Efficacy of therapeutic plasma exchange in a case of severe, generalised canine pemphigus foliaceus.

Successful treatment of pemphigus foliaceus (PF) often requires a multimodal therapeutic approach. The dog described herein underwent four therapeutic plasma exchange treatments for severe, refractory PF, resulting in a 50% reduction of lesional body surface area. This treatment option should be considered for the management of canine PF.

O tratamento bem-sucedido do pênfigo foliáceo (PF) geralmente requer uma abordagem terapêutica multimodal. O cão aqui descrito foi submetido a quatro tratamentos de troca de plasma terapêutica (TPE) para PF grave e refratário, resultando em uma redução de 50% da área corpórea lesional. Esta opção de tratamento deve ser considerada para o manejo do PF canino.

El tratamiento exitoso del pénfigo foliáceo (PF) a menudo requiere un enfoque terapéutico multimodal. El perro aquí descrito se sometió a cuatro tratamientos terapéuticos de intercambio plasmático (TPE) para un PF refractario grave, lo que resultó en una reducción del 50% de la superficie corporal lesionada. Esta opción de tratamiento debe considerarse para el control de PF canino.

Traiter efficacement le pemphigus foliacé (PF) nécessite souvent une approche thérapeutique multimodale. Dans ce rapport clinique, un chie a reçu quatre traitements de plasmaphérèse thérapeutique (EPT) pour le traitement d'un PF sévère et réfractaire, ce qui a permis de réduire de 50 % la surface corporelle lésionnelle. Cette option thérapeutique devrait être envisagée pour la prise en charge du PF canin.

Optical coherence tomography for surgical margin evaluation of excised canine cutaneous and subcutaneous tumours.

Currently, intraoperative tumour margin imaging is not routinely utilized in veterinary medicine. Optical coherence tomography (OCT) allows for real-time assessment of tissue morphology of 1-2 mm depth. The aims of this study were (1) to compare the histologic and OCT features of excised canine skin and subcutaneous specimens, and (2) to determine the diagnostic accuracy of OCT for surgical margin evaluation. The authors hypothesized that OCT imaging would correlate well with histopathology and that OCT would be sensitive for detection of incomplete margins. Eighty dogs were prospectively enrolled. Tumours were excised, and the surgical margins were imaged using a spectral domain OCT system. The tumour type and completeness of excision were determined by histopathology. Nine blinded observers received training in OCT image interpretation and were then given a set of OCT images and videos. The observers assigned each image/video a grade from 1 (no tumour) to 4 (tumour) and the results were compared to histopathology. The overall median sensitivity and specificity of OCT imaging for detection of incomplete margins were 86.7% and 84.6%, respectively. A potential limitation is that observers had varied experience with OCT image interpretation, ranging from no prior experience to participating in a previous OCT project. OCT is sensitive for detection of incomplete margins and could be a promising real-time surgical margin imaging modality. Further study is needed to evaluate intraoperative applications of OCT and its impact on tumour recurrence and long-term outcome.

Lymph Node Abscessation Secondary to Neoplasia in Two Dogs.

A 5-year-old male neutered mixed breed dog and an 8-year-old female spayed golden retriever presented for cervical swelling which was later diagnosed as abscessation of the retropharyngeal lymph node with a malignant round cell tumor and carcinoma with multifocal squamous differentiation, respectively. In veterinary medicine, there is limited published information regarding abscessation of lymph nodes secondary to a neoplastic process. While more common in humans, there are only limited case reports available. Advanced imaging (computed tomography), cytology, surgical excision, and histopathology lead to the final diagnosis. Both dogs underwent surgical extirpation of the lymph nodes and adjuvant chemotherapy protocols. Six weeks postsurgical excision, dog one was euthanized due to quality-of-life concerns. The second dog successfully completed 18 treatments of radiation therapy and was still alive at 388 days postsurgical excision. At the time of manuscript submission, the second dog was doing well clinically.

Pathology in Practice.

In collaboration with the American College of Veterinary Pathologists.

Case Report: Metastatic Parosteal Osteosarcoma in a Dog.

This case report describes a rare form of malignant bone tumor in an 8-year-old Labrador retriever. This dog initially presented for evaluation of a right distal humeral mass. Radiographs of the right elbow and thorax were performed, revealing a smooth mineralized mass adjacent to the lateral aspect of the distal humerus and a 5mm pulmonary nodule. Computed tomography (CT) of the humerus and thorax showed a smooth mineralized lesion adjacent to the lateral humeral epicondyle, and a right cranial lung lobe nodule with a thin mineral rim. Surgical biopsies of both lesions were diagnostic for parosteal osteosarcoma (POSA). The dog was then treated with stereotactic body radiation therapy (SBRT) which controlled the dog's discomfort for 14 months until he became progressively painful and subsequently had his right forelimb amputated. This case report is the first to document the CT imaging characteristics of a metastatic appendicular POSA in a dog and the first dog described with POSA treated with SBRT. The dog lived for 623 days after histopathologic diagnosis and 849 days after initial presentation with pulmonary metastatic disease.

Diagnostic accuracy of optical coherence tomography for surgical margin assessment of feline injection-site sarcoma.

The invasive, locally aggressive nature of feline injection-site sarcomas (FISSs) poses a unique challenge for surgeons to obtain complete margins with surgical excision. Optical coherence tomography (OCT), an imaging technology that uses light waves to generate real-time views of tissue architecture, provides an emerging solution to this dilemma by allowing fast, high-resolution scanning of surgical margins. The purpose of this study was to use OCT to assess surgical margins of FISS and to evaluate the diagnostic accuracy of OCT for detecting residual cancer using six evaluators of varying experience. Five FISSs were imaged with OCT to create a training set of OCT images that were compared with histopathology. Next, 25 FISSs were imaged with OCT prior to histopathology. Six evaluators of varying experience participated in a training session on OCT imaging after which each of the evaluators was given a dataset that included OCT images and videos to score on a scale from cancerous to non-cancerous. Diagnostic accuracy statistics were calculated. The overall sensitivity and specificity for classification of OCT images by evaluators were 78.9% and 77.6%, respectively. Correct classification rate of OCT images was associated with experience, while individual sensitivities and specificities had more variation between experience groups. This study demonstrates the ability of evaluators to correctly classify OCT images with overall low levels of experience and training and also illustrates areas where increased training can improve accuracy of evaluators in interpretation of OCT surgical margin images.

Treatment of indolent cutaneous T-cell lymphoma with hypofractionated radiation therapy in three cats.

BACKGROUND: Feline indolent cutaneous T-cell lymphoma (ICL) is an uncommon neoplastic disease. There is currently no consensus on treatment recommendations for ICL. OBJECTIVE: To report the clinical outcome of three cats with ICL treated with hypofractionated electron-beam radiotherapy (RT). ANIMALS: Three privately owned cats with ICL. MATERIALS AND METHODS: Medical records and client surveys were reviewed. A diagnosis of probable ICL was based on history, clinical presentation and histopathological findings, and confirmed using CD3 immunohistochemical analysis and PCR for antigen receptor gene rearrangement (PARR). All cats were treated with hypofractionated RT (four fractions of 8 Gy). RESULTS: All cats presented with skin lesions characterised by erythema and alopecia that were refractory to previous treatment with systemic glucocorticoids. Before hypofractionated RT treatment, lesions were histologically described as having diffuse infiltration of the dermis with CD3+ T cells. Molecular clonality analysis revealed clonal T-cell receptor gamma gene rearrangement. After RT, two cats showed histological improvement defined by decreased infiltration of lymphocytes, with cellular infiltrate present only in the deeper dermis; one cat had near complete histological resolution of lesions with only minimal residual lymphocytes. One cat was determined to have a complete clinical response while the other showed partial responses. No acute adverse effects of radiation were observed; chronic effects included leukotrichia, partial alopecia and mild fibrosis. All clients reported improvement in quality of life for their cats. CONCLUSIONS AND CLINICAL IMPORTANCE: Clinical and histological improvement in these cats suggests that hypofractionated RT can be a useful treatment modality for cats with ICL.

Is low-level laser therapy useful as an adjunctive treatment for canine acral lick dermatitis? A randomized, double-blinded, sham-controlled study.

BACKGROUND: Conventional therapy for canine acral lick dermatitis (ALD) consists of systemic antibiotics and anti-anxiety medications. Low-level laser therapy (LLLT) is a noninvasive therapy used to treat inflammatory and painful conditions. HYPOTHESIS/OBJECTIVES: The primary objective was to determine whether LLLT with conventional therapy would be beneficial as an adjunct treatment for ALD. We hypothesized that LLLT and conventional therapy combined would result in a greater reduction in licking Visual Analog Score (LVAS) compared to conventional therapy alone. Secondary objectives were to assess change in lesion/ulcer size, thickness and hair growth. ANIMALS: Thirteen dogs with a skin lesion consistent with ALD. METHODS AND MATERIALS: Dogs were randomly assigned to two groups. All dogs received systemic antibiotics and trazodone. The treatment group (TG) received LLLT by laser (130 mW, 2 min) with blue and red light-emitting diodes (LEDs), while the control group (CG) had sham therapy (laser/LEDs off). Treatments were administered three times weekly for two weeks, then twice weekly for two weeks for a total of 10 visits. Descriptive statistics were performed (mean, median); primary and secondary objectives were assessed with nonparametric ANOVA (Kruskal-Wallis test), with significance set at P < 0.05. RESULTS: Thirteen dogs (six CG, seven TG) were enrolled. There were no significant differences in median LVAS, lesion/ulcer size or thickness of the ALD lesion between TG and CG. There was a significantly greater increase (24%) in hair growth in TG (P = 0.0081) compared to CG. CONCLUSIONS AND CLINICAL RELEVANCE: Treatment of ALD requires multimodal therapy. Although combining LLLT with conventional therapy did not result in a significantly greater reduction in LVAS, there was a significant increase in hair growth compared to conventional therapy alone.

Antibody-Mediated Protection against Staphylococcus aureus Dermonecrosis: Synergy of Toxin Neutralization and Neutrophil Recruitment.

The staphylococcal α-hemolysin is critical for the pathogenesis of Staphylococcus aureus skin and soft tissue infection. Vaccine and infection-elicited α-hemolysin-specific antibodies protect against S. aureus‒induced dermonecrosis, a key feature of skin and soft tissue infection. Many interactions between α-hemolysin and host cells have been identified that promote tissue damage and modulate immune responses, but the mechanisms by which protective adaptive responses cross talk with innate responses at the tissue level are not clear. Using an established mouse model of skin and soft tissue infection and a newly developed histopathologic scoring system, we observed pathologic correlates early after infection, predicting protection against dermonecrosis by anti-α-hemolysin antibody. Protection was characterized by robust neutrophilic inflammation and compartmentalization of bacteria into discrete abscesses, which led to the attenuation of dermonecrosis and enhancement of bacterial clearance later in the infection. The ultimate outcome of infection was driven by the recruitment of neutrophils within the first day after infection but not later. Antibody-mediated protection was dependent on toxin neutralization rather than on enhanced opsonophagocytic killing by neutrophils or protection against toxin-mediated neutrophil lysis. Together, these findings advance our understanding of the mechanisms by which the early synergism between antibody-mediated toxin neutralization and tissue-specific neutrophilic inflammation preserve tissue integrity during infection.

The feasibility and utility of optical coherence tomography directed histopathology for surgical margin assessment of canine mast cell tumours.

Histopathologic surgical margin assessment in veterinary patients is an imprecise science with assessment limited to a small proportion of the surgical margin due to time and finances. Incomplete excision of canine mast cell tumours (MCTs) alters treatment recommendations and prognosis. Optical coherence tomography (OCT) is a novel imaging modality that has been reported in a single veterinary study for surgical margin assessment. Twenty-five dogs with 34 MCTs were enrolled in a prospective pilot-study to assess the imaging characteristics of canine MCTs with OCT and to evaluate the feasibility and utility of OCT-guided histopathology. All dogs underwent routine surgical excision of MCTs. OCT imaging was used to assess the entire surgical margin prior to placement in formalin. Either normal areas or areas suspected of incomplete MCT excision were inked. Standard histopathologic sectioning and tangential sectioning of inked areas were performed and compared to OCT results. OCT identified MCT near the surgical margin in 10 of 26 specimens (38.4%). Four specimens suspicious for incomplete margins on OCT had incomplete MCT excision that was missed on standard histopathologic sectioning. Six specimens had OCT-guided sections taken as suspicious, which did not show MCT on histopathology. OCT-guided pathology sections were able to detect incompletely excised MCT near the surgical margin with a sensitivity of 90% and specificity of 56.2% in this preliminary study. OCT imaging shows promise for guiding pathologists to areas of interest to improve the diagnostic accuracy of surgical margin assessment in excised canine MCTs.

Diagnostic accuracy of optical coherence tomography for assessing surgical margins of canine soft tissue sarcomas in observers of different specialties.

OBJECTIVE: To determine the diagnostic accuracy of optical coherence tomography (OCT) to assess surgical margins of canine soft tissue sarcoma (STS) and determine the influence of observer specialty and training. STUDY DESIGN: Blinded clinical prospective study. ANIMALS: Twenty-five dogs undergoing surgical excision of STS. METHODS: In vivo and ex vivo surgical margins were imaged with OCT after tumor resection. Representative images and videos were used to generate a training presentation and data sets. These were completed by 16 observers of four specialties (surgery, radiology, pathology, and OCT researchers). Images and videos from data sets were classified as cancerous or noncancerous. RESULTS: The overall sensitivity and specificity were 88.2% and 92.8%, respectively, for in vivo tissues and 82.5% and 93.3%, respectively, for ex vivo specimens. The overall accurate classification for all specimens was 91.4% in vivo and 89.5% ex vivo. There was no difference in accuracy of interpretation of OCT imaging by observers of different specialties or experience levels. CONCLUSION: Use of OCT to accurately assess surgical margins after STS excision was associated with a high sensitivity and specificity among various specialties. Personnel of all specialties and experience levels could effectively be trained to interpret OCT imaging. CLINICAL SIGNIFICANCE: Optical coherence tomography can be used by personnel of different specialty experience levels and from various specialties to accurately identify canine STS in vivo and ex vivo after a short training session. These encouraging results provide evidence to justify further research to assess the ability of OCT to provide real-time assessments of surgical margins and its applicability to other neoplasms.

Characterization of WWOX expression and function in canine mast cell tumors and malignant mast cell lines.

BACKGROUND: The WW domain-containing oxidoreductase (WWOX) tumor suppressor gene is frequently lost in a variety of solid and hematopoietic malignancies in humans. Dysregulation of WWOX has been implicated as playing a key role in tumor cell survival, DNA damage repair, and genomic stability. The purpose of this study was to characterize WWOX expression in spontaneous canine mast cell tumors (MCTs) and malignant cell lines and investigate the potential contribution of WWOX loss on malignant mast cell behavior. METHODS/RESULTS: WWOX expression is decreased in primary canine MCTs and malignant mast cell lines compared to normal canine bone marrow-cultured mast cells. In transformed canine mastocytoma cell lines, overexpression of WWOX or WWOX knockdown had no effect on mast cell viability. Inhibition of WWOX enhanced clonogenic survival following treatment with ionizing radiation in the C2 mast cell line. Lastly, immunohistochemistry for WWOX was performed using a canine MCT tissue microarray, demonstrating that WWOX staining intensity and percent of cells staining for WWOX is decreased in high-grade MCTs compared to low-grade MCTs. CONCLUSIONS: These data suggest that WWOX expression is attenuated or lost in primary canine MCTs and malignant mast cell lines. Given the observed increase in clonogenic survival in WWOX-deficient C2 mast cells treated with ionizing radiation, further investigation of WWOX and its role in mediating the DNA damage response in malignant mast cells is warranted.

Acquired left-to-right shunting through a valve-incompetent foramen ovale in a cat with hypertrophic cardiomyopathy and congestive heart failure.

CASE SUMMARY: An 8-year-old spayed female domestic shorthair cat was presented for a recheck evaluation of hypertrophic cardiomyopathy and chronic kidney disease. Three years prior to presentation, the patient was diagnosed with obstructive hypertrophic cardiomyopathy and started on atenolol. The left ventricular outflow tract obstruction subsequently resolved. Biochemical analysis a week prior to presentation demonstrated severe azotemia. Transthoracic echocardiograph revealed pericardial effusion, pleural effusion, severe left ventricular concentric hypertrophy, severe left atrial enlargement and continuous left-to-right flow through the interatrial septum near the fossa ovalis. The patient was euthanized owing to poor prognosis, and gross examination at necropsy revealed a valve-incompetent patent foramen ovale secondary to severe left atrial dilation. RELEVANCE AND NOVEL INFORMATION: To our knowledge, this is the first report of an acquired left-to-right shunt through a valve-incompetent foramen ovale in a cat with hypertrophic cardiomyopathy. Severe left atrial dilation was suspected to cause interatrial shunting through the valve-incompetent foramen ovale, and this finding may be relevant to echocardiographic evaluations in other cats.

Stable Attenuation of Human Respiratory Syncytial Virus for Live Vaccines by Deletion and Insertion of Amino Acids in the Hinge Region between the mRNA Capping and Methyltransferase Domains of the Large Polymerase Protein.

Human respiratory syncytial virus (RSV) is the leading viral cause of lower respiratory tract disease in infants and children worldwide. Currently, there are no FDA-approved vaccines to combat this virus. The large (L) polymerase protein of RSV replicates the viral genome and transcribes viral mRNAs. The L protein is organized as a core ring-like domain containing the RNA-dependent RNA polymerase and an appendage of globular domains containing an mRNA capping region and a cap methyltransferase region, which are linked by a flexible hinge region. Here, we found that the flexible hinge region of RSV L protein is tolerant to amino acid deletion or insertion. Recombinant RSVs carrying a single or double deletion or a single alanine insertion were genetically stable, highly attenuated in immortalized cells, had defects in replication and spread, and had a delay in innate immune cytokine responses in primary, well-differentiated, human bronchial epithelial (HBE) cultures. The replication of these recombinant viruses was highly attenuated in the upper and lower respiratory tracts of cotton rats. Importantly, these recombinant viruses elicited high levels of neutralizing antibody and provided complete protection against RSV replication. Taken together, amino acid deletions or insertions in the hinge region of the L protein can serve as a novel approach to rationally design genetically stable, highly attenuated, and immunogenic live virus vaccine candidates for RSV.IMPORTANCE Despite tremendous efforts, there are no FDA-approved vaccines for human respiratory syncytial virus (RSV). A live attenuated RSV vaccine is one of the most promising vaccine strategies for RSV. However, it has been a challenge to identify an RSV vaccine strain that has an optimal balance between attenuation and immunogenicity. In this study, we generated a panel of recombinant RSVs carrying a single and double deletion or a single alanine insertion in the large (L) polymerase protein that are genetically stable, sufficiently attenuated, and grow to high titer in cultured cells, while retaining high immunogenicity. Thus, these recombinant viruses may be promising vaccine candidates for RSV.

Comparative H-gal-GP and H11 specific antibody binding in equine cyathostomins.

The biological-based vaccine (Barbervax®) generates effective antibodies against the biologically essential H-gal-GP and H11 protein complex of the ruminant parasite Haemonchus contortus to target and kill the parasites after taking a blood meal. A comparative analysis of several parasite genera was performed to determine if a similar protein complex or one that is recognized by H-gal-GP and H11 specific antibodies was present. If so, it suggests the vaccine could be effective for other nematode parasites. Ancylostoma caninum, H. contortus, equine cyathostomins, bovine Bunostomum phlebotomum, Dracunculus lutrae, Parascaris sp., Ixodes scapularis, Amblyomma americanum, Dirofilaria immitis and Brugia malayi were evaluated for specific antibody binding using hyperimmunized antibodies against H-gal-GP and H11 native proteins. Of the parasites evaluated, specific and reproducible staining was observed in H. contortus and adult and encysted cyathostomins only. To further evaluate the similar reactivities between cyathostomins and H. contortus, cross-reactivity of equine serum with antibodies to cyathostomins on a H. contortus adult histology cross-section was observed using immunofluorescence. These findings pave the way for future studies on the safety and efficacy of H-gal-GP and H11 protein complex as a potential control for cyathostomins.

Evaluation of microwave ablation for local treatment of dogs with distal radial osteosarcoma: A pilot study.

OBJECTIVE: To evaluate the feasibility and safety of microwave ablation (MWA) as a modality to induce tumor necrosis within distal radial osteosarcoma (OSA). STUDY DESIGN: Pilot study. ANIMALS: Six client-owned dogs with distal radius OSA confirmed by cytological examination. METHODS: Dogs underwent computed tomography for surgical planning before general anesthesia for fluoroscopy-guided ablation. Computed tomography was repeated 48 hours after MWA, before amputation. The ablated tumor was evaluated with histopathology. RESULTS: Six dogs underwent MWA of distal radius OSA. A lower power setting (30 W) was selected for the first two dogs to avoid collateral soft tissue damage. The power was increased to 75 W for the last four dogs. The temperature was maintained between 45°C and 55°C (113 °F-131 °F) at the bone/soft tissue interface. Tumor necrosis varied between 30% and 90% (median, 55%) according to histopathology. No intraoperative or periprocedural complications were observed. CONCLUSION: Microwave ablation induced variable tumor necrosis and did not induce immediate postablation complications in these six dogs with distal radius OSA. CLINICAL SIGNIFICANCE: These results justify further evaluation of MWA as a potential modality to treat primary bone lesions in dogs.

Pharmacokinetics and Tolerability of the Novel Non-immunosuppressive Fingolimod Derivative, OSU-2S, in Dogs and Comparisons with Data in Mice and Rats.

In this study, we characterized the pharmacokinetics of OSU-2S, a fingolimod-derived, non-immunosuppressive phosphatase activator, in mice, rats, and dogs, as well as tolerability and food effects in dogs. Across all species tested, plasma protein binding for OSU-2S was > 99.5%, and metabolic stability and hepatic intrinsic clearance were in the moderate range. OSU-2S did not significantly modulate CYP enzyme activity up until 50 µM, and Caco-2 data suggested low permeability with active efflux at 2 µM. Apparent oral bioavailability in mice was 16% and 69% at 10 and 50 mg/kg, respectively. In rats, bioavailability was 24%, 35%, and 28% at 10, 30, and 100 mg/kg, respectively, while brain/plasma ratio was 36 at 6-h post-dose at 30 mg/kg. In dogs, OSU-2S was well tolerated with oral capsule bioavailability of 27.5%. Plasma OSU-2S exposures increased proportionally over a 2.5-20 mg/kg dose range. After 4 weeks of 3 times weekly, oral administration (20 mg/kg), plasma AUClast (26.1 µM*h), and Cmax (0.899 µM) were nearly 2-fold greater than those after 1 week of dosing, and no food effects were observed. The elimination half-life (29.7 h), clearance (22.9 mL/min/kg), and plasma concentrations of repeated oral doses support a 3-times weekly dosing schedule in dogs. No significant CBC, serum biochemical, or histopathological changes were observed. OSU-2S has favorable oral PK properties similar to fingolimod in rodents and dogs and is well tolerated in healthy animals. This work supports establishing trials of OSU-2S efficacy in dogs with spontaneous tumors to guide its clinical development as a cancer therapeutic for human patients.

Continuous Microevolution Accelerates Disease Progression during Sequential Episodes of Infection.

Bacteria adapt to dynamic changes in the host during chronic and recurrent infections. Bacterial microevolution is one type of adaptation that imparts a selective advantage. We hypothesize that recurrent episodes of disease promote microevolution through genetic mutations that modulate disease severity. We use a pre-clinical model of otitis media (OM) to determine the potential role for microevolution of nontypeable Haemophilus influenzae (NTHI) during sequential episodes of disease. Whole genome sequencing reveals microevolution of hemoglobin binding and lipooligosaccharide (LOS) biosynthesis genes, suggesting that adaptation of these systems is critical for infection. These OM-adapted strains promote increased biofilm formation, inflammation, stromal fibrosis, and an increased propensity to form intracellular bacterial communities (IBCs). Remarkably, IBCs remain for at least one month following clinical resolution of infection, suggesting an intracellular reservoir as a nidus for recurrent OM. Additional approaches for therapeutic design tailored to combat this burdensome disease will arise from these studies.