Determination of the reference interval for urine kidney injury molecule-1 in 50 healthy cats.

OBJECTIVES: The aim of the present study was to establish a reference interval (RI) for urine kidney injury molecule-1 (KIM-1) in healthy cats. METHODS: History, physical examination, blood pressure, and feline immunodeficiency virus and feline leukemia virus serology status were determined. A complete blood cell count, serum biochemical profile, urinalysis and kidney ultrasound were performed, and N-terminal pro-brain natriuretic peptide, total thyroxine (TT4) and urine KIM-1 were measured. An RI was calculated and the effect of age, sex, body condition score (BCS), blood pressure, symmetric dimethylarginine (SDMA), serum creatinine concentration (SCr), phosphorus, TT4, urine specific gravity (USG) and mid-sagittal kidney length on urine KIM-1 was evaluated using a general linear model. RESULTS: Of 69 recruited cats, 50 met the inclusion criteria. There were 35 male cats and 15 female cats, with a median age of 4.3 years (range 1.0-12.3), median weight of 5.11 kg (range 2.52-8.45) and median BCS of 6/9 (range 3-8). The median serum concentrations were SDMA 11.0 µg/dl (range 2-14), SCr 88.5 µmol/l (range 47-136), phosphorus 1.41 mmol/l (range 0.8-2.2) and TT4 32.0 nmol/l (range 17-51). Median USG was 1.057 (range 1.035-1.076), mid-sagittal left kidney length was 3.50 cm (range 2.94-4.45) and mid-sagittal right kidney length was 3.70 cm (range 3.06-4.55). The derived RI for urine KIM-1 was 0.02-0.68. USG was a significant (P <0.001) predictor of urine KIM-1. Individually, age, sex, blood pressure, BCS, SDMA, SCr, phosphorus, TT4 and mid-sagittal kidney length were not significant predictors of urine KIM-1. In a multivariate model, if combined with USG, SDMA concentration was predictive (P = 0.030) of urine KIM-1. CONCLUSIONS AND RELEVANCE: Urine concentration was significantly correlated with urine KIM-1, which will be an important consideration when interpreting findings in cats with potential kidney injury.

Whole-genome sequencing of SARS-CoV-2 from the initial cases of domestic cat infections in Canada.

Two cat nasal swabs from Canada's earliest confirmed SARS-CoV-2 positive domestic cats were sequenced to over 99% SARS-CoV-2 genome coverage. One cat had lineage A.23.1 SARS-CoV-2 not reported before in animals. Both sequences have multiple spike gene mutations and clustered closely with human-derived sequences in the global SARS-CoV-2 phylogenetic tree.

Transcriptomic Profiles of Pectoralis major Muscles Affected by Spaghetti Meat and Woody Breast in Broiler Chickens.

Spaghetti meat (SM) and woody breast (WB) are breast muscle myopathies of broiler chickens, characterized by separation of myofibers and by fibrosis, respectively. This study sought to investigate the transcriptomic profiles of breast muscles affected by SM and WB. Targeted sampling was conducted on a flock to obtain 10 WB, 10 SM, and 10 Normal Pectoralis major muscle samples from 37-day-old male chickens. Total RNA was extracted, cDNA was used for pair-end sequencing, and differentially expressed genes (DEGs) were determined by a false discovery rate of <0.1 and a >1.5-fold change. Principal component and heatmap cluster analyses showed that the SM and WB samples clustered together. No DEGs were observed between SM and WB fillets, while a total of 4018 and 2323 DEGs were found when comparing SM and WB, respectively, against Normal samples. In both the SM and WB samples, Gene Ontology terms associated with extracellular environment and immune response were enriched. The KEGG analysis showed enrichment of cytokine-cytokine receptor interaction and extracellular matrix-receptor interaction pathways in both myopathies. Although SM and WB are macroscopically different, the similar transcriptomic profiles suggest that these conditions may share a common pathogenesis. This is the first study to compare the transcriptomes of SM and WB, and it showed that, while both myopathies had profiles different from the normal breast muscle, SM and WB were similar, with comparable enriched metabolic pathways and processes despite presenting markedly different macroscopic features.

Evaluation of SARS-CoV-2 identification methods through surveillance of companion animals in SARS-CoV-2-positive homes in North Carolina, March to December 2020.

We collected oral and/or rectal swabs and serum from dogs and cats living in homes with SARS-CoV-2-PCR-positive persons for SARS-CoV-2 PCR and serology testing. Pre-COVID-19 serum samples from dogs and cats were used as negative controls, and samples were tested in duplicate at different timepoints. Raw ELISA results scrutinized relative to known negative samples suggested that cut-offs for IgG seropositivity may require adjustment relative to previously proposed values, while proposed cut-offs for IgM require more extensive validation. A small number of pet dogs (2/43, 4.7%) and one cat (1/21, 4.8%) were positive for SARS-CoV-2 RNA, and 28.6 and 37.5% of cats and dogs were positive for anti-SARS-CoV-2 IgG, respectively.

Discordant FeLV p27 immunoassay and PCR test results in 21 cats with hematologic disorders.

CASE SERIES SUMMARY: A total of 1692 medical records from a primary care feline practice and a veterinary referral hospital were evaluated retrospectively to assess discordant feline leukemia virus (FeLV) test results. In total, 73 cats were positive for FeLV using serum in a lateral flow immunoassay (LFI) or laboratory-based ELISA. Of these cats, 21 subsequently tested negative for FeLV proviral DNA by non-quantitative PCR on EDTA whole blood (16/21, 76.2%), bone marrow (4/21, 19%) or both (1/21, 4.7%). The proportional morbidity (an estimate of prevalence in a sample of the total population) for FeLV by LFI/ELISA and PCR assays was 3.1%, consistent with that reported in previous studies for cats in North America. Cats with discordant LFI/ELISA and PCR results had either primary bone marrow disease (18 autoimmune, one neoplastic), a bone marrow insult (hemotrophic mycoplasmosis) or systemic inflammation (pyothorax with a marked neutrophilic leukocytosis). The percentage of cats with a positive LFA/ELISA result and negative PCR assay surviving to discharge was 85.7% (18/21). Of these, 88.9% (16/18) survived 4 months to 6 years. Seven cats (33.3%) were re-tested with LFI or ELISA once primary disease was controlled, and all tested negative. RELEVANCE AND NOVEL INFORMATION: These findings indicate that in cats with bone marrow disease that shares features of progressive FeLV infection, positive LFI and ELISA FeLV test results should be followed up with FeLV proviral DNA PCR testing, particularly in populations where disease prevalence is low.

Quantification of circulating tumour cells over time in dogs with appendicular osteosarcoma.

Enumeration of circulating tumour cells (CTC) has shown promise for prognostication and guidance of therapeutic decisions in human cancers. The objective of this study was to enumerate CTC over time in dogs with naturally occurring osteosarcoma (OSA), and to determine correlation with patient outcome. Twenty-six dogs with OSA and no evidence of metastatic disease at the time of amputation were enrolled. Dogs were assessed for lung metastases and CTC prior to and following amputation, and at each chemotherapy visit. Twenty-one dogs completed the study. Nineteen dogs were euthanized and two were alive and free of metastases. Overall survival time ranged from 88 to 1058 days (median survival time (MST) 374 days). Increased serum alkaline phosphatase activity, advanced age, and higher body weight were significantly associated with lower MST. Dogs with OSA had a mean of 356 (0 to 4443) CTC/106 leukocytes. In 12 of 15 dogs that developed radiographic evidence of metastasis, a pre-metastatic CTC spike was retrospectively detectable on average 36.5 (1-100 days) days prior to metastasis and was associated with significantly shorter MST (301 ± 64 vs. 626 ± 55 days; p = .0107). In a multivariable analysis, dogs with a CTC spike were 10× more likely to die compared with those without. These results suggest that a spike in CTC frequency precedes detection of metastasis in dogs with OSA and is associated with shorter survival. More frequent enumeration of CTC in a larger cohort of dogs with OSA may be warranted.

Diagnosis and management of a peripheral T-cell lymphoproliferative disease in an American pine marten (Martes americana).

A 9-year-old neutered male American pine marten (Martes americana) was referred for further evaluation of suspected lymphoproliferative disease. On physical examination, the pine marten was determined to be in an underconditioned state with an enlarged right mandibular lymph node. Hematology revealed a marked leukocytosis characterized by a lymphocytosis. Flow cytometry performed on peripheral blood was suggestive of a CD4+ T-cell lymphoproliferative disease. Whole-body radiographs demonstrated a large cranial mediastinal mass and splenomegaly. These findings were confirmed using ultrasound, which also identified intra-abdominal lymphadenopathy and splenic nodules. Cytologic evaluation of aspirates from the mediastinal mass was interpreted as possible lymphoma. The pine marten was treated with chlorambucil and prednisolone and achieved a durable partial remission. Twelve months after initial diagnosis, progressive disease was noted and treatment with lomustine was initiated as a rescue protocol until euthanasia, which was carried out 15 mo after the initial diagnosis. Based on a literature search, this is the first case report describing the management of peripheral T-cell lymphoproliferative disease, presumably peripheral lymphoma, in a pine marten; this neoplasm should be considered as a differential diagnosis in pine martens that have abnormal complete blood cell count findings and enlarged lymph nodes. Key clinical message: This report describes the diagnosis and management of a peripheral T-cell lymphoproliferative disease, presumably peripheral lymphoma, in an American pine marten (Martes americana). This is the first report of this disease and its successful treatment in a pine marten.

Diagnostic et prise en charge d'une maladie lymphoproliférative à cellules T périphériques chez une martre d'Amérique ( Martes americana ). Une martre d'Amérique (Martes americana) mâle castré âgé de 9 ans a été référée pour une évaluation plus approfondie d'une suspicion de maladie lymphoproliférative. À l'examen physique, il a été déterminé que la martre était dans un état sous-optimal avec un ganglion lymphatique mandibulaire droit élargi. L'hématologie a révélé une hyperleucocytose marquée caractérisée par une lymphocytose. La cytométrie en flux réalisée sur le sang périphérique était évocatrice d'une maladie lymphoproliférative des lymphocytes T CD4+. Les radiographies du corps entier ont montré une importante masse médiastinale crânienne et une splénomégalie. Ces résultats ont été confirmés par échographie, qui a également identifié une lymphadénopathie intra-abdominale et des nodules spléniques. L'évaluation cytologique des aspirations de la masse médiastinale a été interprétée comme un possible lymphome. La martre d'Amérique a été traitée avec du chlorambucil et de la prednisolone et une rémission partielle durable a été obtenue. Douze mois après le diagnostic initial, une progression de la maladie a été notée et un traitement par lomustine a été initié comme protocole de sauvetage jusqu'à l'euthanasie, qui a été réalisée 15 mois après le diagnostic initial. Sur la base d'une recherche documentaire, il s'agit du premier rapport de cas décrivant la prise en charge d'une maladie lymphoproliférative périphérique à cellules T, vraisemblablement un lymphome périphérique, chez une martre d'Amérique; ce néoplasme doit être considéré comme un diagnostic différentiel chez les martres d'Amérique qui présentent des résultats anormaux de numération globulaire complète et des ganglions lymphatiques hypertrophiés.Message clinique clé :Ce rapport décrit le diagnostic et la prise en charge d'une maladie lymphoproliférative à cellules T périphériques, vraisemblablement un lymphome périphérique, chez une martre d'Amérique (Martes americana). Il s'agit du premier signalement de cette maladie et de son traitement réussi chez une martre d'Amérique.(Traduit par Dr Serge Messier).

Equine alveolar macrophages and monocyte-derived macrophages respond differently to an inflammatory stimulus.

Alveolar macrophages (AMs) are the predominant innate immune cell in the distal respiratory tract. During inflammatory responses, AMs may be supplemented by blood monocytes, which differentiate into monocyte-derived macrophages (MDMs). Macrophages play important roles in a variety of common equine lower airway diseases, including severe equine asthma (SEA). In an experimental model, an inhaled mixture of Aspergillus fumigatus spores, lipopolysaccharide, and silica microspheres (FLS), induced SEA exacerbation in susceptible horses. However, whether equine AMs and MDMs have differing immunophenotypes and cytokine responses to FLS stimulation is unknown. To address these questions, alveolar macrophages/monocytes (AMMs) were isolated from bronchoalveolar lavage fluid and MDMs derived from blood of six healthy horses. Separately, AMMs and MDMs were cultured with and without FLS for six hours after which cell surface marker expression and cytokine production were analyzed by flow cytometry and a bead-based multiplex assay, respectively. Results showed that regardless of exposure conditions, AMMs had significantly higher surface expression of CD163 and CD206 than MDMs. Incubation with FLS induced secretion of IL-1ß, IL-8, TNF-α and IFN-γ in AMMs, and IL-8, IL-10 and TNF-α in MDMs. These results suggest that AMMs have a greater proinflammatory response to in vitro FLS stimulation than MDMs, inferring differing roles in equine lung inflammation. Variability in recruitment and function of monocyte-macrophage populations warrant more detailed in vivo investigation in both homeostatic and diseased states.

Treatment of myeloid neoplasia with doxorubicin and cytarabine in 11 dogs.

Myelodysplastic syndrome (MDS) and acute myeloid leukaemia (AML) are primary myeloid neoplasms in dogs generally considered to have a poor outcome. In this study, we assessed toxicity, efficacy and outcome of concurrent administration of doxorubicin and cytarabine in 11 dogs with myeloid neoplasia. Bone marrow specimens were reviewed by three pathologists and classified as either MDS (n = 2), high grade MDS/early AML (MDS/AML; n = 4) or AML (n = 5). The median number of treatment cycles was 5 (range 1-9) and resolution of cytopenia was reported in 7 of 11 dogs including 2 dogs with MDS, 2 dogs with MDS/AML, and 3 dogs with AML. The median duration of remission in the seven responders was 344 days (range 109-1428) and the median overall survival for all dogs was 369 days. Adverse events consisted of predominantly low-grade gastrointestinal illness and myelosuppression. Three dogs developed grade V toxicity manifesting with heart failure (n = 2) at 369 and 1170 days after diagnosis and acute gastrointestinal side effects (n =1). Despite a limited sample size, these results suggest that a doxorubicin and cytarabine protocol may be considered as a therapeutic option in dogs with myeloid neoplasia. Protocol safety, in particular regarding myocardial toxicity, and efficacy should be further investigated.

Effect of up to 30-days of storage at different temperatures on detection of feline kidney injury molecule-1 in urine.

BACKGROUND: In humans, kidney injury molecule-1 (KIM-1) is a biomarker of acute kidney injury that can be quantified in urine. Preliminary investigation in cats with experimentally induced acute kidney injury showed that KIM-1 urine concentration correlated with kidney injury histopathology scores. A lateral flow assay (LFA) has recently become available for patient-side feline KIM-1 measurement. In vitro parameters of the assay have not yet been determined. The objectives of this study were to determine detection of KIM-1 in urine stored at different temperatures over time, to establish the linear range of the LFA, and to assess the intra-assay repeatability of measurements.  RESULTS: Ten urine samples with a range of KIM-1 concentrations were stored at room temperature (22o C), 4o C or -20o C, and tested with the LFA on days 0, 1, 2, 3, 7, 14, and 30. The concentration of KIM-1 in samples was not significantly different from the day 0 value, except one sample that had been stored for 30 days at room temperature yielded a significantly higher value. The assay results had a correlation coefficient of 0.922. The mean coefficient of variation for all samples was 15.7%. The slope of the curve of expected versus measured values in samples diluted two-fold nine times was 0.908, and results were linear over all dilutions. CONCLUSIONS: The LFA for feline KIM-1 yields consistent results from stored urine samples. These characteristics will allow for KIM-1 to be measured retrospectively if immediate testing is not feasible. Within assay precision was high, and linearity over 9 logs of dilution suggests suitability for a range of subclinical and clinical kidney injuries.

Cyclic hematopoiesis in a mixed-breed dog: case report and brief review.

An 8-wk-old, male, mixed-breed puppy was adopted from a rescue organization. From the time of adoption, the puppy suffered episodes of illness affecting various organ systems, which resolved with supportive therapy but relapsed once medical therapy was discontinued. Review of the hematologic data revealed cyclic fluctuations in circulating blood cells. Cyclicity was most prominent in neutrophils, with recurrent severe neutropenia. Neutropenic episodes lasted 5-6 d, with regular cycles of 11-14 d between nadir neutrophil counts. Genetic testing determined that the patient was homozygous mutant for the frameshift mutation in the adaptor protein complex 3 ß-subunit (AP3B1) gene, originally identified in gray collies with cyclic hematopoiesis (CH). Pedigree information was not available, but the patient's features were phenotypically distinct from those of collies. We describe here a case of the AP3B1 mutation in a mixed-breed dog that did not resemble a collie, undescribed previously, to our knowledge. Our findings indicate that the AP3B1 mutation and CH are present within the general canine population and are not restricted to collies.

Inter-species cell detection - datasets on pulmonary hemosiderophages in equine, human and feline specimens.

Pulmonary hemorrhage (P-Hem) occurs among multiple species and can have various causes. Cytology of bronchoalveolar lavage fluid (BALF) using a 5-tier scoring system of alveolar macrophages based on their hemosiderin content is considered the most sensitive diagnostic method. We introduce a novel, fully annotated multi-species P-Hem dataset, which consists of 74 cytology whole slide images (WSIs) with equine, feline and human samples. To create this high-quality and high-quantity dataset, we developed an annotation pipeline combining human expertise with deep learning and data visualisation techniques. We applied a deep learning-based object detection approach trained on 17 expertly annotated equine WSIs, to the remaining 39 equine, 12 human and 7 feline WSIs. The resulting annotations were semi-automatically screened for errors on multiple types of specialised annotation maps and finally reviewed by a trained pathologist. Our dataset contains a total of 297,383 hemosiderophages classified into five grades. It is one of the largest publicly available WSIs datasets with respect to the number of annotations, the scanned area and the number of species covered.

The Pathobiology of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome: The Case for Neuroglial Failure.

Although myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) has a specific and distinctive profile of clinical features, the disease remains an enigma because causal explanation of the pathobiological matrix is lacking. Several potential disease mechanisms have been identified, including immune abnormalities, inflammatory activation, mitochondrial alterations, endothelial and muscular disturbances, cardiovascular anomalies, and dysfunction of the peripheral and central nervous systems. Yet, it remains unclear whether and how these pathways may be related and orchestrated. Here we explore the hypothesis that a common denominator of the pathobiological processes in ME/CFS may be central nervous system dysfunction due to impaired or pathologically reactive neuroglia (astrocytes, microglia and oligodendrocytes). We will test this hypothesis by reviewing, in reference to the current literature, the two most salient and widely accepted features of ME/CFS, and by investigating how these might be linked to dysfunctional neuroglia. From this review we conclude that the multifaceted pathobiology of ME/CFS may be attributable in a unifying manner to neuroglial dysfunction. Because the two key features - post exertional malaise and decreased cerebral blood flow - are also recognized in a subset of patients with post-acute sequelae COVID, we suggest that our findings may also be pertinent to this entity.

Risk Factors for SARS-CoV-2 Infection and Illness in Cats and Dogs1.

We tested swab specimens from pets in households in Ontario, Canada, with human COVID-19 cases by quantitative PCR for SARS-CoV-2 and surveyed pet owners for risk factors associated with infection and seropositivity. We tested serum samples for spike protein IgG and IgM in household pets and also in animals from shelters and low-cost neuter clinics. Among household pets, 2% (1/49) of swab specimens from dogs and 7.7% (5/65) from cats were PCR positive, but 41% of dog serum samples and 52% of cat serum samples were positive for SARS-CoV-2 IgG or IgM. The likelihood of SARS-CoV-2 seropositivity in pet samples was higher for cats but not dogs that slept on owners' beds and for dogs and cats that contracted a new illness. Seropositivity in neuter-clinic samples was 16% (35/221); in shelter samples, 9.3% (7/75). Our findings indicate a high likelihood for pets in households of humans with COVID-19 to seroconvert and become ill.

Effects of equine SALSA on neutrophil phagocytosis and macrophage cytokine production.

Salivary scavenger and agglutinin (SALSA) is a secreted protein with various immunomodulatory roles. In humans, the protein agglutinates and inactivates microorganisms, and inhibits the release of pro-inflammatory cytokines. Saliva, which is rich in SALSA, accelerates bacterial phagocytosis, but SALSA's contribution is unclear. In horses, the functions of SALSA in inflammation remain undetermined, so they were investigated through phagocytosis and cytokine assays. Equine SALSA was purified from duodenal tissue, which contains abundant SALSA. To assess phagocytosis, fluorescently-labelled bacteria were incubated with 20, 10, 5, or 2.5 µg/mL of SALSA or phosphate buffered saline (PBS), and then incubated at 37°C or on ice with whole blood from seven healthy horses. Fluorescence was measured by gating on neutrophils using a flow cytometer, and compared between groups. To assess effects on cytokine production, alveolar macrophages were isolated from bronchoalveolar lavage fluid of five healthy horses and cultured in serum-free media for 24 hours with different concentrations of SALSA plus 1 µg/mL lipopolysaccharide (LPS), only LPS, or only media. Cytokines were measured in supernatant using an equine-specific multiplex bead immunoassay. There was significantly greater phagocytosis in samples incubated at 37°C compared to incubation on ice. Samples incubated with 20 µg/mL of SALSA at 37°C had less phagocytosis compared to samples with 10 or 2.5 µg/mL SALSA, or PBS. Alveolar macrophages incubated with SALSA plus LPS released significantly less CXC motif chemokine ligand 1, interleukin-8, interleukin-10, and tumor necrosis factor α, and more granulocyte colony stimulating factor (G-CSF), compared to macrophages incubated with LPS alone. These findings indicate anti-inflammatory effects, which may be due to interference with toll-like receptor 4 recognition of LPS or downstream signaling. Increase in G-CSF following incubation with SALSA suggests a novel mechanism for immunoregulation of alveolar macrophages by SALSA, addressing a knowledge gap regarding its functions in horses.

RNA-Seq analysis of gene expression in 25 cases of canine lymphoma undergoing CHOP chemotherapy.

OBJECTIVES: Canine lymphoma, the most common hematological cancer in dogs, shares many molecular and clinical characteristics with human Non-Hodgkin lymphoma (NHL). The standard treatment for canine lymphoma is "CHOP" multiagent chemotherapy protocol consisting of Cyclophosphamide, Doxorubicin (Hydroxydaunorubicin), Vincristine (Oncovin™), and Prednisone. Approximately 70-85% of patients treated with CHOP achieve clinical remission. However, duration of remission varies and the majority of dogs eventually relapse. To identify possible biomarkers for patients failing to achieve remission, we performed RNA-Seq analysis on 25 cases of canine lymphoma obtained prior the start of their CHOP therapy regime and assessed gene expression associated with patient progression free survival (PFS). DATA DESCRIPTION: The data consists of (1) raw RNA-Seq reads in 75 bp fastq format from fine needle aspirate samples of enlarged lymph nodes from canine patients with naturally occurring lymphoma; (2) Fragments Per Kilobase Million (FPKM) values for each sample; (3) raw transcript counts for each sample; (4) anonymized patient details including PFS; (5) heat map of gene expression and (6) Cox proportional hazard analysis showing significantly expressed genes. These data may be useful for comparative analysis of gene expression in human NHL and analysis of gene expression associated with disease outcome in canine lymphoma.

Characteristics of broiler chicken breast myopathies (spaghetti meat, woody breast, white striping) in Ontario, Canada.

Spaghetti meat (SM), woody breast (WB), and white striping (WS) are myopathies affecting breast muscle of broiler chickens, and are characterized by a loss of myofibers and an increase in fibrous tissue. The conditions develop in intensive broiler chicken production systems, and cause poor meat process-ability and negative customer perception leading to monetary losses. The objectives of the present study were to describe the physical and histological characteristics of breast myopathies from commercial broiler chicken flocks in Ontario, Canada, and to assess the associations between the severity of myopathies with the physical and histological characteristics of the affected breast muscle fillets. Chicken breast fillets (n = 179) were collected over 3 visits from a processing plant and scored macroscopically to assess the severity of myopathies, following an established scoring scheme. For each fillet, the surface area, length, width, thickness, weight, and hardness (compression force) were measured. A subset of 60 fillets was evaluated microscopically. Multinomial logistic regression models were built to evaluate associations between physical parameters and macroscopic scores. The odds of SM co-occurring with severe WB (SM1WB2) were significantly associated with increased fillet thickness (OR = 1.59, 95% CI 1.31-1.94) and weight (OR = 1.06, 95% CI 1.03-1.09). Histologically, myopathies had overlapping lesions consisting of polyphasic myodegeneration, perivascular inflammatory cuffing and accumulation of fibrous tissue and fat. The pairwise correlation between macroscopic and microscopic scores was moderate (rho 0.45, P < 0.001). This is the first study to characterize breast myopathies in Canadian broiler flocks. Results show that the morphologic and microscopic changes of fillets from this cohort are similar to data from other countries, and provide database to benchmark these parameters in future studies. Our standardized categorization can be applied to broiler breast fillets in other regions of the world.

Effect of intrabronchial administration of autologous adipose-derived mesenchymal stem cells on severe equine asthma.

BACKGROUND: Severe equine asthma (SEA) is a common chronic respiratory disease and a significant health and well-being problem in horses. Current therapeutic strategies improve pulmonary function and clinical signs in some horses, but in the long-term, return to full athletic function appears to be rare. The aim of this study was to assess the safety and the effect of intrabronchial administration of adipose-derived mesenchymal stem cells (AD-MSC) on pulmonary inflammatory and clinical parameters in horses with SEA. METHODS: This was a randomized controlled trial. Twenty adult horses diagnosed with SEA were randomly divided into two groups (n = 10), and treated either with a single intrabronchial application of autologous AD-MSC or oral dexamethasone for three weeks. A targeted clinical examination with determination of clinical score, maximal change in pleural pressure during the breathing cycle, and an endoscopic examination of the airways were performed at baseline and three weeks after treatment. Bronchoalveolar lavage fluid was analyzed cytologically, and IL-1ß, IL-4, IL-8, IL-17, TNFα and IFNγ mRNA and protein concentrations were measured at baseline and three weeks. The horses were then monitored over one year for recurrence of SEA. A non-inferiority analysis and a linear mixed-effects model were performed to assess differences between treatments. RESULTS: The non-inferiority of AD-MSC treatment was not established. However, AD-MSC administration significantly ameliorated the clinical score (P = 0.01), decreased the expression of IL-17 mRNA (P = 0.05) and IL-1ß (P ≤ 0.001), IL-4 (P ≤ 0.001), TNFα (P = 0.02) protein levels, and had a positive long-term effect on SEA-associated clinical signs (P = 0.02). CONCLUSIONS: Intrabronchial administration of AD-MSC had limited short-term anti-inflammatory effects but improved the clinical signs of SEA at one year.

Flow cytometric analysis of equine bronchoalveolar lavage fluid cells in horses with and without severe equine asthma.

Severe equine asthma (SEA) is a common, debilitating lower airway inflammatory disorder of older horses. Alveolar macrophages (AMs) survey inhaled particulates from barn sources causing them to switch from an anti-inflammatory to a proinflammatory phenotype, resulting in neutrophil recruitment to the lung. This proinflammatory switch may contribute to the development and prolongation of SEA. Validated antibodies to identify the cells involved in the pathogenesis of SEA are lacking. In this study, monoclonal antibodies against CD90, CD163, and CD206 were tested for reactivity with equine leukocytes by immunocytochemistry and flow cytometry. A multi-color flow cytometric assay was developed to identify leukocytes in equine bronchoalveolar lavage fluid (BALF). Four control and 4 SEA-susceptible horses had BALF collected before and after a 48-hour moldy hay challenge. Antibodies against CD90 uniquely labeled equine neutrophils, and antibodies against CD163 and CD206 identified equine macrophages. Postchallenge AM surface expression of CD163 increased in both groups of horses, but the increase was statistically significant in only the SEA-susceptible group (P = .02). The surface expression of CD206 on AMs increased significantly in the SEA-susceptible group (P = .03) but was unchanged in the control group (P = .5). Increased expression of CD163 and CD206 during exacerbation of SEA suggested an association between AM phenotype and lung inflammation. However, functions of AMs in the pathogenesis of SEA remain to be elucidated.

Bronchial brush cytology, endobronchial biopsy, and SALSA immunohistochemistry in severe equine asthma.

Horses with severe equine asthma (SEA), also known as heaves and recurrent airway obstruction, have persistent neutrophilic inflammation of the lower airways. Cytologic evaluation of bronchoalveolar lavage (BAL) fluid is commonly used to confirm the clinical diagnosis of SEA. However, the utility of microscopic assessment of bronchial brushings, endobronchial biopsies, and immunohistochemical detection of disease-associated biomarkers for the diagnosis of SEA remain poorly characterized. Salivary scavenger and agglutinin (SALSA) has anti-inflammatory properties and downregulated gene expression in SEA; therefore, it was investigated as a tissue biomarker for airway and systemic inflammation. Six asthmatic and 6 non-asthmatic horses were exposed to an inhaled challenge. Before and after challenge, samples of BAL fluid, bronchial brushing, and endobronchial biopsy were collected. Location of SALSA in biopsies was determined, and immunohistochemical label intensity was computed using image analysis software. Serum amyloid A (SAA) was measured to assess systemic inflammation. After challenge, neutrophil proportions were significantly higher in asthmatic versus non-asthmatic horses in BAL fluid (least squares means, 95% confidence interval: 80.9%, 57.2% to 93.1%, vs 3.6%, 1.1% to 10.7%) and in brush cytology slides (39.5%, 7.7% to 83.6%, vs 0.2%, 0% to 2.3%), illustrating the potential of brush cytology as an alternate modality to BAL for assessing intraluminal inflammation. Bronchial histopathologic findings and intensity of SALSA immunolabeling in surface and glandular epithelium were similar in asthmatic and non-asthmatic horses, indicating limited changes in bronchial tissue from the inhaled challenge. Increases in SAA indicated systemic inflammation, but SALSA immunolabeling did not change significantly.