Bilateral bronchoalveolar lavage cytology profiles in a warmblood horse population during a 1-year period.

BACKGROUND: Bronchoalveolar lavage (BAL) cytology results from 1 lung might not be representative of both lungs. OBJECTIVES: To determine whether the lung site sampled would influence the horse's BAL cytology profile, and if a pooled BAL sample would be superior with regard to BAL cytology diagnosis in a cohort of healthy and subclinical asthmatic warmblood horses. ANIMALS: Fifty-nine horses in 2021 and 70 horses in 2022, the follow-up included 53 of the same in each year. METHODS: A cross-sectional study with follow-up included BAL cytology samples from individual lungs and from pooled BAL samples. The BAL samples were enumerated and differential cell count were applied to categorize the horses as control or with airway inflammation (AI). RESULTS: Bronchoalveolar lavage mast cell count was higher in left lung compared to right lung (2021; median 1.6 [range, 0.6-3.3] vs 1.2 [0.7-1.5] P = .009, 2022; median 3.1 [2.1-4.2] vs 2.4 [1.7-3.4], P < .001) and compared to pooled samples (2022; median 2.6 [1.7-3.7], P < .001). Between year 2021 and 2022, 17 of the horses had changes in BAL cytology from control to AI or vice versa. CONCLUSIONS AND CLINICAL IMPORTANCE: Pooled BAL sample was the least reliable for detecting AI, and was not representative of the overall lung condition.

Atrial fibrillation as a risk factor for exercise-induced pulmonary haemorrhage following a standardised exercise test.

BACKGROUND: Atrial fibrillation (AF) has been proposed as a risk factor for exercise-induced pulmonary haemorrhage (EIPH) due to increased pressure in the left atrium. OBJECTIVE: To evaluate if AF was associated with EIPH following a standardised exercise test (SET) to fatigue. STUDY DESIGN: Two-arm controlled experiment. METHODS: Ten untrained Standardbred mares mean (standard deviation [SD]) age 6 (2) years performed a SET on the treadmill in sinus rhythm (SR) (SET1) and 25-44 days after induction of self-sustained AF (SET2). AF was induced by tachypacing using a pacing device. Endoscopy, including tracheal wash and bronchoalveolar lavage (BAL), was performed 48-72 h before and 24 h after the two SETs. In addition, endoscopic grading of tracheal blood was performed 2 h after each SET. RESULTS: After SET1, none of the horses showed blood in the trachea, and two horses showed erythrophagocytosis. Following SET2, two horses had grade 1 blood in the trachea and free erythrocytes and erythrophagocytosis in the BAL, while another two horses had erythrophagocytosis in the BAL. In SET2, the overall performance on the treadmill was decreased with a lower maximum velocity (SET1 10.3 ± 0.8 m/s vs. SET2 8.9 ± 0.9 m/s, p = 0.004), a higher heart rate (284 ± 21 vs. 221 ± 18 bpm, p = 0.003) and more abnormal QRS complexes (p < 0.001) compared with SET1. CONCLUSIONS: Two horses showed signs of EIPH, resulting in visible blood in the trachea, when exercising in AF compared with SR. However, a possible link between EIPH, pulmonary pressure and AF needs to be further elucidated.

Evaluation of Treatment With Respiratory Gene Technology and Serum in a Group of Standard Bred Racehorses With Cytological Evidence of Mild Equine Asthma.

Alternative treatment options to glucocorticoids for equine asthma is desirable due to withdrawal time. The objective was to evaluate if serum and Respiratory Gene Technology (RGT), a commercial kit to produce autologous conditioned serum, was effective in reducing bronchoalveolar lavage (BAL) neutrophils and mast cells in racehorses with cytological evidence of mild equine asthma . Thirty-six Standardbred trotters in active training were enrolled in this randomized clinical trial; a healthy control group (n=11), a RGT group (n=12) and a serum group (n=13). Endoscopy including tracheal wash (TW) and BAL was performed before (T0), after a 6-week treatment period including 12 intramuscular injections of RGT or serum (T6) and as a follow-up 10 weeks after treatment (T16). A significant decrease in BAL neutrophils for the RGT group was found between T0 and T6 (P = .002, d=-1.51, CI: -2.43;-0.59) and for the serum group between T0-T6 (P = .002, d=-1.36, CI: -2.26;-0.46). Further, a significant decrease in BAL mast cells between T0-T6 for the both the RGT group (P = .019, d=-1.23, CI: -1.22;-0.34) and the serum group (P= .004, d=-0.81, CI: -1.65;0.04), and further between T0-T16 (RGT P= .011, d=-1.55, CI: -2.62;-0.48; serum P= .044, d=-0.65, CI: -1.68;-0.37). No significant difference in TW cytology was found for any of the time-points. Pro- and anti-inflammatory cytokines were regulated according to treatment. The control group showed no cytological differences between any time-point. Study results showed that intramuscular treatment with both RGT and serum was effective associated with reduction of BAL neutrophils and mast cells in horses with cytological evidence of mild equine asthma. Further large-scale studies are necessary.

Evaluation of Dynamic Structural Disorders in the Upper Airways and Applied Rein Tension in Healthy Dressage Horses During Riding in Different Gaits and Head-Neck Positions.

Flexion of the horse's head and neck during dressage riding reduces the pharyngeal lumen with the risk of increased upper airway resistance and upper airway obstructions. According to the Fédération Equestre Internationale, hyperflexion is achieved through force, whereas the position low-deep-round is nonforced. The objectives of this study were to evaluate (1) applied rein tension and (2) dynamic structural disorders in the upper airways in dressage horses in different gaits and different head-neck positions (HNPs). Overground endoscopy (OGE) and rein tension were evaluated in 13 clinically healthy and high-performance Warmblood dressage horses while being ridden in a standardized program comprised of four different gaits (halt, walk, trot, and canter) and in four HNPs (unrestrained, competition frame, hyperflexion, and low-deep-round). All included horses were able to achieve the desired HNPs. The HNP low-deep-round showed significantly lower rein tension than competition frame (P < .001) and hyperflexion (P < .001). An association was found between dynamic structural disorders in the upper airway tract evaluated by OGE and head-neck flexion, but this association was not linked to the degree of flexion. The HNP hyperflexion was neither associated with greater rein tension nor severe dynamic structural disorders than the HNP competition frame. This study confirms that low-deep-round is a nonforced position, in contrast to hyperflexion. Further studies are needed to evaluate whether dynamic structural disorders are a result of flexion or if the degree of flexion has an impact.

Bronchoalveolar lavage fluid cytokine, cytology and IgE allergen in horses with equine asthma.

The pathophysiology of equine asthma (EA) is still not fully described, but the involvement of an allergic reaction is strongly suspected. This theory has led to the use of allergen-specific immunoglobulin (Ig)E tests to support a diagnosis of asthma. The objective of this descriptive study was to evaluate the correlation between four subgroups of EA (mastocytic mild equine asthma [MEA], neutrophilic MEA, mixed MEA, and severe equine asthma [SEA]), allergen specific IgE (measured in both serum and BALF) and mRNA expression of selected genes in bronchoalveolar lavage fluid (BALF). Serum and BALF were collected from 64 horses with a history of lower airway problems with or without poor performance. Differential cell counts from BALF were used to assign horses to one of four groups (mastocytic MEA; neutrophilic MEA, mixed MEA, and SEA). The expression of messenger RNA (mRNA) coding for IL4, IL5, IL8, IL10, TGFB, TNFA, toll-like receptor (TLR)4, IL1RA, IL1B, matrix metalloproteinase 8 (MMP8), TLR9, chemokine ligand 5 (CCL5) and cluster of differentiation (CD)14 in BALF were measured using reverse transcriptase (RT) quantitative PCR (qPCR). Allergen-specific IgE was measured in serum and BALF using an allergen-specific IgE ELISA test with the screening panel: house mites, storage mites, mould and pollen. As expected, the BALF neutrophil differential count correlated with mRNA expression of MMP-8 (r = 0.611, p < 0.001), TLR-4 (r = 0.540, p < 0.001), IL-1RA (r = 0.490, p < 0.001), IL-1ß (r = 0.463, p < 0.001) and IL-8 (r = 0.302, p = 0.015). Cytokine expression of IL-1ß (p = 0.014), MMP8 (p = 0.028) and IL-1RA (p = 0.037) was significantly higher in the SEA group compared to the MEA subgroups. The BALF mast cell count was correlated with allergen-specific IgE for insects (r = 0.370, p = 0.002) and pollen (r = 0.313, p = 0.011). Eosinophils in BALF were correlated with BALF mRNA expression of IL-4 (r = 0.340, p = 0.006) together with a significant correlation between BALF eosinophils and allergen-specific IgE for mites (r = 0.930, p < 0.001) and pollen in BALF (r = 0.837, p < 0.001). No correlation was found between allergen-specific IgE in serum and BALF for any of the allergen in the screening panel. Based on these results from allergen-specific IgE in horses with EA is not found in systemic circulation, and only the mastocytic and mixed subgroups of horses with EA had allergen-specific IgE in BALF. Further studies are needed to clarify the relationships identified here.

Effect of standard clinical procedures on blood glucose concentration in clinically healthy horses.

BACKGROUND: Blood glucose concentrations fluctuate with stress, but little is known on how it is influenced by clinical procedures. The objective was to investigate the effect of clinical procedures on blood glucose concentration in healthy horses. MATERIALS AND METHODS: Prospective, experimental study. Seven hospital-owned research horses were included in the study. A total of 4 horses were sampled either during a control sedation trial or during 3 different student workshops (prepurchase, oral, and rectal examination-the latter 2 with sedation). Blood samples were taken every 15 minutes and glucose concentration in whole blood was measured immediately with a previously validated handheld glucometer until normalization after the end of the workshops. No food was provided during sampling periods. KEY FINDINGS: All measured blood glucose concentrations remained within reference interval. A significant increase in blood glucose concentration between baseline and peak was found during sedation (P = 0.005) and the oral workshop (P = 0.031). A decrease was found during prepurchase examination (P = 0.006; before exercising). Peak glucose concentration values between the sedation trial and both the oral (P = 0.065) and rectal workshop (P = 0.709) were not statistically different. Glucose measurements returned to baseline 1 hour after completion of the workshops. SIGNIFICANCE: No impact of different clinical procedures on the blood glucose concentration over the effects of sedation was found. It is advisable to wait 1 hour after a procedure to measure blood for glucose concentration in horses.

Age-related dynamics of pro-inflammatory cytokines in equine bronchoalveolar lavage (BAL) fluid and peripheral blood from horses managed on pasture.

The objectives of this study were to evaluate the natural age-related variation and compare the level of pro-inflammatory cytokines in the peripheral blood and lower airways of horses. The mRNA expression of IL-1ß, IL-6, IL-8 TNF-α TLR-4 in bronchoalveolar lavage (BAL) fluid and peripheral blood mononuclear cells (PBMC) were studied by quantitative real time polymerase chain reaction (PCR) and differential cell count cytology from 44 horses of different ages. A significant age-related increase was found for the mRNA expression of IL-6, IL-8, TLR-4 and TNF-α in stimulated BAL cells and for TNF-α in stimulated PBMC. Furthermore, a significant decrease was found in the mRNA expression of IL-1ß and TNF-α in stimulated BAL cells compared to stimulated PBMC. In conclusion, continued low antigen exposure of horses in the pasture environment could lead to a tight regulation of mRNA gene expression in the airway space compared to the peripheral blood and favour an age-related accumulation of mRNA genes.

Equine airway inflammation in loose-housing management compared with pasture and conventional stabling.

Icelandic horses are often stabled in loose-housing systems, and to date this type of stabling has not been evaluated with regard to its potential impact on respiratory health. The objective was to assess if differences in management systems (eg, conventional stable, loose housing and pasture only) affect the degree of airway inflammation, evaluated by cytology of tracheal aspirate (TA) and bronchoalveolar lavage (BAL) fluid. In total, 84 Icelandic horses (aged 8.1±4.6 years) housed under three different management systems (conventional stables [n=29], loose-house systems [n=29] and pasture [n=26]) were included. Endoscopy including mucus scoring, TA and BAL was performed. TA and BAL cytologies were evaluated by performing both the total cell count (TCC) and the differential cell count (DCC). Significantly higher BAL neutrophil DCC (P=0.032, P=0.040) and TA TCC (P=0.007, P=0.028) were found for each of the two groups of horses with indoor access (conventional stable and loose housing) compared with the pasture group. Regardless of stabling environment, weak positive correlations were found between TA and BAL TCC (r=0.37, P<0.001), between TA TCC and TA neutrophil ratio (r=0.33, P=0.002), as well as between TA and BAL neutrophil ratio (r=0.39, P=<0.001). A larger proportion of horses with indoor access showed evidence of subclinical airway inflammation characterised by an increase in TA and BAL neutrophil ratios.

Serum insulin-like growth factor 1 in the aging horse.

BACKGROUND: Insulin-like growth factor 1 (IGF-1) has important roles in anabolic processes in the musculoskeletal system and has been reported to decrease with age in both people and horses. OBJECTIVES: The objective of this study was to determine serum IGF-1 levels in the aging horse from early to late adulthood (age range 5-27 years). METHODS: Healthy horses (n = 72) were used in a cross-sectional study, while 37 paired serum samples were available for a longitudinal study. Serum IGF-1 protein was determined using an ELISA kit validated for use in equine samples. RESULTS: No association was found between serum IGF-1 levels and age in the cross-sectional study. In the longitudinal study, a latent variable model fitted to the data revealed that horses in general experienced a 5.2% increase of serum IGF-1 levels over a 5-year period, but horses crossing a change point around 9 years of age between the 2 samples experienced an 11.0% decrease. CONCLUSIONS: In this study, there was no evidence for aging being a factor in changes of IGF-1 levels in an adult and old horse population.

Age-related changes in intracellular expression of IFN-γ and TNF-α in equine lymphocytes measured in bronchoalveolar lavage and peripheral blood.

Diseases of the lower airways represent some of the most common conditions affecting horses of all ages, but the type and severity tends to follow the horses' age. The age-related dysregulation of pro-inflammatory cytokines may, in part contribute to the development of the diseases. Therefore, we hypothesize that the expression of pro-inflammatory cytokines increases with age. Peripheral blood mononuclear cells (PBMC) and bronchoalveolar lavage (BAL) cells from clinically healthy horses of different ages were used for the investigation. The cells were stimulated and the production of IFN-γ and TNF-α measured using flow cytometry. The frequency of IFN-γ producing lymphocytes in both BAL and PBMCs from old horses was significantly increased compared to the young horses. The age-related increase of TNF-α production was also found in PBMCs but not in BAL cells. In conclusion, the productions of certain pro-inflammatory cytokine are age-associated. This age-associated increase of pro-inflammatory cytokines production may be a co-factor for the pathogenesis of equine airway diseases.

Acute phase response to surgery of varying intensity in horses: a preliminary study.

OBJECTIVE: To evaluate the postoperative inflammatory response of horses to elective surgery of varying intensity. STUDY DESIGN: Prospective longitudinal study. ANIMALS: Horses referred to 2 hospitals for either arthroscopic removal of a unilateral osteochondritic lesion in the tibiotarsal joint (minimal surgical trauma, n=11), correction of recurrent laryngeal neuropathy by laryngoplasty and ventriculectomy (intermediate surgical trauma, n=10) or removal of an ovarian tumor by laparotomy (major surgical trauma, n=5). METHODS: Horses had a thorough clinical examination every day. White blood cell (WBC) counts and concentrations of serum amyloid A (SAA), fibrinogen, and iron were assessed in blood samples obtained before, and 1-3, 5, 7, 9, and 11 days after surgery. Differences in levels of the inflammatory markers between the 3 surgical groups were analyzed using repeated measures ANOVA. RESULTS: Postoperative concentrations of SAA and fibrinogen were significantly higher in horses that had laparotomy and ovariectomy than in horses that had laryngoplasty and ventriculectomy, or arthroscopy. Iron concentrations decreased to lower levels after intermediate and major surgical trauma than after small surgical trauma. WBC count did not differ between the 3 groups. CONCLUSIONS: Levels of SAA, fibrinogen, and iron reflected the intensity of the surgical trauma, whereas WBC count did not. CLINICAL RELEVANCE: Postoperative measurements of SAA, fibrinogen, and iron may be useful for comparing surgical trauma associated with new and established surgical techniques. Moreover, knowledge of the normal postoperative acute phase response is essential, if acute phase reactants are to be used for monitoring occurrence of postoperative infections.

Evaluation of pain and inflammation associated with hot iron branding and microchip transponder injection in horses.

OBJECTIVE: To compare effects of hot iron branding and microchip transponder injection regarding aversive behavioral reactions indicative of pain and inflammation in horses. ANIMALS: 7 adult horses. PROCEDURES: In a randomized controlled clinical crossover study, behavioral reactions to hot iron branding and microchip transponder injection were scored by 4 observers. Local and systemic inflammation including allodynia were assessed and compared by use of physiologic and biochemical responses obtained repeatedly for the 168-hour study period. Serum cortisol concentration was measured repeatedly throughout the first 24 hours of the study. Sham treatments were performed 1 day before and 7 days after treatments. RESULTS: Hot iron branding elicited a significantly stronger aversive reaction indicative of pain than did microchip transponder injection (odds ratio [OR], 12.83). Allodynia quantified by means of skin sensitivity to von Frey monofilaments was significantly greater after hot iron branding than after microchip transponder injection (OR, 2.59). Neither treatment induced signs of spontaneously occurring pain that were observed during the remaining study period, and neither treatment induced increased serum cortisol concentrations. Comparison with sham treatments indicated no memory of an unpleasant event. The hot iron branding areas had significantly increased skin temperature and swelling (OR, 14.6). Systemic inflammation as measured via serum amyloid A concentration was not detected after any of the treatments. CONCLUSIONS AND CLINICAL RELEVANCE: Microchip transponder injection induced less signs of pain and inflammation and did not seem to pose a higher long-term risk than hot iron branding. Consequently, results indicated that hot iron branding does inflict more pain and should be abandoned where possible.