Immunohistochemical localization of alpha 2-beta 1-glycoprotein in horses

Alpha 2-beta 1-glycoprotein may be found free in horse serum or complexed with alpha-1-proteinase inhibitor to form pre-alpha 2-elastase inhibitor. There has been little information published concerning alpha 2-beta 1-glycoprotein and its possible tissue sources in horses. A peroxidase-antiperoxidase technique was used to identify alpha 2-beta 1-glycoprotein in buffy coat and bone marrow neutrophils of healthy horses. Macrophages and neutrophils in bronchoalveolar lavage samples from clinically normal horses and from horses with chronic pulmonary disease also were positive for alpha 2-beta 1-glycoprotein. Alpha 2-beta 1-glycoprotein was identified in some instances in normal equine hepatocytes of formalin-fixed liver sections. In formalin-fixed lung sections from horses with chronic, small-airway disease and chronic bronchointerstitial pneumonia, alpha 2-beta 1-glycoprotein was observed in some airway secretions and in macrophages.     

Immunohistochemical localisation of alpha-1-protease inhibitor in the horse

Using a peroxidase anti-peroxidase technique alpha-1-protease inhibitor (alpha-1-PI) was identified in normal equine hepatocytes in formalin-fixed liver sections, and in airway secretions and macrophages in formalin-fixed lung sections of horses with chronic small airway disease and chronic bronchointerstitial pneumonia. In addition, it was identified occasionally in macrophages in bronchoalveolar lavage samples from clinically healthy horses and from horses with chronic small airway disease. Equine peripheral blood leucocytes and formalin-fixed lung sections with normal histology were negative for alpha-1-PI.     

Evaluation of plasma alpha-2-macroglobulin and interactions with tumour necrosis factor-alpha in horses with endotoxemic signs.

The electrophoretic position and behavior of the native and activated forms of equine plasma alpha-2-macroglobulin (alpha 2M) were characterized and compared to human alpha 2M by nondenaturing polyacrylamide-gel electrophoresis (PAGE). Plasma alpha 2M was also compared between 6 normal horses and 6 horses with clinical signs of colic and endotoxemia due to volvulus or enteritis. Native and activated forms of alpha 2M were quantified by PAGE and densitometry. Binding of radio-labeled recombinant human tumour necrosis factor-alpha (125I-rhTNF-alpha) to native and activated forms of equine alpha 2M was also evaluated by autoradiography and densitometry of PAGE. Equine plasma alpha 2M migrated as a single band at a position equivalent to native human alpha 2M. Methylamine-reacted equine plasma samples resulted in faster migration of alpha 2M in a similar position to activated human alpha 2M. However, in methylamine-reacted equine plasma, an intermediate alpha 2M band was consistently present between the bands corresponding to native and activated alpha 2M. Amounts of plasma alpha 2M were similar in normal and endotoxemic horses, and remained in the electrophoretically slow or unreacted native form. The vast majority of 125I-rHuTNF-alpha did not bind to alpha 2M or other equine plasma proteins. 125I-rHuTNF-alpha bound weakly to both native and fast methylamine-reacted equine forms of alpha 2M, although binding was better to the activated form. This study indicates that: (1) equine plasma alpha 2M behaves similarly to human alpha 2M on PAGE, (2) plasma alpha 2M of horses can be activated to electrophoretically fast forms, but it is neither activated nor depleted during endotoxemia, and (3) the binding interactions between equine alpha 2M and TNF-alpha are too low to implicate equine alpha 2M as a regulator of TNF-alpha during endotoxemia in horses.     

The activity and expression of chitinase in the equine lung and its activity in normal horses and animals with recurrent airway obstruction

Recurrent airway obstruction (RAO) is a chronic inflammatory condition in equine lung, which may share a common immunological basis with human asthma, in which dysregulated Th2 responses occur. Mammals express chitinases and chitinase-like proteins, two of which are active enzymes, chitotriosidase and acidic mammalian chitinase (AMCase). Both enzymes are upregulated in a range of inflammatory conditions, including asthma. We investigated the activity of chitinase in bronchoalveolar lavage fluid from horses with and without RAO in response to organic dust challenges. No significant differences were found in activity, although in one study RAO animals had elevated chitinase activity that fell short of statistical significance. The pH optimum and pH lability of the activity was consistent with the presence of chitotriosidase. RT-PCR amplification of the mRNA encoding chitotriosidase and AMCase in normal equine lung showed that chitotriosidase, but not AMCase, is expressed in trachea, bronchi, and peripheral lung tissue. The gene for chitotriosidase was identified from the Equus caballus (horse) genome 1.1 database and its similarity to the same genes from other species was determined. The results of this study indicate that the involvement of chitotriosidase in RAO is uncertain.     

Pulmonary function,airway cytology and bronchoalveolar lavage fluid drug concentration after aerosol administration of cefquinome to horses

The administration of antibiotics by aerosol to horses suffering from respiratory infections may partially circumvent the limitations of antimicrobial therapy, e.g. large injection volumes, low bioavailability and risk of diarrhoea. Only injectable formulations are available currently and usually contain other substances that could irritate the mucosa and induce coughing and bronchospasm. In addition, the quality of the aerosol, particularly in terms of the delivery of antibiotics to the deep parts of the lung, is unknown. Although used under field conditions, cefquinome delivered by aerosol has never been studied in horses. This study examined the safety of cefquinome injectable solution, administered by aerosol at a dose of 225 mg/inhalation to 7 healthy horses, by assessing (1) pulmonary function before and 15 min after a single inhalation, at the first day (Day 1) and the fifth day (Day 5) of a 5 day period treatment; and (2) the inflammatory status of the lung, i.e. percentage neutrophils and myeloperoxidase concentration, based on bronchoalveolar lavage (BAL) at D1 and D5. In addition, cefquinome concentrations were measured in bronchoalveolar lavage fluid after aerosol, intravenous (i.v.) and intramuscular (i.m.) administrations. A single aerosol of cefquinome injectable solution did not induce any immediate nor delayed pulmonary side effects in healthy horses and produced cefquinome concentrations in bronchoalveolar lavage (BAL) within 30 min that were higher than the minimal inhibitory concentration of the main equine respiratory pathogens. These results should stimulate further studies, especially in horses suffering from bronchial hyper‐reactivity. Aerosol delivery of antibiotics may well have a role in equine therapeutics.     

Cloning and expression of the extra-cellular part of the alpha chain of the equine high-affinity IgE receptor and its use in the detection of IgE

The high-affinity receptor for IgE (FcepsilonRI) plays a central role in IgE-mediated allergic reactions. Cross-linking of FcepsilonRI by IgE-antigen complexes results in the activation of mast cells and basophils and is thought to contribute to the immunopathology of Heaves, a chronic obstructive pulmonary disease of horses. Recombinant protein corresponding to the extra-cellular portion of the FcepsilonRI alpha subunit, cloned and sequenced previously, was expressed using both mammalian cells and insect cells. The yield of expressed protein was considerably greater using insect cells and the baculovirus expression system. The recombinant proteins differed in size between the two systems, presumably due to differences in the extent of glycosylation. However, recombinant protein from both cell systems bound equine IgE present in bronchoalveolar lavage fluid from horses with Heaves. These results suggest that the recombinant extra-cellular part of FcepsilonRI should be a useful tool with which to study equine IgE responses.     

Arachidonate metabolites in bronchoalveolar lavage fluid from horses with and without COPD.

Arachidonate metabolites were measured in bronchoalveolar lavage fluid (BALF) from horses with (N = 4) and without (N = 7) chronic obstructive pulmonary disease (COPD). Prostaglandin (PG) D2, leukotriene (LT) B4 and LTC4 were present in highest concentrations in BALF from clinically normal horses. Concentrations of PGE2 and PGF were significantly higher in BALF from horses with COPD than in BALF from normal horses, but no differences were detected in thromboxane B2, 6-keto-PGF1 alpha, PGD2, LTB4 or LTC4.     

Interaction of transforming growth factor-beta-1 with alpha-2-macroglobulin from normal and inflamed equine joints.

Binding between equine plasma alpha-2-macroglobulin (alpha 2M) and several cytokines known to participate in inflammatory reactions in other species was initially examined. Plasma was obtained from 5 horses with various abnormalities. Samples, both untreated and after reaction with methylamine, were incubated with exogenous, radiolabeled, porcine-derived transforming growth factor-beta-1 (125I-TGF-beta 1), recombinant human interleukin-1-beta (125I-IL-1 beta), and recombinant human tumor necrosis factor-alpha (125I-rhTNF-alpha). They were then subjected to nondenaturing polyacrylamide gel electrophoresis (PAGE). Binding of the native (slow) and activated (fast) forms of alpha 2M to each cytokine was subjectively evaluated with autoradiography. Equine alpha 2M bound 125I-TGF-beta 1. However, poor or no binding was observed between alpha 2M and either of 125I-rhTNF-alpha or 125I-IL-1 beta. Synovial fluid was then obtained from 6 normal horses, 6 horses with septic arthritis, and 6 horses with degenerative joint disease. Untreated and methylamine-reacted samples were quantitatively examined for binding with 125I-TGF-beta 1, using the autoradiographic techniques described above and densitometry. Native and activated alpha 2M were also quantified by densitometry of PAGE gels. Native alpha 2M was significantly elevated in septic arthritis (6.4% to 29.5% of total protein detected) and degenerative joint disease (2.8% to 12.3%), compared to normal joints (0.9% to 4.2%). Activated alpha 2M, however, was not detected in untreated synovial fluid samples. In all plasma and joint fluid samples, whether untreated or reacted with methylamine, 125I-TGF-beta 1 bound predominantly to alpha 2M, and preferentially to the activated form of alpha 2M. In synovial fluid, the amount of 125I-TGF-beta 1 binding was proportional to the quantity of alpha 2M present. These results indicate that: 1) equine alpha 2M binds TGF-beta 1; 2) the native form of alpha 2M is present in both equine plasma and synovial fluid, and 3) alpha 2M is a major binding protein for TGF-beta 1 in equine synovial fluid. Therefore, alpha 2M may play a role in regulating this mediator of inflammation in equine joints.     

A novel model for equine recurrent airway obstruction

Equine recurrent airway obstruction (RAO; a term combining both chronic obstructive pulmonary disease (COPD) and summer pasture associated obstructive pulmonary disease (SPAOPD)) is one of the most common equine respiratory diseases with up to 50% of horses affected worldwide. The etiopathogenesis of RAO is unknown although pulmonary hypersensitivity to inhaled mold antigens may be involved. Recent work in our laboratory demonstrating elevated levels of IL-4 and IL-13 mRNA in the airways and peripheral blood of horses with RAO is consistent with an atopic component to RAO. Little is known regarding the earliest phases of RAO in horses. Here we describe the development of a novel airway model for equine RAO that utilizes ovalbumin-coated polystyrene beads for airway sensitization and challenge. Aerosol challenge of sensitized ponies with OVA-coated microbeads resulted in decreased airway compliance, increased percentage of lymphocytes and neutrophils in the bronchoalveolar lavage fluid, and evidence of a Th2 cytokine response in the bronchoalveolar cells. These results suggest that this approach may be useful in describing the initial stages of RAO development in the horse.     

Neutrophil migration induced by equine respiratory secretions, bronchoalveolar lavage fluids and culture supernatants of pulmonary lavage cells

Supernatants of equine respiratory secretions enhanced the migration of equine neutrophils into the lower compartments of Boyden chambers. Checkerboard analysis revealed that the neutrophil migration promoting activity (NMPA) of secretion specimens was in great part caused by chemokinesis, irrespective of the neutrophil score of the specimen. The NMPA of respiratory secretions was correlated neither with the neutrophil score of the secretion specimen nor with the severity of the chronic pulmonary disease. Respiratory secretions collected while horses were kept under low dust or under dusty housing conditions induced migration of neutrophils in the same order of magnitude. The number of migrated neutrophils and the procoagulant activity (PCA) within respiratory secretion specimens was positively correlated; however, the meaning of this finding is not yet clear. None of the nine cell-free supernatants of bronchoalveolar lavage fluid, which were assayed undiluted, induced significant neutrophil migration, although some samples contained up to 4.0 x 10(5) neutrophils/ml. In vitro culture of lung lavage cells, which mainly comprised macrophages and lymphocytes, without stimulation or with the addition of low doses of phytohemagglutinin (PHA) resulted in the secretion of NMPA which was in great part chemotactic. However, culture supernatants of lung cell preparations which were stimulated by lipopolysaccharide (LPS) or by PHA-prestimulated lymphocytes reduced the migration of neutrophils compared with the supernatants of control cells. NMPA within culture supernatants had a highly significant negative correlation with the PCA of macrophages within the lung cell preparations. Our results imply that a complicated and sophisticated regulation underlies neutrophil accumulation within the airways of horses affected with chronic pulmonary disease. Future experiments are required to assess the biological significance of the factors modulating neutrophil migration which are present in the respiratory secretions and in the culture supernatants of equine lung lavage cells.     

Comparison of transtracheal aspirate and bronchoalveolar lavage cytology in 50 horses with chronic lung disease

Comparisons were made between transtracheal aspirate (TTA) and bronchoalveolar lavage (BAL) cytology obtained from 50 horses with chronic lung disease and from 10 control horses. There was no significant correlation between the TTA cytology and the BAL cytology, suggesting that the cell population in the trachea is not representative of the cell population in the lower airways. In control horses the range of differential cell counts obtained from TTA fluid was remarkably large, whereas the variability in cell populations observed in BAL fluid was smaller. In the principal horses the total and differential cell counts of the TTA and BAL fluids were within the 95 per cent confidence interval in 38 and 24 per cent of cases, respectively; and an increase in percentage neutrophils was most common. It was concluded that BAL may be a useful diagnostic aid when evaluating horses with chronic lung disease, but that the clinical usefulness of cytological evaluations of TTA fluid may be limited in these cases.     

Comparison of bronchoalveolar lavage and respiratory secretion cytology in horses with histologically diagnosed pulmonary disease

Equine bronchoalveolar lavage (BAL) fluid collected from 70 horses and respiratory secretions (RS) obtained from 61 of these horses were evaluated cytologically and grouped according to the histological diagnosis of the lungs from which they were obtained. The histological categories included: normal lung (8 horses); pulmonary eosinophilic infiltration (9 horses); interstitial pneumonia (5 horses); pulmonary hemorrhage (5 horses); and mild (12 horses), moderate (7 horses) and severe (24 horses) chronic small airway disease. In horses with pulmonary disease, all BAL samples and all but one RS sample differed cytologically to those obtained from normal horses; however, the type and severity of the pulmonary disease could not always be determined using either BAL or RS cytology. There was a positive association between the percentage of neutrophils in BAL and the neutrophil scores in RS specimens; there was no positive association between other cell types.     

Multinodular pulmonary fibrosis in five horses

CASE DESCRIPTION: 5 horses were evaluated because of decreased appetite, weight loss, fever, cough, tachypnea, and respiratory distress. CLINICAL FINDINGS: Tachycardia, tachypnea, increased respiratory effort, lethargy, fever, poor body condition, and nasal discharge were detected in various combinations on initial physical examination. Evaluation of the lower portion of the respiratory tract via radiography and ultrasonography revealed a severe nodular interstitial pattern. Histologic examination of lung tissue revealed interstitial expansion of alveolar parenchyma with collagen, intraluminal accumulation of neutrophils and macrophages within the alveoli, and occasional intranuclear inclusion bodies within alveolar macrophages. Equine herpesvirus type 5 was detected in samples of lung tissue, bronchoalveolar lavage fluid, or both via polymerase chain reaction assay in all cases. A diagnosis of equine multinodular pulmonary fibrosis (EMPF) was established. TREATMENT AND OUTCOME: Horses were provided supportive treatment and were administered a variety of medications including corticosteroids and acyclovir. Two horses survived and returned to their previous level of activity. Three horses were euthanized because of either deterioration of clinical condition (n=2) or failure to improve within 4 weeks of initiation of treatment (1). CLINICAL RELEVANCE: EMPF should be considered as a differential diagnosis for adult horses with interstitial pneumonia and should be suspected on the basis of characteristic radiographic, ultrasonographic, and histopathologic findings. Equine herpesvirus type 5 is found in association with EMPF; although the exact pathogenic role this virus plays in EMPF is unknown, equine herpesvirus type 5 may be an etiologic agent or cofactor in the development of EMPF.     

The role of alveolar macrophages in the pathogenesis of recurrent airway obstruction in horses

When challenged with allergens and pro-inflammatory agents, such as Aspergillus fumigatus (AF), hay dust solution (HDS) and lipopolysaccharide (LPS), the innate immune response will not only activate the immune system but also increase the amount of pro-inflammatory cytokines in the bronchoalveolar space. The aim of this study was to assess the response of equine alveolar macrophages to different aerosolized challenges and to investigate the differences in this response between horses susceptible or nonsusceptible to recurrent airway obstruction (RAO). Seven susceptible and 5 nonsusceptible horses were challenged with saline, LPS, HDS, or AF, and bronchoalveolar lavage (BAL) cytology, total cell counts, and lung function were assessed. In addition, alveolar macrophages were isolated 6 and 24 hours after challenge, and macrophage mRNA expression of tumor necrosis factor (TNF)-alpha and interleukins (IL) IL-1beta, IL-6, IL-8, and IL-10 were measured by means of real-time (RT) polymerase chain reaction (PCR). There was a significant difference in lung function, neutrophil ratios, and total cell counts in the bronchoalveolar lavage fluid between RAO-susceptible and nonsusceptible horses. In addition, the expression of TNF-alpha, IL-1beta, and IL-8 by alveolar macrophages after challenges were higher in susceptible horses, than in nonsusceptible horses. In contrast, I1-6, considered an anti-inflammatory cytokine, showed a higher expression in nonsusceptible horses 6 hours after inhalation challenge with allergens and pro-inflammatory antigens. These data suggest that the differences between susceptible and nonsusceptible horses to RAO are not only dependent on adaptive immunity but also start with an innate immune response.     

Antigen challenge increases adherence of circulating neutrophils in horses with chronic obstructive pulmonary disease

Activation of circulating neutrophils has been observed following challenge of horses with chronic obstructive pulmonary disease (COPD) and may facilitate the accumulation of these cells in the airways. In this study, no significant difference was observed between adherence to protein coated plastic of blood neutrophils from asymptomatic COPD-susceptible and normal horses stimulated by the mediators PAF, human recombinant (hr)IL-8 and hrC5a. Twenty-four hours after the start of a 7 h antigen challenge, adherence of unstimulated neutrophils from COPD-susceptible horses increased from 2.5 (0.5-4.1)% and 3.4 (0.6-6.6)% to 19.6 (16.9-20.3)% and 21.8 (10.6-23.1)% adherence for cells in medium containing 0.1% or 0.2% BSA, respectively; (median [range]; n = 4). Adherence of cells from normal horses remained unchanged. Addition of an anti-CD18 monoclonal antibody, H20A, inhibited the increase in adherence at 24 h by 96 (45-100)%, n = 3. The percentage of neutrophils in bronchoalveolar lavage fluid at 24 h increased from 1 (0-2) to 80 (65-94), (median (range), n = 4). These results suggest that antigen challenge results in exposure of circulating equine neutrophils to one or more factors that prime, or activate, these cells, which may enhance their recruitment to the lungs. Inhibition of circulating neutrophil activation may therefore represent a therapeutic target.     

Effects of aerosolized albuterol on physiologic responses to exercise in standardbreds.

OBJECTIVE: To examine the effects of an aerosolized beta2-adrenoreceptor agonist, albuterol, on performance during a standardized incremental exercise test in clinically normal horses. ANIMALS: 8 Standardbred pacing mares. PROCEDURE: Clinically normal horses, as judged by use of physical examination, hematologic findings, serum biochemical analysis, and airway endoscopy, were randomly assigned to 2 groups and were given 900 microg of albuterol via a metered-dose inhaler 30 minutes before beginning a standardized incremental exercise test in a crossover design with a 7-day minimum washout. Further examination included measurement of baseline lung mechanics, response to histamine bronchoprovocation, and bronchoalveolar lavage. RESULTS: No significant differences (albuterol vs placebo) were seen for any incremental exercise test variables (ie, maximum oxygen consumption, maximum carbon dioxide consumption, respiratory quotient, treadmill speed at heart rate of 200 beats/min, or number of steps completed during an incremental exercise protocol). Mast cell percentage was significantly (r = -0.84) associated with the concentration of aerosolized histamine that evoked a 100% increase in total respiratory system resistance. No other direct correlations between bronchoalveolar lavage fluid cell types and any indices of exercise capacity or airway reactivity were found. CONCLUSIONS AND CLINICAL RELEVANCE: Although no horse had exercise intolerance, 4 horses had airway hyperreactivity with bronchoalveolar lavage fluid mastocytosis; these horses may have been subclinically affected with inflammatory airway disease. In our study, albuterol did not enhance performance in 8 clinically normal racing-fit Standardbreds.     

Comparison of TGF-beta 1 concentrations in bronchoalveolar fluid of horses affected with heaves and of normal controls

Airway remodeling may play an important role in heaves pathophysiology. Transforming growth factor-beta 1 (TGF-beta1) is a potent profibrotic cytokine, which might contribute to airway wall thickening and fibrosis of bronchiolar and alveolar submucosa. An ELISA designed for the measurement of human TGF-beta1 was used to measured total TGF-beta1 released in bronchoalveolar lavage fluid (BALF) of normal horses and of those affected with heaves in remission. The specificity of the assay for TGF-beta1 of the horse was confirmed using recombinant equine TGF-beta1. The influence of hay exposure on TGF-beta1 release in the airways was also examined by stabling horses in a dusty environment. TGF-beta1 was found in the BALF of all horses. However, no significant difference between basal concentration of TGF-beta1 in BALF of control horses versus that of horses affected with heaves was found. Furthermore, no differences were identified in these populations 1 and 9 days after allergen challenge. In conclusion, these data indicate that TGF-beta1 is released in BALF fluid of horses in biologically active concentrations. Other studies are necessary for a better definition of the role of this cytokine within the lung, as our study does not establish a causal relationship between TGF-beta1 and the pathophysiology of heaves in the horse.     

Myeloperoxidase concentration in bronchoalveolar lavage fluid from healthy horses and those with recurrent airway obstruction

The aim of this work was to measure the myeloperoxidase (MPO) concentration in bronchoalveolar lavage (BAL) fluid collected from horses with recurrent airway obstruction (RAO), both in crisis and in remission, as well as from healthy horses. Seven horses with RAO were exposed to moldy hay until the maximum change in pleural pressure was greater than 1.5 kPa. At that point, BAL was performed, and the total cell counts and percentages in the fluid were immediately determined. To measure the MPO concentration in BAL-fluid supernatant, we used a specific enzyme-linked immunosorbent assay with polyclonal antibodies against equine MPO. The tests were repeated on the horses with RAO after they had spent 2 mo on pasture. Six healthy horses serving as controls underwent the same tests. The absolute and relative neutrophil counts and the MPO concentration in the BAL fluid were significantly greater in the horses with an RAO crisis than in the control horses. After 2 mo on pasture, the horses that had been in RAO crisis were clinically normal, and their neutrophil counts and MPO levels in BAL fluid had significantly decreased; during remission their neutrophil counts were not significantly different from those in the healthy horses, but their MPO concentration remained significantly higher. This study showed that determining the MPO concentration in a horse's BAL fluid is technically possible and that during remission from RAO the concentration remains higher than normal. Thus, MPO may be a marker of neutrophil presence and activation in the lower airways.     

Temporal regulation of cytokine mRNA expression in equine recurrent airway obstruction

Acute and chronic inflammation of the airway remains an important health problem for equids. "Heaves" or recurrent airway obstruction (RAO) remains one of the most commonly diagnosed conditions affecting the lung of older horses in Europe and the United States. The typical clinical signs of RAO include non-productive coughing, serous nasal discharge, labored expiratory effort, and flaring of the nostrils. Auscultation of the lungs of the affected horse often reveals abnormal respiratory sounds, described as crackles and wheezes, throughout the area of the lung field. These clinical signs occur secondary to an inflammatory response that results in bronchospasm, excessive mucus production and airway obstruction. This inflammatory response is characterized by the presence of excessive mucus and inflammatory cells, primarily neutrophils, in the small airways. Most evidence suggests that RAO is the result of a pulmonary hypersensitivity to inhaled antigens. Exposure of affected horses to hay dust, pollens, and mold spores leads to neutrophil accumulation in the lung and bronchospasm. The identification of allergen-specific IgE in bronchoalveolar lavage (BAL) fluid and sera of affected horses supports the involvement of a late phase, IgE-mediated, hypersensitivity reaction in the pathogenesis of equine RAO. The production of IgE antibodies is regulated by the cytokines IL-4 and IL-13. Using a quantitative PCR method we have reported that horses with RAO exhibit a modified Type 2 cytokine response characterized by the production of IL-4 and IL-13 mRNA, but not IL-5 mRNA in BAL cells. Interferon-gamma mRNA was also elevated, suggesting a mixed response. While these results are consistent with equine RAO being the result of an aberrant Type 2 cytokine response to inhaled allergens, others have failed to find any evidence of elevated Type 2 cytokine mRNA in BAL from horses with "heaves". It is likely that these disparate results could be the result of differences in the clinical stage of the affected animals or the timing of sample collection. Here, we report a diverse pattern of cytokine gene expression when sampling a group of affected horses over a period of time.     

Evaluation of clinical signs of disease, bronchoalveolar and tracheal wash analysis, and arterial blood gas tensions in 13 horses with chronic obstructive pulmonary disease treated with prednisone, methyl sulfonmethane, and clenbuterol hydrochloride.

We evaluated the efficacy of 3 treatments for chronic obstructive pulmonary disease in horses: prednisone (400 mg/horse, PO, daily; n = 7), methyl sulfonmethane (10 g/horse, PO, q 12 h; n = 6), and clenbuterol hydrochloride (0.4 mg/horse, PO, q 12 h; n = 7). A fourth group acted as controls (n = 6) and was not treated. The treatment period lasted 10 days. Each horse was a member of 2 different groups for 10 days, separated by an 18-day interval of no treatment. All horses were housed together in an outdoor pen without bedding. Horses were fed alfalfa/grass hay mix ad libitum from a large feeder. The same batch of hay was fed throughout the study. Multiple physical and laboratory variables were monitored prior to, during, and at the end of each 10-day trial period. Changes in lung sounds, respiratory effort, degree of anal movement, nasal discharge, temperature, respiratory rate, or heart rate were not significant. Changes in arterial blood gas tensions, tracheal wash or bronchoalveolar lavage cytologic findings, or phagocyte function were not significant. All horses were tachypneic and most were tachycardic. The median value for PaO2 was below normal for all horses. All tracheal wash and most bronchoalveolar lavage cytologic findings represented a suppurative response. Negative linear correlation was observed between PaO2 and degree of respiratory effort in these horses (eg, as PaO2 decreased, the degree of respiratory effort increased).