heart diseases are limited in number. This study aimed to identify significantly dysregulated circulating miRNAs and characterize them as novel biomarkers in dogs with heart diseases. Materials and Methods: Circulating levels of 11 miRNAs were investigated in serum samples of 82 dogs (72 with heart diseases and 10 healthy dogs) using quantitative reverse transcription-polymerase chain reaction. The results were correlated to clinical data including echocardiographic results and N-terminal pro B-type natriuretic peptide (NT-proBNP) levels. Results: Upregulation of cfa-miR-130b was observed in dogs with myxomatous mitral valve degeneration (MMVD) stage B, patent ductus arteriosus, and pulmonic stenosis. In dogs with MMVD stage B, cfa-miR-130b was upregulated and correlated with clinical indices. In receiver operating characteristic (ROC) analysis, cfa-miR-130b accurately distinguished dogs with diseases from healthy dogs. We also observed that cfa-miR-375 and cfa-let-7b were upregulated in dogs with concentric cardiac hypertrophy. The cfa-miR-375 was correlated with concentric hypertrophy indices and was an accurate indicator of concentric hypertrophy in ROC analysis. Conclusions: The miRNAs identified in this study may be used as novel biomarkers and possible candidates for therapeutic targets in various canine heart diseases.

Evaluation of circulating PD-1 and PD-L1 as diagnostic biomarkers in dogs with tumors.

BACKGROUND: Programmed cell death protein-1 (PD-1) and programmed cell death ligand-1 (PD-L1) have important roles in tumor evasion of the immune system. OBJECTIVES: This study aimed to assess the diagnostic utility of circulating PD-1 and PD-L1 levels in healthy dogs and dogs with tumors. METHODS: Circulating PD-1 and PD-L1 levels in the serum of 71 dogs with tumors were compared with those of 52 healthy dogs by performing enzyme-linked immunosorbent assay (ELISA). RESULTS: The ELISA results revealed higher circulating PD-1 and PD-L1 levels in dogs with tumors (2.9 [2.2-3.7] ng/mL; median [IQR] and 2.4 [1.4-4.4] ng/mL, respectively) than in healthy dogs (2.4 [1.9-3.0] ng/mL; p = 0.012 and 1.4 [0.9-2.1] ng/mL; p < 0.001, respectively). Especially, there was a significant difference in circulating PD-1 levels between healthy dogs and dogs with malignant epithelial tumors (2.4 [1.9-3.0] ng/mL and 3.1 [2.6-4.4] ng/mL, respectively; p < 0.01). In addition, there was a significant difference in circulating PD-L1 levels between healthy dogs and dogs with lymphomas (1.4 [0.9-2.1] ng/mL and 2.7 [1.6-5.8] ng/mL, respectively; p < 0.001). CONCLUSION: This study indicates that circulating PD-1 and PD-L1 have potential as tumor diagnostic biomarkers in dogs with tumors.

Expression Profile of Circulating MicroRNAs in Dogs With Cardiac Hypertrophy: A Pilot Study.

This study aimed to identify the expression profile of circulating microRNAs in dogs with eccentric or concentric cardiac hypertrophy. A total of 291 microRNAs in serum samples of five dogs with myxomatous mitral valve degeneration (MMVD) and five dogs with pulmonic stenosis (PS) were compared with those of five healthy dogs using microarray analysis. Results of microarray analysis revealed up-regulation of cfa-miR-130b [fold change (FC) = 2.13, p = 0.014), down-regulation of cfa-miR-375 (FC = 1.51, p = 0.014), cfa-miR-425 (FC = 2.56, p = 0.045), cfa-miR-30d (FC = 3.02, p = 0.047), cfa-miR-151 (FC = 1.89, p = 0.023), cfa-miR-19b (FC = 3.01, p = 0.008), and cfa-let-7g (FC = 2.53, p = 0.015) in MMVD group which showed eccentric cardiac hypertrophy, up-regulation of cfa-miR-346 (FC = 2.74, p = 0.032), down-regulation of cfa-miR-505 (FC = 1.56, p = 0.016) in PS group which showed concentric cardiac hypertrophy, and down-regulation of cfa-miR-30c (FC = 3.45, p = 0.013 in MMVD group; FC = 3.31, p = 0.014 in PS group) and cfa-let-7b (FC = 11.42, p = 0.049 in MMVD group; FC = 5.88, p = 0.01 in PS group) in both MMVD and PS groups. In addition, the unsupervised hierarchical clustering of differentially expressed microRNAs in each group resulted in complete separation of healthy dogs from dogs with heart diseases. Therefore, eleven microRNAs among 291 microRNAs were identified as differentially expressed circulating microRNAs related to MMVD or PS in dogs. This pilot study demonstrates that the microRNAs identified in this study could be possible candidates for novel biomarker or therapeutic target related to cardiac hypertrophy in dogs.

Case Report: Long-Term Survival of a Dog With Chronic Lymphocytic Leukemia Treated With Chlorambucil, Prednisolone, and Imatinib.

A 7-year-old castrated male Poodle dog presented with chronic progressive lymphocytosis. Hematologic and peripheral blood smear findings included remarkable lymphocytosis with well-differentiated small lymphocytes. Cytology of bone marrow aspirate showed hypercellular integrity with infiltration of small mature lymphocytes, accounting for 45% of all nucleated cells. Flow cytometry of blood and marrow samples revealed neoplastic lymphocytes predominantly expressing the CD21 molecule. B-cell chronic lymphocytic leukemia (CLL) was diagnosed on an immunophenotypic analysis. Administrations of prednisolone and chlorambucil were initiated and the response was unremarkable. Therefore, additional treatment with imatinib was provided, which resolved the hematologic abnormalities associated with CLL. Flow cytometry after ~1 year of treatment showed normalization of the count of lymphocytes positive for CD21 and resolved hematologic lymphocytosis. The dog was followed-up for 2 years, and there were no severe adverse effects. This case indicates that imatinib may be a good option as an adjunctive therapy with prednisolone and chlorambucil treatment for CLL in dogs without treatment response.

Whole-exome and whole-transcriptome sequencing of canine mammary gland tumors.

Studies of naturally occurring cancers in dogs, which share many genetic and environmental factors with humans, provide valuable information as a comparative model for studying the mechanisms of human cancer pathogenesis. While individual and small-scale studies of canine cancers are underway, more generalized multi-omics studies have not been attempted due to the lack of large-scale and well-controlled genomic data. Here, we produced reliable whole-exome and whole-transcriptome sequencing data of 197 canine mammary cancers and their matched controls, annotated with rich clinical and biological features. Our dataset provides useful reference points for comparative analysis with human cancers and for developing novel diagnostic and therapeutic technologies for cancers in pet dogs.

Genome-wide association study of degenerative mitral valve disease in Maltese dogs.

Genome-wide association study (GWAS) is a powerful tool for identifying the genetic causes of various diseases. This study was conducted to identify genomic variation in Maltese dog genomes associated with degenerative mitral valve disease (DMVD) development and to evaluate the association of each biological condition with DMVD in Maltese dogs. DNA was extracted from blood samples obtained from 48 Maltese dogs (32 with DMVD and 16 controls). Genome-wide single nucleotide polymorphism (SNP) genotyping was performed. The top 30 SNPs from each association of various conditions and genetic variations were mapped to their gene locations. A total of 173,662 loci were successfully genotyped, with an overall genotype completion rate of 99.41%. Quality control analysis excluded 46,610 of these SNPs. Manhattan plots were produced using allelic tests with various candidate clinical conditions. A significant peak of association was observed between mitral valve prolapse (MVP) and SNPs on chromosome 17. The present study revealed significant SNPs in several genes associated with cardiac function, including PDZ2, Armadillo repeat protein detected in velo-cardio-facial syndrome, catenin (cadherin-associated protein) alpha 3, low-density lipoprotein receptor class A domain containing protein 4, and sterile alpha motif domain containing protein 3. To our knowledge, this is the first study of a genetic predisposition to DMVD in Maltese dogs. Although only a limited number of cases were analyzed, these data could be the basis for further research on the genetic predisposition to MVP and DMVD in Maltese dogs.

Increased Expression of Osteopontin in the Degenerating Striatum of Rats Treated with Mitochondrial Toxin 3-Nitropropionic Acid: A Light and Electron Microscopy Study.

The mycotoxin 3-nitropropionic acid (3NP) is an irreversible inhibitor that induces neuronal damage by inhibiting mitochondrial complex II. Neurodegeneration induced by 3NP, which is preferentially induced in the striatum, is caused by an excess influx and accumulation of calcium in mitochondria. Osteopontin (OPN) is a glycosylated phosphoprotein and plays a role in the regulation of calcium precipitation in the injured brain. The present study was designed to examine whether induction of OPN protein is implicated in the pathogenesis of 3NP-induced striatal neurodegeneration. We observed overlapping regional expression of OPN, the neurodegeneration marker Fluoro-Jade B, and the microglial marker ionized calcium-binding adaptor molecule 1 (Iba1) in the 3NP-lesioned striatum. OPN expression was closely associated with the mitochondrial marker NADH dehydrogenase (ubiquinone) flavoprotein 2 in the damaged striatum. In addition, immunoelectron microscopy demonstrated that OPN protein was specifically localized to the inner membrane and matrix of the mitochondria in degenerating striatal neurons, and cell fragments containing OPN-labeled mitochondria were also present within activated brain macrophages. Thus, our study revealed that OPN expression is associated with mitochondrial dysfunction produced by 3NP-induced alteration of mitochondrial calcium homeostasis, suggesting that OPN is involved in the pathogenesis of striatal degeneration by 3NP administration.