Effects of low-dust forages on dust exposure, airway cytology, and plasma omega-3 concentrations in Thoroughbred racehorses: A randomized clinical trial.

BACKGROUND: Racehorses commonly develop evidence of mild asthma in response to dust exposure. Diets deficient in omega-3 polyunsaturated fatty acids (Ω-3) might exacerbate this response. HYPOTHESIS: To compare dust exposure, bronchoalveolar lavage fluid (BALF) cytology, and plasma Ω-3 and specialized pro-resolving mediators (SPM) concentrations amongst racehorses fed dry hay, steamed hay, and haylage. ANIMALS: Forty-three Thoroughbred racehorses. METHODS: Prospective clinical trial. Horses were randomly assigned to be fed dry hay, steamed hay, or haylage for 6 weeks. Measures of exposure to dust in the breathing zone were obtained twice. At baseline, week-3, and week-6, BALF cytology was examined. Plasma lipid profiles and plasma SPM concentrations were examined at baseline and week 6. Generalized linear mixed models examined the effect of forage upon dust exposure, BALF cytology, Ω-3, and SPM concentrations. RESULTS: Respirable dust was significantly higher for horses fed hay (least-square mean ± s.e.m. 0.081 ± 0.007 mg/m3 ) when compared with steamed hay (0.056 ± 0.005 mg/m3 , P = .01) or haylage (0.053 ± 0.005 mg/m3 , P < .01). At week 6, BALF neutrophil proportions in horses eating haylage (3.0% ± 0.6%) were significantly lower compared with baseline (5.1 ± 0.7, P = .04) and horses eating hay (6.3% ± 0.8%, P < .01). Plasma eicosapentaenoic acid to arachidonic acid ratios were higher in horses eating haylage for 6 weeks (0.51 ± 0.07) when compared with baseline (0.34 ± 0.05, P < .01) and horses eating steamed (0.24 ± 0.02, P < .01) or dry hay (0.25 ± 0.03, P < .01). CONCLUSIONS AND CLINICAL IMPORTANCE: Steamed hay and haylage reduce dust exposure compared with dry hay, but only haylage increased the ratio of anti-inflammatory to pro-inflammatory lipids while reducing BAL neutrophil proportions within 6 weeks.

A randomised, controlled trial to determine the effect of levothyroxine on Standardbred racehorses.

BACKGROUND: The use of thyroid supplement is pervasive in athletic horses although its effects on measures of performance are not known. OBJECTIVES: One purpose of this study was to determine whether supra-physiologic doses of levothyroxine affect the velocities at which blood lactate was greater than 4 mmol/L (VLa4 ) and heart rate was over 150 (V150 ) and 200 (V200 ) beats per minute respectively. Additionally, a survey of post-race blood samples was also conducted to determine whether high thyroxine concentrations were common in racehorses. STUDY DESIGN: A randomised, crossover, trial was performed in six healthy Standardbred racehorses. METHODS: Study 1: T4 was determined in 50 post-race samples from a single Standardbred meet. Study 2: Research horses were trained to fitness and then randomised to one of three treatments: carrier, 0.1 mg/kg thyroxine or 0.25 mg/kg thyroxine for 2 weeks. Horses completed a standardised exercise treadmill test (SET) to fatigue on the last day of treatment. Serum free and total thyroxine and triiodothyronine were determined on the day of SET testing. Blood lactate and ECG data were collected during the SET at 6, 8, 10, 11, and 12 m/s and during recovery. The effect of treatment and SET on heart rate and blood lactate was examined using generalised linear mixed models. Post hoc analysis was adjusted for multiple comparisons using Tukey's Test. Data were expressed as mean ± standard deviation and P < .05 was considered significant. RESULTS: Study 1: The median T4 value in this population of horses was 2.00 µg/dL (laboratory's normal range 1.5-4.5 µg/dL) and 3 of 50 racehorses had values above the laboratory reference range. Study 2: Levothyroxine at 0.25 mg/kg resulted in higher heart rates during SET (199 ± 30, 223 ± 17 and 239 ± 9 bpm at 6, 8 and 10 m/s respectively) and recovery (144 ± 20 and 119 ± 15 at 5 and 15 min) as compared to placebo (176 ± 18, 203 ± 10 and 219 ± 6 bpm at 6, 8, and10 m/s and 126 ± 5, 102 ± 11 at 5-15 minutes respectively). Three of six horses developed cardiac arrhythmias including atrial fibrillation. MAIN LIMITATIONS: A relatively small number of animals were used and a SET is not identical to actual racing conditions. CONCLUSIONS: Supra-physiologic thyroxine supplementation caused a decreased V200 during a standard exercise test and may result in cardiac arrhythmias.

Effects of forages, dust exposure and proresolving lipids on airway inflammation in horses.

OBJECTIVE: To investigate the role of omega-3 polyunsaturated fatty acids (Ω-3)-derived proresolving lipid mediators (PRLM) in the resolution of mild airway inflammation in horses. ANIMALS: 20 horses with mild airway inflammation. PROCEDURES: Horses previously eating hay were fed hay pellets (low Ω-3 content; n = 10) or haylage (high Ω-3 content; 9) for 6 weeks. Dust exposure was measured in the breathing zone with a real-time particulate monitor. Bronchoalveolar lavage (BAL) was performed at baseline, week 3, and week 6. The effect of PRLM on neutrophil apoptosis and efferocytosis was examined in vitro. BAL fluid inflammatory cell proportions, apoptosis of circulating neutrophils, efferocytosis displayed by alveolar macrophages, and plasma lipid concentrations were compared between groups fed low and high amounts of Ω-3 by use of repeated measures of generalized linear models. RESULTS: Dust exposure was significantly higher with hay feeding, compared to haylage and pellets, and equivalent between haylage and pellets. BAL fluid neutrophil proportions decreased significantly in horses fed haylage (baseline, 11.8 ± 2.4%; week 6, 2.5 ± 1.1%) but not pellets (baseline, 12.1 ± 2.3%; week 6, 8.5% ± 1.7%). At week 6, horses eating haylage had significantly lower BAL neutrophil proportions than those eating pellets, and a significantly lower concentration of stearic acid than at baseline. PRLM treatments did not affect neutrophil apoptosis or efferocytosis. CLINICAL RELEVANCE: Despite similar reduction in dust exposure, horses fed haylage displayed greater resolution of airway inflammation than those fed pellets. This improvement was not associated with increased plasma Ω-3 concentrations. Feeding haylage improves airway inflammation beyond that due to reduced dust exposure, though the mechanism remains unclear.

The effect of lower airway inflammation on inflammatory cytokine gene expression in bronchoalveolar lavage fluid and whole blood in racing Thoroughbreds.

BACKGROUND: Immunological mechanisms involved in the pathogenesis of mild to moderate equine asthma (MEA) are not completely understood. There are limited data on bronchoalveolar lavage fluid (BALF) and blood inflammatory cytokine profiles in racehorses with MEA, and the effect of racing on inflammatory cytokines is unknown. HYPOTHESIS/OBJECTIVES: We hypothesized that inflammatory cytokine gene expression in BALF and resting blood would be higher in racehorses with lower airway inflammation compared to healthy controls, and that gene expression in blood collected immediately post-race would be increased compared to resting blood in racehorses with lower airway inflammation. ANIMALS: 38 racing Thoroughbreds (samples: 30 resting blood, 22 post-race BALF, 41 post-race blood). METHODS: Prospective observational study. Inflammatory cytokine gene expression was determined in resting blood, post-race BALF and post-race blood from racehorses with lower airway inflammation and controls. RESULTS: Lower airway inflammation was diagnosed in 79 % of racehorses (23 % neutrophilic, 67 % mastocytic, and 10 % mixed). There was no difference in gene expression in BALF or resting blood between racehorses with lower airway inflammation and controls. IL-8 gene expression was higher in post-race blood compared to resting peripheral blood, regardless of disease (p = 0052). BALF neutrophil proportions increased with increasing IL-1ß gene expression in all sample types (p = 0.0025). BALF mast cell proportions increased with increasing TNF-α gene expression in post-race blood (p = 0.015). CONCLUSIONS AND CLINICAL IMPORTANCE: Lower airway inflammation was common in a population of racehorses without respiratory signs or exercise intolerance. Exercise alone increased peripheral blood IL-8 gene expression. Inflammatory cytokine gene expression was not increased in BALF or resting blood in horses with subclinical lower airway inflammation, precluding its diagnostic utility in clinical practice.

Equine respiratory viruses, airway inflammation and performance in thoroughbred racehorses.

Equine asthma is a common cause of poor performance in racehorses but it is unclear if respiratory viruses contribute to its etiology. The objective of the study was to determine if respiratory viruses were associated with clinical signs and bronchoalveolar lavage fluid (BALF) cytology in Thoroughbred racehorses. Equine herpesviruses (EHV-1, 2, 4, 5) and equine rhinitis A and B viruses (ERBV, ERAV) genomes were quantified by qPCR in nasopharyngeal, tracheal, and BALF samples collected after racing. The relationships between virus detection and load and clinical signs, performance, BALF cytology, and environmental exposures were examined with generalized linear mixed models. Ninety-two samples were collected from 31 horses. EHV-1 and ERAV were not found; EHV-4 was detected in only one sample. EHV-2, EHV-5 and ERBV were more likely to be detected in upper airway samples than in BALF (P < 0.0001). Neither respiratory virus detection nor load was associated with clinical signs or performance. Nasopharyngeal detection and load of ERBV and tracheal detection and load of EHV-5 were associated with increased proportions of neutrophils in BALF (P < 0.003). However, nasopharyngeal detection and load of EHV-5 was not (P = 0.11). Nasopharyngeal detection and load of EHV-2 were associated with decreased BALF mast cell proportions. Respirable dust exposures were significantly higher in horses with detection of ERBV when compared to horses with no detectable ERBV (P < 0.001). Our results suggest that ERBV, EHV-2 and EHV-5 are commonly present in upper airways of healthy racehorses; however, the role they play in the etiology of equine asthma remains unclear.

Equine Asthma: Current Understanding and Future Directions.

The 2019 Havemeyer Workshop brought together researchers and clinicians to discuss the latest information on Equine Asthma and provide future research directions. Current clinical and molecular asthma phenotypes and endotypes in humans were discussed and compared to asthma phenotypes in horses. The role of infectious and non-infectious causes of equine asthma, genetic factors and proposed disease pathophysiology were reviewed. Diagnostic limitations were evident by the limited number of tests and biomarkers available to field practitioners. The participants emphasized the need for more accessible, standardized diagnostics that would help identify specific phenotypes and endotypes in order to create more targeted treatments or management strategies. One important outcome of the workshop was the creation of the Equine Asthma Group that will facilitate communication between veterinary practice and research communities through published and easily accessible guidelines and foster research collaboration.

An observational study of environmental exposures, airway cytology, and performance in racing thoroughbreds.

BACKGROUND: Mild equine asthma is presumed to arise in response to environmental exposures but the relative impact of differing inflammatory phenotypes upon performance are largely unexplored. HYPOTHESES: Airway inflammation negatively affects performance and cytological phenotype varies with environmental exposure. ANIMALS: Thoroughbred racehorses in active training and racing. METHODS: Thoroughbreds were recruited 24-48 hours before racing. Each horse was eligible for re-enrollment with each race entry. Within one hour of race completion, physical examination, respiratory endoscopy, and BAL were performed. Respirable and inhalable dust, respirable endotoxin, and respirable ß-glucan exposures were measured at the breathing zone within one week after racing. Controlling for age, trainer, and pulmonary hemorrhage, the relationship between performance, bronchoalveolar lavage fluid (BALF) cytology, and measures of exposure were modeled. RESULTS: Performance and BALF data were collected on 64 individual horses from 8 stables for a total of 98 race performances and 79 dust exposure assessments. Evidence of mild equine asthma was found in 80% (78/98) of BALF samples from 52/64 horses. For each percent increase in BALF mast cell and neutrophil proportions, speed figures were reduced by 2.9 (P = .012) and 1.4 (P = .046) points, respectively. Respirable dust concentration was associated with BALF neutrophil proportions (P = .015). Bronchoalveolar lavage fluid mast cell proportions were only associated with respirable ß-glucan exposures (P = .030). CONCLUSIONS AND CLINICAL IMPORTANCE: Mild equine asthma is common in racing horses and negatively impacts performance. The data support that respirable, rather than inhalable, dust exposure measures are pertinent to equine airway health.

Gentamicin concentrations in synovial fluid obtained from the tarsocrural joints of horses after implantation of gentamicin-impregnated collagen sponges.

OBJECTIVE: To determine synovial fluid gentamicin concentrations and evaluate adverse effects on the synovial membrane and articular cartilage of tarsocrural joints after implantation of a gentamicin-impregnated collagen sponge. ANIMALS: 6 healthy adult mares. PROCEDURES: A purified bovine type I collagen sponge impregnated with 130 mg of gentamicin was implanted in the plantarolateral pouch of 1 tarsocrural joint of each horse, with the contralateral joint used as a sham-operated control joint. Gentamicin concentrations in synovial fluid and serum were determined for 120 hours after implantation by use of a fluorescence polarization immunoassay. Synovial membrane and cartilage specimens were collected 120 hours after implantation and evaluated histologically. RESULTS: Median peak synovial fluid gentamicin concentration of 168.9 microg/mL (range, 115.6 to 332 microg/mL) was achieved 3 hours after implantation. Synovial fluid gentamicin concentrations were < 4 microg/mL by 48 hours. Major histologic differences were not observed in the synovial membrane between control joints and joints implanted with gentamicin-impregnated sponges. Safranin-O fast green stain was not reduced in cartilage specimens obtained from treated joints, compared with those from control joints. CONCLUSIONS AND CLINICAL RELEVANCE: Implantation of a gentamicin-impregnated collagen sponge in the tarsocrural joint of horses resulted in rapid release of gentamicin, with peak concentrations > 20 times the minimum inhibitory concentration reported for common pathogens that infect horses. A rapid decrease in synovial fluid gentamicin concentrations was detected. The purified bovine type I collagen sponges did not elicit substantial inflammation in the synovial membrane or cause mechanical trauma to the articular cartilage.