Adjunct ambrisentan therapy had clinical benefits in 5 dogs with sildenafil-refractory pulmonary hypertension.

Although sildenafil is used in dogs with severe pulmonary hypertension, they sometimes become resistant and clinical signs deteriorate over time. The objective of this study was to determine the benefits of adjunct ambrisentan therapy in dogs with sildenafil-refractory pulmonary hypertension. In 5 dogs with severe pulmonary hypertension with deteriorating clinical signs despite ongoing sildenafil treatment, adding ambrisentan improved appetite, activity, and respiratory functions. Although peak tricuspid valve regurgitation velocity, as measured by Doppler echocardiography, did not necessarily decrease after ambrisentan administration, there was improved partial pressure of arterial oxygen and the alveolar-arterial oxygen gradient, with no apparent side effects. We concluded that ambrisentan has potential as an adjunct treatment in dogs with pulmonary hypertension that are refractory to sildenafil therapy. Key clinical message: Ambrisentan improved clinical signs in dogs with sildenafil-refractory pulmonary hypertension.

Le traitement d'appoint à l'ambrisentan a eu des avantages cliniques chez cinq chiens atteints d'hypertension pulmonaire réfractaire au sildénafil. Bien que le sildénafil soit utilisé chez les chiens souffrant d'hypertension pulmonaire sévère, ils deviennent parfois résistants et les signes cliniques s'aggravent avec le temps. L'objectif de cette étude était de déterminer les avantages d'un traitement d'appoint à l'ambrisentan chez les chiens souffrant d'hypertension pulmonaire réfractaire au sildénafil. Chez cinq chiens souffrant d'hypertension pulmonaire sévère avec détérioration des signes cliniques malgré un traitement continu au sildénafil, l'ajout d'ambrisentan a amélioré l'appétit, l'activité et les fonctions respiratoires. Bien que la vitesse maximale de régurgitation de la valve tricuspide, mesurée par échocardiographie Doppler, n'ait pas nécessairement diminué après l'administration d'ambrisentan, la pression partielle d'oxygène artériel et le gradient alvéolo-artériel d'oxygène ont été améliorés, sans effets secondaires apparents. Nous avons conclu que l'ambrisentan a un potentiel en tant que traitement d'appoint chez les chiens souffrant d'hypertension pulmonaire qui sont réfractaires au traitement par le sildénafil.Message clinique clé:L'ambrisentan a amélioré les signes cliniques chez les chiens souffrant d'hypertension pulmonaire réfractaire au sildénafil.(Traduit par Dr Serge Messier).

Establishment of a novel experimental model for muscle-invasive bladder cancer using a dog bladder cancer organoid culture.

In human and dogs, bladder cancer (BC) is the most common neoplasm affecting the urinary tract. Dog BC resembles human muscle-invasive BC in histopathological characteristics and gene expression profiles, and could be an important research model for this disease. Cancer patient-derived organoid culture can recapitulate organ structures and maintains the gene expression profiles of original tumor tissues. In a previous study, we generated dog prostate cancer organoids using urine samples, however dog BC organoids had never been produced. Therefore we aimed to generate dog BC organoids using urine samples and check their histopathological characteristics, drug sensitivity, and gene expression profiles. Organoids from individual BC dogs were successfully generated, expressed urothelial cell markers (CK7, CK20, and UPK3A) and exhibited tumorigenesis in vivo. In a cell viability assay, the response to combined treatment with a range of anticancer drugs (cisplatin, vinblastine, gemcitabine or piroxicam) was markedly different in each BC organoid. In RNA-sequencing analysis, expression levels of basal cell markers (CK5 and DSG3) and several novel genes (MMP28, CTSE, CNN3, TFPI2, COL17A1, and AGPAT4) were upregulated in BC organoids compared with normal bladder tissues or two-dimensional (2D) BC cell lines. These established dog BC organoids might be a useful tool, not only to determine suitable chemotherapy for BC diseased dogs but also to identify novel biomarkers in human muscle-invasive BC. In the present study, for the 1st time, dog BC organoids were generated and several specifically upregulated organoid genes were identified. Our data suggest that dog BC organoids might become a new tool to provide fresh insights into both dog BC therapy and diagnostic biomarkers.

Cloning of carrier cells infected with oncolytic adenovirus driven by midkine promoter and biosafety studies.

BACKGROUND: A549 carrier cells infected with oncolytic adenovirus can induce complete tumor reduction of subcutaneous ovarian tumors but not intraperitoneal disseminated ovarian tumors. This appears to be a result of the insufficient antitumor effect of A549 carrier cells. Therefore, in the present study, we cloned a novel carrier cell with the aim of improving the antitumor effects. METHODS: Carrier cells infected with oncolytic adenovirus AdE3-midkine with a midkine promoter were cloned by limiting dilution. We examined the antitumor effects of these cells on subcutaneous and intraperitoneal OVHM ovarian tumors in a syngeneic mouse model. Biosafety tests were conducted in beagle dogs and rabbits. RESULTS: We cloned EHMK-51-35 carrier cells with 10-fold higher antitumor effects compared to A549 carrier cells in vitro. EHMK-51-35 carrier cells co-infected with AdE3-midkine and Ad-mGM-CSF induced a 100% complete tumor reduction in subcutaneous tumors and a 60% reduction of intraperitoneal disseminated tumors. Single-dose acute toxicity test on beagle dogs with EHMK-51-35 carrier cells co-infected with AdE3-midkine and Ad-cGM-CSF showed no serious side effects. Biologically active adenoviruses were not detected in the blood, saliva, feces, urine or whole organs. In a chronic toxicity test, VX2 tumors in rabbits were injected five times with EHMK-51-35 carrier cells infected with AdE3-midkine and these rabbits showed no serious side effects. CONCLUSIONS: Significant antitumor effects and safety of cloned EHMK-51-35 carrier cells were confirmed in intraperitoneal ovarian tumors and toxicity tests, respectively. These findings will be extended to preclinical efficacy studies using dogs and cats, with the aim of conducting human clinical trials on refractory solid tumors.

Characterization of feline cytochrome P450 2B6.

1. Little is known about drug metabolism in carnivores. Although the domestic cat (Felis catus) is an obligate carnivore and is the most common companion animal, usage and dosage of many drugs are determined according to information obtained from humans and dogs. We determined the complete cDNA sequence of CYP2B6 from the feline lung. 2. Feline CYP2B6 consists of 494 deduced amino acids, showing highest identity with the dog CYP2B ortholog, followed by those of horse, pig, primate and human. 3. Feline CYP2B6 transcripts were expressed predominantly in the lung and slightly in the small intestine but not in the liver without significant sex-dependent differences. Western blot analysis with an anti-human CYP2B6 antibody confirmed the presence of CYP2B protein in the lung but not in the liver. 4. Feline CYP2B6 proteins heterologously expressed in Escherichia coli metabolized several substrates specific to human CYP2B6, including 7-ethoxy-4-(trifluoromethyl) coumarin (EFC). The metabolic activity was strongly inhibited by medetomidine and atipamezole, potent inhibitors of canine CYP2B11 (now officially CYP2B6) as well as by ticlopidine and sertraline, inhibitors selective to human CYP2B6. 5. The results suggest that feline CYP2B6 is a functional CYP2B ortholog that plays a role in the local defense mechanism in the cat respiratory system and intestine.

Identification and functional characterization of novel feline cytochrome P450 2A.

1. Cytochrome P450s are the major metabolizing enzymes for xenobiotics in humans and other mammals. Although the domestic cat Felis catus, an obligate carnivore, is the most common companion animal, the properties of cytochrome P450 subfamilies are largely unknown. 2. We newly identified the feline CYP2A13, which consists of 494 deduced amino acids, showing the highest identity to CYP2As of dogs, followed by those of pigs, cattle and humans. 3. The feline CYP2A13 transcript and protein were expressed almost exclusively in the liver without particular sex-dependent differences. 4. The feline CYP2A13 protein heterogeneously expressed in Escherichia coli showed metabolic activity similar to those of human and canine CYP2As for coumarin, 7-ethoxycoumarin and nicotine. 5. The results indicate the importance of CYP2A13 in systemic metabolism of xenobiotics in cats.

Ultrasonographic findings related to prognosis in canine transitional cell carcinoma.

In human bladder cancer patients, ultrasonography is extensively used not only to identify tumor masses but also to evaluate tumor size, shape, echogenicity, location, and degree of tumor invasion into the bladder wall. The information revealed by ultrasonography delineates the tumor's biological features and facilitates prediction of prognosis. However, in veterinary medicine the feasibility of using ultrasonography for these purposes has not been fully investigated. In this retrospective study, we reviewed cases of dogs with histologically confirmed bladder mass lesions, including transitional cell carcinoma (n = 22) and polypoid cystitis (n = 5), to determine whether ultrasonography could reliably predict bladder wall involvement. By following patients with transitional cell carcinoma until death, we also determined whether ultrasonographic tumor size, shape, echogenicity, and mass location were related to prognosis. Wall involvement as revealed by ultrasound was significantly (P = 0.00005) associated with histological muscular layer involvement with a sensitivity of 93% (95% Confidence interval, 79-98%) and specificity of 92% (95% Confidence interval, 76-98%). Ultrasonographic wall involvement (P = 0.03, vs. noninvolvement), heterogeneous mass (P = 0.02, vs. homogeneous mass), and trigone location (P = 0.01, vs. other locations) characteristics were significantly associated with shorter survival times in transitional cell carcinoma cases. Findings indicated that ultrasonographic characteristics such as wall involvement, heterogeneous mass, and trigone location could be reliable prognostic indicators in canine transitional cell carcinoma.

A new treatment for human malignant melanoma targeting L-type amino acid transporter 1 (LAT1): a pilot study in a canine model.

L-type amino acid transporter 1 (LAT1), an isoform of amino acid transport system L, transports branched or aromatic amino acids essential for fundamental cellular activities such as cellular growth, proliferation and maintenance. This amino acid transporter recently has received attention because of its preferential and up-regulated expression in a variety of human tumors in contrast to its limited distribution and low-level expression in normal tissues. In this study, we explored the feasibility of using LAT1 inhibitor as a new therapeutic agent for human malignant melanomas (MM) using canine spontaneous MM as a model for human MM. A comparative study of LAT expression was performed in 48 normal tissues, 25MM tissues and five cell lines established from MM. The study observed LAT1 mRNA levels from MM tissues and cell lines that were significantly (P<0.01) higher than in normal tissues. Additionally, MM with distant metastasis showed a higher expression than those without distant metastasis. Functional analysis of LAT1 was performed on one of the five cell lines, CMeC-1. [(3)H]l-Leucine uptake and cellular growth activities in CMeC-1 were inhibited in a dose-dependent manner by selective LAT1 inhibitors (2-amino-2-norbornane-carboxylic acid, BCH and melphalan, LPM). Inhibitory growth activities of various conventional anti-cancer drugs, including carboplatin, cyclophosphamide, dacarbazine, doxorubicin, mitoxantrone, nimustine, vinblastine and vincristine, were significantly (P<0.05) enhanced by combination use with BCH or LPM. These findings suggest that LAT1 could be a new therapeutic target for MM.

Chemosensitivity of three patient-derived primary cultures of canine pericardial mesothelioma by single-agent and combination treatment.

Introduction: Canine mesothelioma is a rare malignant tumor that mostly affects body cavities, such as the pericardial and pleural cavities. Chemotherapy plays a crucial role in the treatment of canine mesotheliomas. We aimed to compare the antitumor effects of single-agent and combination chemotherapeutic agents on patient-derived primary cultures of canine pericardial mesothelioma established in this study. We planned to generate xenograft models for future studies. Material and methods: Effusion samples were collected from three dogs with histologically diagnosed pericardial mesothelioma and used for primary culture. Cultured cells were characterized by immunostaining for pan-cytokeratin AE1/AE3, vimentin, Wilms' tumor suppressor gene 1 (WT1), and cytokeratin 5 (CK5). To assess the tumorigenic properties of cells in the effusion and generate a xenograft model, the cell suspension was injected into a severe combined immunodeficient (SCID) mouse either subcutaneously (SC) or intraperitoneally (IP). Lastly, chemosensitivity of established primary cultures against four drugs, doxorubicin, vinorelbine, carboplatin, and gemcitabine, by single-agent treatment as well as combination treatment of carboplatin at a fixed concentration, either 10 or 100 µM, and gemcitabine at different concentrations ranging from 0-1000 µM was assessed by cell viability assay. Results: Primary cultures were successfully generated and characterized by dual positivity for AE1/AE3 and vimentin and positive staining for WT-1 and CK5, confirming the mesothelial origin of the cells. In the xenograft models, SC mouse developed a subcutaneous mass, whereas IP mouse developed multiple intraperitoneal nodules. The masses were histopathologically consistent with mesotheliomas. The chemosensitivity assay revealed that carboplatin had the highest anti-tumor effects among the four tested single-agent treatments. Furthermore, carboplatin at 100 µM combined with gemcitabine at clinically relevant doses demonstrated the augmented anti-tumor effects compared to single-agent treatment. Discussion and conclusion: Primary cultures and xenograft models generated in this study could be useful tools for in vitro and in vivo studies of canine mesothelioma. Carboplatin is a highly effective chemotherapeutic agent against canine mesothelioma when used as a sole agent and in combination with gemcitabine.

Establishment of an experimental model of canine malignant mesothelioma organoid culture using a three-dimensional culture method.

Canine malignant mesothelioma (cMM) is a rare and drug-resistant malignant tumor. Due to few patients and experimental models, there have not been enough studies to demonstrate the pathogenesis of the disease and novel effective treatment for cMM. Since cMM resembles human MM (hMM) in histopathological characteristics, it is also considered a promising research model of hMM. Compared with conventional 2-dimensional (2D) culture methods, 3-dimensional (3D) organoid culture can recapitulate the properties of original tumor tissues. However, cMM organoids have never been developed. In the present study, we for the first time generated cMM organoids using the pleural effusion samples. Organoids from individual MM dogs were successfully generated. They exhibited the characteristics of MM and expressed mesothelial cell markers, such as WT-1 and mesothelin. The sensitivity to anti-cancer drugs was different in each strain of cMM organoids. RNA sequencing analysis showed cell adhesion molecule pathways were specifically upregulated in cMM organoids compared with their corresponding 2D cultured cells. Among these genes, the expression level of E-cadherin was drastically higher in the organoids than that in the 2D cells. In conclusion, our established cMM organoids might become a new experimental tool to provide new insights into canine and human MM therapy.

Derivation of a new model of lung adenocarcinoma using canine lung cancer organoids for translational research in pulmonary medicine.

Canine primary lung cancer (cPLC) is a rare malignant tumor in dogs, and exhibits poor prognosis. Effective therapeutic drugs against cPLC have not been established yet. Also, cPLC resembles human lung cancer in histopathological characteristics and gene expression profiles and thus could be an important research model for this disease. Three-dimensional organoid culture is known to recapitulate the tissue dynamics in vivo. We, therefore, tried to generate cPLC organoids (cPLCO) for analyzing the profiles of cPLC. After samples from cPLC and the corresponding normal lung tissue were collected, cPLCO were successfully generated, which recapitulated the tissue architecture of cPLC, expressed lung adenocarcinoma marker (TTF1), and exhibited tumorigenesis in vivo. The sensitivity of cPLCO to anti-cancer drugs was different among strains. RNA-sequencing analysis showed significantly upregulated 11 genes in cPLCO compared with canine normal lung organoids (cNLO). Moreover, cPLCO were enriched with the MEK-signaling pathway compared with cNLO. The MEK inhibitor, trametinib decreased the viability of several strains of cPLCO and inhibited the growth of cPLC xenografts. Collectively, our established cPLCO model might be a useful tool for identifying novel biomarkers for cPLC and a new research model for dog and human lung cancer.

Establishment of an experimental model of normal dog bladder organoid using a three-dimensional culture method.

Dog bladder cancer (BC) is mostly muscle-invasive (MI) with poor prognosis, and its pathogenesis is close to human MIBC. Three-dimensional (3D) organoid culture ensures novel knowledge on cancer diseases including BC. Recently, we have established dog BC organoids (BCO) using their urine samples. BCO recapitulated the epithelial structures, characteristics, and drug sensitivity of BC-diseased dogs. However, organoids from dog normal bladder epithelium are not established yet. Therefore, the present study aimed to establish dog normal bladder organoids (NBO) for further understanding the pathogenesis of dog BC and human MIBC. The established NBO underwent various analyzes including cell marker expressions, histopathological structures, cancer-related gene expression patterns, and drug sensitivity. NBO could be produced non-invasively with a continuous culturing and recapitulated the structures and characteristics of the dog's normal bladder mucosal tissues. Different drug sensitivities were observed in each NBO. The analysis of RNA sequencing revealed that several novel genes were changed in NBO compared with BCO. NBO showed a higher expression of p53 and E-cadherin, but a lower expression of MDM2 and Twist1 compared with BCO. These results suggest that NBO could be a promising experimental 3D model for studying the developmental mechanisms of dog BC and human MIBC.

Measurement of Pulmonary Artery Wave Reflection Before and After Mitral Valvuloplasty in Canine Patients With Pulmonary Hypertension Caused by Myxomatous Mitral Valve Disease.

Background: Pulmonary arterial wave reflection provides novel information about pulmonary artery hemodynamics in pulmonary hypertension (PH). PH is common in dogs with myxomatous mitral valve disease (MMVD), though research examining the relationship between pulmonary arterial wave reflection and MMVD with PH is lacking. Hypothesis/Objective: This study investigated conventional echocardiographic parameters and pulmonary artery wave reflection parameters before and after mitral valvuloplasty in canine patients with PH due to MMVD. The parameters were backward pressure (Pb), forward pressure (Pf), and the reflection coefficient calculated as the ratio of peak Pb to peak Pf (RC). Animals: The study subjects were 10 client-owned dogs receiving mitral valvuloplasty for MMVD with PH. Methods: Conventional echocardiographic parameters and pulmonary artery wave reflection parameters were measured before and after mitral valvuloplasty. The relationships between pulmonary artery wave reflection parameters and echocardiographic parameters, estimation of pulmonary artery systolic pressure, and right atrium pressure (RAP) gained by catheter in mitral valvuloplasty were also investigated. Post-operative echocardiography and the measurement of pulmonary arterial wave reflection were performed 2 weeks after mitral valvuloplasty. Results: The parameters of normalized left ventricular internal diameter at end-diastole (LVIDDN), E velocity, and the estimation of pulmonary artery systolic pressure were significantly reduced post-operatively compared with baseline measurements (p < 0.05). Post-operative Pb decreased significantly compared with pre-operative measurements (8.8 ± 5.9 to 5.0 ± 3.2 mmHg, p = 0.037) as did RC (0.37 ± 0.15 to 0.22 ± 0.11, p < 0.01). A statistically significant positive correlation existed between wave reflection parameters and RAP, an estimation of pulmonary artery systolic pressure. Conclusions: Results demonstrate that mitral valvuloplasty can be used to treat secondary PH caused by MMVD, resulting in the improvement of post-operative echocardiographic and wave reflection parameters and a decrease in the right afterload. In some patients, some degree of vascular admittance mismatch persisted, despite the improvement of left atrial pressure. This may be indicative of residual pulmonary arterial disease, which may continue to adversely affect interactions between the right ventricle and the vasculature.

Prognostic role of ΔNp63 expression in canine transitional cell carcinoma of the urinary bladder.

BACKGROUND: Decreased p63 protein expression in canine transitional cell carcinoma (TCC) of the urinary bladder is associated with vascular invasion of the tumor, metastasis, and shortened survival. ΔNp63, an isoform of p63, is downregulated in high-grade invasive urothelial carcinoma in humans. However, the clinical significance of ΔNp63 expression in canine urinary bladder tumors is unknown. Therefore, it is essential to investigate ΔNp63 expression patterns in TCC, the most common urinary bladder tumor in dogs. AIM: This study aimed to evaluate the expression and role of ΔNp63 in canine TCC of the urinary bladder. METHODS: ΔNp63 expression was compared between the normal canine urinary bladder, polypoid cystitis, and TCC. The correlation of ΔNp63 expression with histopathological and clinical findings were further evaluated, and its usefulness as a prognostic factor was examined. RESULTS: We observed that ΔNp63 was highly expressed in dogs' normal urinary bladder and polypoid cystitis, and its expression levels were low in TCC. Furthermore, low levels of ΔNp63 expression were associated with vascular invasion, metastasis, and shortened survival in dogs with TCC. CONCLUSION: These results indicate that ΔNp63 expression could serve as a valuable biomarker for invasion, metastasis, and prognosis of canine TCC of the urinary bladder.

Evaluation of the Safety and Feasibility of Apheresis in Dogs: For Application in Metastatic Cancer Research.

In patients with solid tumors, circulating tumor cells (CTCs) spread in their blood and function as a seed for metastases. However, the study of CTCs has been limited by their rarity, low frequency, and heterogeneity. The efficient collection of CTCs will contribute to further research of metastatic cancers. Apheresis is a process in which the whole blood of an individual is passed through a machine that isolates a particular constituent and returns the remainder to the circulation. In the present study, we investigated the safety and feasibility of apheresis to separate peripheral blood monocytes (PBMCs), whose density is closely similar to that of CTCs, and to capture intravenously administered human breast cancer cells, MCF7s, from the dogs. No life-threatening events were observed in dogs during the apheresis process. The changes in the hemogram were transient and recovered gradually within a few days after apheresis. During apheresis, 50 mL of PBMCs could be collected from each dog. Notably, a thrombus was formed along the circuit wall during apheresis, which decreased the blood collection pressure. MCF7 cells were successfully captured by the apheresis machine. The captured cells were regrown in vitro and characterized compared with the original cells. In conclusion, apheresis could be safely performed in dogs to isolate CTCs with precautions to maintain hemodynamic stability.

Anti-tumor effect of trametinib in bladder cancer organoid and the underlying mechanism.

Bladder cancer (BC), a main neoplasm of urinary tract, is usually inoperable and unresponsive to chemotherapy. As a novel experimental model for muscle-invasive BC, we previously established a culture method of dog BC organoids. In the present study, the detailed in vitro and in vivo anti-tumor effects of trametinib were investigated by using this model. In each BC organoid strain, epidermal growth factor receptor (EGFR)/ERK signaling was upregulated compared with normal bladder cells. Trametinib even at a low concentration inhibited the cell viability of BC organoids and the activation of ERK through decreasing expression of c-Myc, ELK1, SIK1, and PLA2G4A. Trametinib arrested cell cycle of BC with few apoptosis. Dual treatment of BC organoids with trametinib and YAP inhibitor, verteporfin extremely inhibited the cell viability with apoptosis induction. Moreover, trametinib induced basal to luminal differentiation of BC organoids by upregulating luminal markers and downregulating basal ones. In vivo, trametinib decreased the tumor growth of BC organoids in mice and the xenograft-derived organoids from trametinib-administered mice showed enhanced sensitivity to carboplatin due to MSH2 upregulation. Our data suggested a new strategy of trametinib-YAP inhibitor or trametinib-carboplatin combination as a promising treatment of BC.

Antimuscarinic action of doxorubicin does not involve free-radical formation in isolated guinea pig hearts.

It has been proposed that the cardiotoxicity of anthracycline anticancer drugs involves free-radical formation. One early manifestation of toxicity appears to be caused by the antimuscarinic actions of these drugs. Accordingly, we examined whether the antimuscarinic action of one of these drugs, doxorubicin, is altered by antioxidants. In electrically stimulated left atrial muscle preparations obtained from guinea pig hearts, doxorubicin significantly increased the tissue concentration of thiobarbituric acid-reactive substance indicating increased lipid peroxidation. This effect of doxorubicin was significantly suppressed by the antioxidants alpha-tocopherol, dexrazoxane, and epigallocatechin gallate. Carbachol produced a concentration-dependent negative inotropic effect in our atrial preparations. Doxorubicin caused a seemingly parallel rightward shift of the concentration-response curve for carbachol. Neither alpha-tocopherol, dexrazoxane, nor epigallocatechin gallate reversed this effect of doxorubicin. The results indicate that in extirpated heart tissue, doxorubicin causes lipid peroxidation through the formation of free radicals. However, this effect of doxorubicin is unrelated to its antimuscarinic action.

Establishment of 2.5D organoid culture model using 3D bladder cancer organoid culture.

Three-dimensional (3D) organoid culture holds great promises in cancer precision medicine. However, Matrigel and stem cell-stimulating supplements are necessary for culturing 3D organoid cells. It costs a lot of money and consumes more time and effort compared with 2D cultured cells. Therefore, the establishment of cheaper and Matrigel-free organoid culture that can maintain the characteristics of a part of 3D organoids is demanded. In the previous study, we established a dog bladder cancer (BC) 3D organoid culture system by using their urine samples. Here, we successfully isolated cells named "2.5D organoid" from multiple strains of dog BC 3D organoids using 2.5 organoid media. The cell proliferation speed of 2.5D organoids was faster than parental 3D organoid cells. The expression pattern of stem cell markers was close to 3D organoids. Injection of 2.5D organoid cells into immunodeficient mice formed tumors and showed the histopathological characteristics of urothelial carcinoma similar to the injection of dog BC 3D organoids. The 2.5D organoids had a similar sensitivity profile for anti-cancer drug treatment to their parental 3D organoids. These data suggest that our established 2.5D organoid culture method might become a reasonable and useful tool instead of 3D organoids in dog BC research and therapy.

Sjögren's-like syndrome in a dog.

A neutered male Golden Retriever was referred with a 2-week history of dry mouth. Multiple and bilateral enlargement of the lacrimal and salivary glands showing heterogeneous internal enhancement was identified on contrast-enhanced computed tomography (CT). Ultrasonographic examination detected multifocal hypoechoic areas within the swollen submandibular salivary glands, which were histopathologically diagnosed as lymphoplasmacytic sialoadenitis. As both imaging and histopathological findings were in accordance with those in human Sjögren's syndrome, a provisional diagnosis of Sjögren's-like syndrome was made. Immunosuppressive drugs promptly improved clinical signs concurrently with the abnormal sonographic findings, indicating the feasibility of ultrasonography in monitoring therapeutic outcomes. Herein, we discuss a proposed criteria set for diagnosis of Sjögren's-like syndrome in veterinary medicine.

Pericardial Mesothelioma in a Dog: The Feasibility of Ultrasonography in Monitoring Tumor Progression.

A 6-year-old neutered male Yorkshire Terrier presented with recurrent pericardial effusion. Although clinical examinations including computed tomography were inconclusive, an exploratory thoracotomy revealed multiple small nodules and plaques on the inner surface of the pericardial sac (Day 1). A subtotal pericardiectomy was performed to prevent cardiac tamponade due to the increasing pericardial effusion, and the resected section of the pericardium was histopathologically diagnosed with mesothelioma. After surgery, chemotherapy with intrathoracic carboplatin was commenced. During the course of the treatment, a detailed follow-up ultrasonographic scan was performed to detect early lesions disseminated on the pleura, originating from the primary pericardial mesothelioma. On Day 101, the minute pleural nodules, which were disseminated lesions as predicted, were successfully imaged by ultrasonography. As the clinical stage advanced, the nodules were observed to gradually increase in size and number, implying tumor progression. These observations highlight the feasibility of ultrasonography in detecting minute disseminated lesions at an early stage, monitoring tumor progression, and thereby, predicting the prognosis of canine pericardial mesothelioma.

A case of feline primary duodenal carcinoid with intestinal hemorrhage.

A 15-year-old neutered male Persian cat was presented with recurrent hematemesis and melena. Abdominal ultrasonography and computed tomography revealed a mass in the proximal descending duodenal wall. Endoscopic examination revealed hemorrhage on the luminal side of the mass. Fine-needle aspiration of the mass was performed. Microscopic analysis revealed a cluster of cells with oval nuclei and indistinct cell borders, suggesting a neoplastic disease of neuroendocrine origin. The mass located near the major duodenal papilla was partially resected, and the bleeding was stopped by cauterization. However, the surgical procedures could not control the hemorrhage from the tumor mass, and the cat died of severe anemia. Immunohistopathological analysis revealed that the tumor was a duodenal carcinoid.