Tumor necrosis factor activity in the circulation of horses given endotoxin

Serum and plasma from horses injected with endotoxin was examined for cytotoxic activity. Each of the cell lines, L929 and WEHI 164 clone 13, was sensitive to the cytotoxic effects of equine serum; however, a precipitation artifact caused by the use of isopropanol in the WEHI assay limited the use of this assay to samples containing less than 2 mg of protein/ml. In foals treated with a sublethal IV bolus of 5 micrograms of lipopolysaccharide (LPS)/kg and in adult horses given a low-dose continuous infusion of LPS (30 ng/kg/h for 4 hours), cytotoxic activity was detected in all serum or plasma samples taken between 30 minutes and 4 hours after LPS infusion began. In horses given either continuous or bolus LPS infusions, circulating cytotoxic activity peaked at 1 to 2 hours before decreasing sharply. The onset of pyrexia after LPS infusion coincided with the appearance of circulating cytotoxic activity, but the temperature remained high, even after cytotoxic activity disappeared. Treatment of horses with flunixin meglumine (1 mg/kg) appeared to blunt the pyrexic effect of low-dose continuous LPS infusion, but had no significant effect on circulating cytotoxic activity. Incubation of serum samples with an antibody raised against a portion of human tumor necrosis factor (TNF) resulted in the removal of greater than 90% of serum cytotoxicity, suggesting strongly that the cytotoxic activity was attributable to TNF. These findings are consistent with the hypothesis that TNF is an early acting mediator of the effects of endotoxin in the horse.     

Microvascular circulation of the small intestine in horses.

The microvascular anatomic features of the small intestine was described by correlating results of microangiography, light microscopy, gross studies, and scanning electron microscopy of vascular replicas in 14 horses. After heparinization, the horses were euthanatized, a length of jejunum was transected, and blood was flushed free of the circulation, using isotonic NaCl solution. In six horses, the circulatory system was perfused with a modified radiopaque medium and evaluated radiographically. These sections were then evaluated by standard histologic methods. Sections from 8 horses were perfused with 1 of 2 types of plastics and studied grossly or by scanning electron microscopy. The marginal arterial arcade gives rise to vessels that enter the jejunum at the mesenteric angle. These vessels penetrated either directly, by branching and entering on both sides of the mesenteric angle, or supplying only 1 side of the mesenteric angle. All these vessels continued in the submucosa branching extensively, forming a submucosal plexus. This submucosal plexus supplied the tunica muscularis, tunica serosa, and the mucosa. Vessels within the 2 muscle layers ran parallel to the muscle fibers and, consequently, perpendicular to each other. The arterial supply to the mucosa penetrated the muscularis mucosae and branched to supply 2 mucosal capillary networks. An eccentrically placed arteriole penetrated the base of the villus and spiralled to the tip where it "fountained" into a mesh-like capillary network, which descended peripherally in the villus to drain via 1 to 3, but most commonly 2 venules. Venules from adjacent villi united and drained via the submucosal veins.(ABSTRACT TRUNCATED AT 250 WORDS)     

Microvascular circulation of the descending colon in horses.

The microvascular circulation of the descending colon was studied in 5 adult horses, using microangiography and light microscopy combined with gross studies and scanning electron microscopy of vascular replicas. After heparinization, horses were euthanatized, and 3 segments of the descending colon and its mesentery containing 1 vascular arcade were removed from each horse. The fecal balls were gently massaged from the lumen, and the blood was flushed free of the circulation with isotonic NaCl. In 5 segments, the vascular system was injected with a modified radiopaque medium and evaluated radiographically. Specimens examined radiographically also were prepared for histologic examination, using standard methods. Ten segments were injected with 1 of 2 types of plastics and studied grossly or by scanning electron microscopy. Arcuate arteries gave rise to a descending colonic rete that surrounded the vein and supplied numerous descending colonic lymph nodes. The rete also supplied the mesocolon and the descending colonic tissue. Short filamentous vessels arising from the rete directly penetrated the mesenteric tenia to supply an intermuscular plexus between the longitudinal and circular muscle layers of the muscularis externa. Larger vessels arising from either side of the rete divided into the long- and short-terminal arteries that supplied an extensive submucosal plexus, which was continuous around the circumference. The submucosal plexus supplied the mucosa, the tunica muscularis, and the serosa. Vessels running centrifugally from the submucosal plexus formed an intermuscular plexus between the longitudinal and circular muscle layers of the muscularis externa. The intermuscular plexus at the mesenteric angle also was supplied by vessels branching from the short-terminal arteries as they penetrated the muscularis externa.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isolation and characterization of alpha 1-acid glycoprotein from horses, and its evaluation as an acute-phase reactive protein in horses.

Equine alpha 1-acid glycoprotein (alpha 1AG) was isolated from equine serum by successive ammonium precipitation, anion- and cation-exchange chromatographies, and gel filtration. Purified equine alpha 1AG had a molecular weight of 46,000 +/- 1,000, and contained 31.4% carbohydrate. Gel isoelectric focusing revealed an isoelectric point range of 2.8 to 3.7. With immunoelectrophoresis, it was found that alpha 1AG migrated to the alpha 1-globulin region. Single radial immunodiffusion was used for quantitative measurement of alpha 1AG in equine serum. In clinically normal foals, serum alpha 1AG was undetectable (less than or equal to 20 micrograms/ml) in less than or equal to 7-day-old foals, but was detected by 14 days. The alpha 1AG concentration (mean +/- SD) increased to reach mean adult values of 99.23 +/- 26.90 micrograms/ml by 1 year of age. The alpha 1AG concentration in pregnant mares decreased at 2 to 3 months before parturition, then gradually increased until 1 day after parturition, when a brief decrease was observed. The concentration increased again at 2 weeks after foaling, then a decrease was observed, after which the alpha 1AG concentration increased again by 2 to 4 months after parturition. The concentration of serum alpha 1AG quickly rose to peak values 2 to 3 days after castration and jejunojejunostomy in adult horses, returning to baseline values by 14 to 28 days after surgery. The alpha 1AG was concluded to be an acute-phase reactive protein in horses.     

Microvascular circulation of the cecum in horses

The microvascular circulation of the cecum was studied in 15 adult horses, using microangiography and light microscopy combined with gross studies and scanning electron microscopy of vascular replicas. After heparinization, the horses were euthanatized and the cecum was transected at the cecocolic junction. Blood was flushed free of the circulation with isotonic NaCl and the cecal lumen was slightly distended. In 6 horses, the vascular system was injected with a modified radiopaque medium and evaluated radiographically. Sections evaluated radiographically were also prepared for histologic examination by standard methods. Eight horses were injected with 1 of 2 types of plastics and studied grossly or by scanning electron microscopy. In 1 additional horse, the lateral and medial arteries were injected with different colored plastic for gross studies. The lateral and medial cecal arteries appear to supply the respective areas of the cecum with minimal mixing of the 2 circulations. The major vascular supply to the cecal apex appeared to be through the medial cecal artery. Both the lateral and medial cecal arteries gave rise to cecal retia, which formed a mesh-like network around the respective veins. Vessels from these retia supplied the cecal tissue and the cecal lymph nodes. The continuation of the retia was through long terminal arteries that coursed around the circumference in the submucosa, forming an extensive submucosal plexus. This plexus supplied both the mucosa, and the tunica muscularis and serosa. Vessels within the longitudinal and circular muscle layers of the muscularis externa ran parallel to the muscle fibers, and consequently, perpendicular to each other.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hydrocortisone levels in the urine and blood of horses treated with ACTH.

An investigation was undertaken to demonstrate whether therapeutic treatment with ACTH raises hydrocortisone (cortisol) levels in horse urine above the limit (1000 ng/ml) established by the International Conference of Racing Authorities with the aim of controlling the abuse of cortisol and ACTH in equine sports. ACTH (200 iu) was administered i.m. to 3 Thoroughbred horses; urine and blood samples were collected at intervals afterwards and analysed by an immunoenzymatic system (ELISA) and HPLC-MS. To ascertain post exercise cortisol levels in untreated horses, 101 urine and 103 serum samples were taken from horses immediately after racing and analysed by ELISA. The peak urine level of cortisol, detected 8 h after ACTH administration, was around 600 ng/ml using either ELISA or HPLC-MS. The peak serum cortisol concentration was found to be around 250 ng/ml by ELISA, but consistently less by HPLC-MS. Mean cortisol levels in post race horses were 135.1+/-72.1 ng/ml in urine and 90.1+/-41.7 ng/ml in serum. High levels of the metabolite 20beta-dihydrocortisol in urine and the cortisol precursor 11beta-desoxycortisol in serum were found. The latter showed high cross-reactivity with cortisol on ELISA. In our experiment, treatment with ACTH 200 iu i.m. did not raise urinary cortisol levels above the 1000 ng/ml threshold proposed by the ICRA.     

Microvascular circulation of the ascending colon in horses

Microvascular circulation of the ascending colon in healthy horses was studied using microangiography, light microscopy, and scanning electron microscopy. The pelvic flexure with 30 cm of ventral and dorsal colon attached was removed from 14 adult horses immediately after horses were euthanatized. The lumen was flushed with warm water, and this section of the ascending colon was placed in a 37-C bath of isotonic NaCl. In sections from 8 horses, colic vessels were perfused with a radio-opaque medium for microangiography. After angiographic evaluation, tissue sections were prepared for light microscopic observation, using standard histologic methods. In sections from 6 horses, injection replicas were made by perfusing the vessels with 2 types of plastics. The results of microangiography, light microscopy, and scanning electron microscopy of vascular replicas were correlated, providing a comprehensive documentation of the microvasculature of the ascending colon at the pelvic flexure. Arteries branched from mesenteric colic vessels approximately every 2 cm toward the colonic tissue. Immediately after branching, arterial vessels formed an anastomotic plexus, the colonic rete. However, each branch from the colic vessel eventually continued into the colonic tissue. A second set of vessels originated from the colonic tissue. A second set of vessels originated from the colonic rete and supplied the mesenteric lymph nodes. Arterial vessels penetrated the tunica muscularis into the submucosa 3 to 4 cm toward the antimesenteric border forming a submucosal vascular network. From the submucosal arterioles, branching took place at right angles to supply the mucosal capillaries. Capillaries surrounded the colonic glands and anastomosed at the luminal surface, forming a superficial luminal honeycomb-appearing vascular plexus.(ABSTRACT TRUNCATED AT 250 WORDS)     

Metoclopramide ameliorates the effects of endotoxin on gastric emptying of acetaminophen in horses.

The effect of metoclopramide on gastric emptying of a liquid marker in horses was evaluated by measuring serum concentrations of acetaminophen. Gastric emptying was determined in normal, fasted horses (n = 7), horses given endotoxin intravenously (n = 7), and horses given intravenous metoclopramide plus endotoxin (n = 6). The mean time to reach maximum serum acetaminophen concentration (Tmax), the maximum serum concentration (Cmax), and the area under the serum acetaminophen concentration vs time curve (AUC) were compared among treatment groups. Endotoxin caused a profound delay in gastric emptying, and pretreatment with metoclopramide significantly (P < 0.05) ameliorated this effect.     

Effects of infusion of adenosine triphosphate-magnesium chloride on cardiopulmonary and clinicopathologic variables, cytokine activity, and endothelin concentration in horses administered a low dose of endotoxin

OBJECTIVE: To evaluate systemic effects of i.v. infusion of ATP-MgCl2 subsequent to infusion of a low dose of endotoxin in horses. ANIMALS: 12 adult horses. PROCEDURE: Horses were administered endotoxin (lipopolysaccharide [LPS]) or saline (0.9% NaCl) solution i.v., during a 30-minute period. Immediately thereafter, horses in each group were infused i.v. with ATP-MgCl2 or saline solution. Two weeks later, horses were administered the opposite solution (LPS or saline solution), but it was followed by the same infusion as 2 weeks previously (ie, ATP-MgCl2 or saline solution). Cardiopulmonary and clinicopathologic variables, cytokine activity, and endothelin (ET) concentrations were recorded. RESULTS: IV infusion of ATP-MgCl2 after administration of a low dose of endotoxin failed to attenuate the cardiopulmonary, clinicopathologic, and cytokine alterations that develop secondary to endotoxin exposure. The combination of LPS and ATP-MgCl2 potentiated pulmonary hypertension, leukopenia, and neutropenia when compared with the combination of LPS and saline solution. The combination of LPS and ATP-MgCl2 resulted in thrombocytopenia. Endothelin concentration was increased in jugular venous and pulmonary arterial plasma in horses receiving LPS and ATP-MgCl2. Similar increases were not observed with LPS and saline solution. CONCLUSIONS AND CLINICAL RELEVANCE: Administration of ATP-MgCl2 did not protect horses from systemic effects of experimentally induced endotoxemia. Furthermore, the use of ATP-MgCl2 during endotoxemia may worsen the cardiopulmonary and clinicopathologic status of affected horses. Because ATP and other adenine nucleotides are released from cells during shock, their potential role in the development of hemodynamic derangements, leukocyte adherence, and coagulopathies during endotoxemic episodes warrants further investigation.     

Interleukin-6 and tumour necrosis factor in synovial fluid from horses with carpal joint pathology

The carpal joints are common sites of traumatic arthritis and osteoarthritis (OA) in athletic horses. The pro-inflammatory cytokines interleukin (IL)-6 and tumour necrosis factor (TNF) may be of great importance in the development of intra-articular lesions. The aim of the present study was to investigate possible associations between synovial fluid levels of bioactive IL-6 and TNF and different types of joint lesions seen in traumatic arthritis and OA. Synovial fluid was collected from horses with carpal lameness immediately before arthroscopic surgery. Articular cartilage, synovial membranes and intra-articular ligaments were assessed macroscopically at arthroscopy. Synovial fluid levels of IL-6 and TNF were determined by bioassays, and the cytokine levels between different grades of morphologic changes in each type of assessed tissue were compared. The highest levels of IL-6 were detected in joints with chip fractures. All joints with chip fractures also showed some degree of synovitis. Tumour necrosis factor bioactivity was low and not associated with any joint lesion. Hence, TNF is not useful as a biomarker indicating a specific joint lesion in equine traumatic arthritis or OA. We conclude that a dramatic increase of IL-6 in synovial fluid indicates the presence of osteochondral fragmentation, although low or undetectable levels of IL-6 do not exclude chip fractures. The role of IL-6 in the disease process of osteochondral fragmentation needs further investigation.     

Values of urine specific gravity for thoroughbred horses treated with furosemide prior to racing compared with untreated horses.

The distribution of specific gravity values for 2,599 urine samples collected from racing Thoroughbred horses that were known to have received furosemide prior to racing was compared with that for 1,669 urine samples from racing Thoroughbred horses that reportedly had not received furosemide. Values of specific gravity for furosemide-treated horses were significantly lower (P < 0.001) than those for horses that had not received furosemide, and the proportion of horses with urine specific gravity either <1.010 or <1.012 was significantly greater (P < 0.001) among the furosemide-treated horses. These data indicate that evaluation of urine specific gravity would be a useful component of drug testing programs for regulation of furosemide use.     

Yohimbine ameliorates the effects of endotoxin on gastric emptying of the liquid marker acetaminophen in horses.

The effect of yohimbine pretreatment on gastric emptying of a liquid marker in horses was evaluated by measuring serum concentrations of acetaminophen. Gastric emptying was determined in normal, fasted horses, in horses given endotoxin (E. coli 055 B5; 0.2 microg/kg) intravenously, and in horses given yohimbine (0.25 mg/kg, IV, over 30 minutes) plus endotoxin. Acetaminophen (20 mg/kg) was given by stomach tube 15 minutes after the endotoxin infusion. Blood samples for acetaminophen analysis were collected, and time to reach the peak serum concentration (Tmax), the maximum serum concentration (Cmax) and the area under the acetaminophen serum concentration versus time curve (AUC) were determined for each treatment group. Endotoxin significantly increased Tmax, indicating a profound delay in gastric emptying and yohimbine pretreatment significantly (P < or = 0.05) prevented this effect.     

Characterization of the inflammatory infiltrate and cytokine expression in the skin of horses with recurrent urticaria

Background – Recurrent urticaria (RU) is a common skin disease of horses, but little is known about its pathogenesis. Hypothesis/Objective – The aim of this study was to characterize the inflammatory cell infiltrate and cytokine expression pattern in the skin of horses with RU. Animals – Biopsies of lesional and nonlesional skin of horses with RU (n = 8) and of skin from healthy control horses (n = 8) were evaluated. Methods – The inflammatory cell infiltrate was analysed by routine histology. Immunohistochemistry was used to identify T cells (CD3), B cells (CD79), macrophages (MAC387) and mast cells (tryptase). Expression of T‐helper 2 cytokines (interleukins IL‐4, IL‐5 and IL‐13), a T‐helper 1 cytokine (interferon‐γ), IL‐4 receptor α and thymic stromal lymphopoietin was assessed by quantitative RT‐PCR. Results – In subepidermal lesional skin of RU‐affected horses, increased numbers of eosinophils (P ≤ 0.01), CD79‐positive (P ≤ 0.01), MAC387‐positive (P ≤ 0.01) and tryptase‐positive cells (P ≤ 0.05) were found compared with healthy horses. Subepidermal lesional skin of RU‐affected horses contained more eosinophils (P ≤ 0.05) and tryptase‐positive cells (P ≤ 0.05) compared with nonlesional skin. There was no significant difference in infiltrating cells between nonlesional skin and skin of healthy horses. Expression of IL‐4 (P ≤ 0.01), IL‐13 (P ≤ 0.05), thymic stromal lymphopoietin (P ≤ 0.05) and IL‐4 receptor α (P ≤ 0.05) was increased in lesional skin of RU‐affected horses compared with control horses. Expression of IL‐4 was higher (P ≤ 0.05) in lesional compared with nonlesional RU skin. Conclusions and clinical importance – Analysis of cytokine expression and inflammatory infiltrate suggests that T‐helper 2 cytokines, eosinophils, mast cells and presumptive macrophages play a role in the pathogenesis of equine RU.     

Cyathostome fecal egg count trends in horses treated with moxidectin, ivermectin or fenbendazole

Commercial preparations of fenbendazole (Safe-Guard, Intervet), ivermectin (Eqvalan, Merial) or moxidectin (Quest, Fort Dodge) were administered once to horses scheduled for routine parasiticide treatment. In total, 93 horses from six cooperating farms were used in the study. Computer generated, random allocation of horses to treatment group was conducted at each farm. Fecal egg counts were determined for all horses on trial days 0, 56, 84 and 112, with corresponding calendar dates that were unique to each farm. Only strongyle egg counts from animals which were positive at day 0 were used for analysis of variance and comparisons. Counts for the three treatment groups were similar at day 0, moxidectin相似文献   

Plasma and urine nitric oxide concentrations in horses given below a low dose of endotoxin.

OBJECTIVE: To quantify plasma and urine nitric oxide (NO) concentrations before and after low-dose endotoxin infusion in horses. ANIMALS: 11 healthy adult female horses. Procedure-Eight horses were given endotoxin (35 ng/kg of body weight,i.v.) over 30 minutes. Three sentinel horses received an equivalent volume of saline (0.9% NaCl) solution over the same time. Clinical signs of disease and hemodynamic variables were recorded, and urine and plasma samples were obtained to measure NO concentrations prior to endotoxin infusion (t = 0) and every hour until postinfusion hour (PIH) 6, then every 2 hours until PIH 24. Blood for hematologic and metabolic analyses and for serum cytokine bioassays were collected at 0 hour, every hour until PIH 6, every 2 hours through PIH 12, and finally, every 6 hours until PIH 24. RESULTS: Differences in plasma NO concentrations across time were not apparent, but urine NO concentrations significantly decreased at 4 and 20 to 24 hours in endotoxin-treated horses. Also in endotoxin-treated horses, alterations in clinical signs of disease, and hemodynamic, metabolic, and hematologic variables were significant and characteristic of endotoxemia. Serum interleukin-6 (IL-6) activity and tumor necrosis factor (TNF) concentrations were increased above baseline values from 1 to 8 hours and 1 to 2 hours, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Plasma and urine NO concentrations did not increase in horses after administration of a low dose of endotoxin, despite induction of an inflammatory response, which was confirmed by increased TNF and IL-6 values characteristic alterations in clinical signs of disease, and hematologic, hemodynamic and metabolic variables.     

Exercise-induced pulmonary haemorrhage in the horses: results of a detailed clinical, post mortem and imaging study. VII. Ventilation/perfusion scintigraphy in horses with EIPH

Detailed post mortem examination of the lungs of horses with exercise-induced pulmonary haemorrhage (EIPH) has demonstrated significant small airway disease and intense bronchial arterial proliferation in the dorsocaudal lungfields. The purpose of this study was to investigate ventilation and perfusion distribution in the lungs of a similar group of horses to compare changes in the live animal with the previously reported post mortem findings. Thoracic radiography and ventilation/perfusion (V/Q) scintigraphy were performed on five racing Thoroughbreds with recent histories of EIPH. Parametric images of V/Q ratios for left and right lungfields were also generated from the scan images. In all horses, ventilation and perfusion deficits were demonstrated in the dorsocaudal areas of the lung corresponding closely to the observed radiographic lesions. In particular, the perfusion images and V/Q ratio displays indicated that, in affected areas of lung, pulmonary arterial perfusion was the more seriously impaired. This finding appears to confirm the post mortem evidence of reduced pulmonary arterial perfusion and bronchial arterial dominance in these areas. Ventilation deficits in the same areas also confirmed the likelihood of partial airway obstruction consistent with the small airway disease noted in previous post mortem observations. These results suggest that the vascular and airway lesions demonstrated in detailed post mortems of horses with EIPH are also functionally important in affected horses, even at rest. As a consequence of the apparent persistent, insidious and progressive nature of the lesions associated with EIPH there are serious long term implications for management of the condition.     

Influence of an omega-3 fatty acid-enriched ration on in vivo responses of horses to endotoxin.

Because certain inflammatory processes are dependent on the fatty acid composition of the cellular membrane, dietary manipulations that replace omega-6 fatty acids with omega-3 fatty acids may modify inflammatory responses. We investigated the effect of supplemental dietary linseed oil, containing the omega-3 fatty acid, alpha-linolenic acid, on in vivo responses of horses to endotoxin. One group of horses (n = 6) was fed a control pelleted ration (0% linseed oil), and another group of horses (n = 6) was fed an 8% linseed oil pelleted ration. After 8 weeks of consuming these rations, all horses were given 0.03 micrograms of Escherichia coli 055:B5 endotoxin/kg of body weight, infused over 30 minutes. Horses were monitored over 24 hours. Compared with baseline values within each ration group, endotoxin infusion caused significant (P less than 0.05) increase in rectal temperature, heart rate, and plasma concentration of thromboxane B2, 6-keto-prostaglandin F1 alpha, and fibrinogen and significant (P less than 0.05) decrease in total WBC count. Compared with baseline values within each ration group, endotoxin infusion failed to cause significant changes in prothrombin, activated partial thromboplastin, thrombin, or whole blood recalcification times, serum concentration of fibrin degradation products, PCV, or plasma total protein concentration. Before and after endotoxin infusion, horses given the linseed oil ration had longer mean whole blood recalcification time and activated partial thromboplastin time than did horses fed the control ration.     

Evaluation of cytokine mRNA expression in bronchoalveolar lavage cells from horses with inflammatory airway disease

Inflammatory airway disease (IAD) is a common disorder of performance horses and is associated with poor performance and accumulation of mucus and inflammatory cells in lower airway secretions. Horses with IAD frequently have increased relative counts of neutrophils in bronchoalveolar lavage fluid (BALF); less commonly relative counts of eosinophils and/or mast cells may be increased. The aetiopathogenesis of IAD is unknown and may involve innate and/or acquired immune responses to various factors including respirable dust constituents, micro-organisms, noxious gases and unconditioned air. The molecular pathways and role of the immune system in the pathogenesis of IAD remain poorly defined and it is unknown whether polarised T cell responses occur in the disease, as have been reported to occur in equine recurrent airway obstruction and asthma in humans. Elucidating cytokine responses that develop in horses with IAD may allow a greater understanding of the possible aetiopathological pathway(s) involved and could contribute to development of novel treatments. We compared the mRNA expression of tumour necrosis factor-alpha (TNF-α), interferon-gamma (IFN-γ), interleukin (IL)-1β, IL-2, IL-4, IL-8, IL-13, IL-17 and IL-23 in cell pellets extracted from BALF of horses with IAD (n=21) and horses free of respiratory tract disease (n=17). Horses with IAD had significantly increased levels of TNF-α, IL-1β and IL-23 mRNA; no significant differences in the other cytokine mRNAs were detected. The results of this study indicate that IAD of horses is associated with increased mRNA expression of pro-inflammatory cytokines in BALF cells, which may reflect stimulation of the innate immune responses to inhaled antigens. There was no evidence of a polarised T-cell cytokine response suggesting hypersensitivity responses may not be involved in the aetiopathogenesis of IAD.