Cytological analysis of equine bronchoalveolar lavage fluid. Part 3: The effect of time,temperature and fixatives

Bronchoalveolar lavage fluid (BALF) samples are often subject to time delays, possibly with temperature fluctuations, between collection and processing. The aim of this study was to evaluate the effects of time, temperature and 2 different fixatives on equine BALF cytology, in order to develop guidelines for optimal equine BALF storage conditions. Total nucleated cell count (TCC), differential cell counts (DCC), absolute cell counts (ACC), cell viability, cell morphology and bacterial growth of BALF samples stored at 4, 18 (+/- addition of formalin- or alcohol-based fixatives) and 38 degrees C were monitored serially over a 72 h period. The time taken for a significant reduction in TCC and cell viability of unfixed BALF samples decreased as the storage temperature increased. There was no diagnostically significant difference in DCC or ACC over this time-course at any temperature. Unfixed BALF samples showed significant bacterial growth by 24 h at 4 degrees C, and 8 h at 18 and 38 degrees C; and poor morphology by 48 h at 4 degrees C, 24 h at 18 degrees C and 8 h at 38 degrees C. Fixed BALF samples showed poor morphology with Leishman's stain compared to unfixed samples.     

Cytological analysis of equine bronchoalveolar lavage fluid. Part 2: Comparison of smear and cytocentrifuged preparations

The aim of this study was to develop a diagnostically useful smear method for preparation of equine bronchoalveolar lavage fluid (BALF) for use by practitioners. A smear method for equine BALF preparation which included the addition of serum was developed, and cell morphology, differential cell counts (DCC) and repeatability of counting DCC compared with those of cytocentrifuged BALF preparations. BALF samples (n = 21) were collected from 5 control horses and 5 heaves-susceptible horses. Smear preparations of BALF produced smaller, darker, staining cells, making cytological identification more difficult than on cytocentrifuged preparations. There was a significantly higher (P<0.01) macrophage DCC and lower lymphocyte DCC on cytocentrifuged compared to smear preparations. Mast cell and eosinophil DCC were significantly higher (P<0.05) on cytocentrifuged compared to smear preparations of BALF. Smear preparations were shown to be reliable for the cytological diagnosis of equine neutrophilic pulmonary disease and offer practitioners an alternative to sending equine BALF to a laboratory for processing and cytological analysis.     

Airway inflammation and mucus in two age groups of asymptomatic well-performing sport horses

REASONS FOR PERFORMING STUDY: Horses housed in conventional stable environments are exposed to high dust loads. Long-term exposure to inhaled dust may lead to subclinical airway disease. HYPOTHESES: Airway inflammation and mucus accumulation are 1) common in asymptomatic well-performing sport horses, 2) increased in older compared to younger individuals and 3) correlated. METHODS: Mucus quantity and quality (endoscopic scoring) and BALF differential cytology were assessed in 13 younger horses (mean age 5 years) and 13 older horses (mean age 15 years), which had no historical or clinical evidence of lower airway disease, were performing well and were housed permanently in a conventional stable environment. RESULTS: 1) Airway inflammation and/or mucus accumulation were very common. All sport horses showed evidence of subclinical inflammatory airway disease (IAD). 2) Older horses, having been exposed for 10 more years to a conventional stable environment, did not show increased subclinical airway inflammation or mucus accumulation. The only differences between the age groups were increased BALF total and lymphocyte cell counts in the younger horses. 3) Mucus quantity and quality scores were not significantly correlated with each other or with BALF neutrophil percentages and absolute numbers. CONCLUSIONS: Although clinically healthy and performing well, all of the examined horses housed in a conventional stable environment showed evidence of IAD. The degree of IAD was not increased in older horses that had not developed clinical signs. POTENTIAL RELEVANCE: The effect of subclinical degrees of IAD on performance in dressage and show jumping should be investigated further to better understand the clinical significance of IAD in sport horses.     

Lung function and airway cytologic profiles in horses with recurrent airway obstruction maintained in low-dust environments

BACKGROUND: The effects of long-term environmental management on airway obstruction and inflammation in horses with recurrent airway obstruction (RAO) are unknown. HYPOTHESIS: Horses with RAO maintained in low-dust environments have persistent airway obstruction and neutrophilic inflammation. ANIMALS: Study horses were treated for RAO and then maintained in low-dust environments with no medical management. Horses were classified into 3 groups by years after diagnosis: 1 year (time 1, n = 9), 2-3 years (time 2, n = 7), and 5-6 years (time 3, n = 8). The comparison groups were age-matched healthy horses. METHODS: In this cross-sectional study, a clinical examination was performed, and the clinical score was calculated. Standard lung function, forced expiratory maneuvers, and the cytology of bronchoalveolar lavage fluid (BALF) were evaluated. RESULTS: The clinical scores of the RAO horses were higher than those of the non-RAO horses at time 2 (P = .018). Standard lung function data were not different between the groups at any time point. The forced expiratory flow between 75-95% of exhaled vital capacity was lower in RAO horses than in non-RAO horses at all time points (P < .02), indicating persistent peripheral airway obstruction. Cytologic evaluation of BALF revealed no difference in total nucleated cell numbers or differential cell counts between RAO and non-RAO horses at any time point. CONCLUSIONS AND CLINICAL IMPORTANCE: The peripheral airway obstruction detected in horses with RAO maintained in low-dust environments likely is due to irreversible airway remodeling but is not associated with cytologic evidence of airway inflammation.     

Bronchoalveolar lavage fluid cytologic findings in horses with pneumonia or pleuropneumonia

Bronchoalveolar lavage was performed in 22 horses with pneumonia or pleuropneumonia. All horses had clinical evidence of pneumonia or pleuropneumonia on the basis of physical, radiographic, ultrasonographic, tracheobronchial aspirate or post-mortem findings. Results of lavage fluid analysis were normal in 9 horses, equivocal in 3 horses, and abnormal in 10 horses. Abnormal lavage fluid had increased total cell count, increased relative and absolute neutrophil counts, degenerative neutrophils, and decreased relative and absolute macrophage and lymphocyte counts.     

Post-exercise endoscopic and cytologic diagnosis of equine asthma syndrome in asymptomatic Brazilian pacers

The objectives were: 1. to investigate the occurrence of equine asthma syndrome (EAS) in a group of Campolina and Mangalarga Marchador horses through post-exercise respiratory endoscopy and cytology; 2. compare the efficiency of different diagnostic criteria; and 3. compare the incidence of EAS by breed, age group and sex. A standardised exercise test was conducted in the field, followed by respiratory endoscopy and bronchoalveolar lavage and fluid collection 30 min post-exercise. Clinical evaluation pre- and post-exercise, respiratory endoscopy post-exercise and bronchoalveolar lavage, also post-exercise, were conducted in 30 horses (15 Campolina and 15 Mangalarga Marchador) aged 3–18 years, without a history of respiratory illness and without clinical evidence of active respiratory disease. All horses were in active training and were considered fit to participate in equestrian competition characteristic of these breeds. Post-collection, the horses were subdivided into healthy or EAS-positive, depending on several diagnostic criteria. Statistical comparisons between breeds, age brackets (up to 7 years of age and above 7 years of age) and sex were conducted using appropriate statistical tests. According to bronchoalveolar lavage fluid (BALF) cytology, 56.7% of the horses showed at least one inclusion criteria that contributed to classification as EAS-positive. Of those, only one was classified as moderate EAS, while the others were classified as mild. Total cell count in the BALF above 530 cells/µL was the single most common diagnostic criteria (100%), followed by increased tracheal mucus (53%) at endoscopy, eosinophilia (47%) and neutrophilia (35%) in BALF. Significant differences were found between macrophage and lymphocyte numbers according to breed, but no other differences between breeds, age brackets, sex or status (healthy vs. EAS-positive). A high occurrence of EAS was found in competition-ready Mangalarga and Campolina pacer horses in this study.     

Platelet‐Activating Factor and Evidence of Oxidative Stress in the Bronchoalveolar Fluid of Thoroughbred Colts during Race Training

Background: Inflammatory airway disease (IAD) is prevalent in young racehorses during training, being the 2nd most commonly diagnosed ailment interrupting training of 2‐year‐old Thoroughbred racehorses. Hypothesis: That stabling and exercise cause oxidative stress, release of platelet‐activating factor (PAF) and inflammation in airways of Thoroughbred colts. Animals: Colts in breeding farms (NC, n = 45), stabled for 30 days (EC, n = 40), and race trained (EX, n = 34). Methods: Cytological profile and parameters of bronchoalveolar lavage fluid (BALF) related to oxidative stress, bioactivity of the proinflammatory mediator PAF, catalase activity, and alveolar macrophage function. Results: Percentages of neutrophils and eosinophils in the BALF of the EX group were higher (5.4 ± 6.4% versus 0.9 ± 1.2%) than the upper limits for normal horses (3–5%). BALF from the EX group (45.6 ± 2.8 cells/μL of BALF) also displayed significantly (P= .017) higher total nucleated cell count. PAF bioactivity and the total protein concentration in the BALF were higher in the EX group (0.0683 ± 0.076 versus 0.0056 ± 0.007 340 : 380 nm ratio P= .0039, 0.36 ± 0.30 versus 0.14 ± 0.15 mg of proteins/mL of BALF P < .001). Concentration of BALF hydroperoxides was higher in the EC group (104.7 ± 80.0 versus 35.2 ± 28.0 nmol/mg of proteins, P= .013) and catalase activity was higher in the EX group (0.24 ± 0.16 versus 0.06 ± 0.02 μmol H₂O₂/min/mg of proteins, P= .0021). Alveolar macrophage phagocytosis (P= .048) as well as production of superoxide anion (P= .0014) and hydrogen peroxide (P= .0011) were significantly lower in EX group. Conclusions and Clinical Importance: Further studies should be performed to elucidate the role of PAF in the pathophysiology of IAD. Its presence in bronchoalveolar fluid of young athletic horses makes it a potential therapeutic target to be investigated.     

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.     

Inhalation of organic dusts and lipopolysaccharide increases gelatinolytic matrix metalloproteinases (MMPs) in the lungs of heaves horses

We report the effects of mouldy hay/straw exposure, inhaled hay dust suspension (HDS) and inhaled lipopolysaccharide (LPS) on bronchoalveolar lavage fluid (BALF) gelatinolytic matrix metalloproteinase (MMP) levels and degree of activation in healthy (n = 6) and heaves- (previously termed chronic obstructive pulmonary disease) affected (n = 6 or 7) horses. Gelatinolytic MMPs in BALF were quantified by zymography, and gelatinases were shown by Western immunoblotting to be MMP-2 and MMP-9. Hay/straw and HDS challenges increased BALF total gelatinolytic activity only in heaves horses, with the majority of gelatinolytic activity comprising pro- and active MMP-9. The 5 h duration hay/straw challenge increased BALF gelatinolytic MMP activity in heaves horses at 5 and 24 h after the start of this challenge, with activity returning to baseline by Day 4. In contrast to hay/straw and HDS challenges, LPS inhalation increased BALF gelatinolytic MMP activity in both groups. For all challenges, absolute BALF neutrophil counts were highly significantly correlated (P<0.0001) with levels of proMMP-9 and active MMP-9, but not with levels of MMP-2 (P>0.05). As gelatinolytic MMPs are pro-inflammatory agents, they may contribute to lung dysfunction and tissue destruction in heaves horses exposed to airborne organic stable dusts.     

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.     

Assessment of respiratory herd health in weaner pigs by measuring cellular composition of bronchoalveolar lavage fluid

Cell differentiation of bronchoalveolar lavage fluid (BALF) was compared among 11 herds having a history of recurrent respiratory disease in weaner pigs and nine herds lacking such a history. In every herd, 20 pigs aged 8-10 weeks were lavaged. The two groups differed significantly on median percentage of macrophages, neutrophils and lymphocytes, but not on white blood cell count of the BALF. Logistic regression showed the percentage of samples per herd exceeding the reference value for neutrophils of 0-8% to be the most promising parameter to assess the health status in weaner pigs.     

Effect of repetitive bronchoalveolar lavage on cytologic findings in healthy dogs

OBJECTIVE: To determine reference values for cytologic examination results of bronchoalveolar lavage fluid (BALF) and to investigate effects of repeated lavages on pulmonary health and on results of cytologic examination of BALF in dogs. ANIMALS: 16 healthy adult Beagles. PROCEDURE: All dogs underwent pulmonary lavage to obtain BALF. Eleven dogs were repeatedly lavaged 6 times at 5- to 7-week intervals. Analyses for total and differential cell counts and for viability of cells before and after cell processing were performed. Arterial blood gas analysis before and after bronchoalveolar lavage was used to study the safety of the lavage procedure. Histologic and radiologic examinations were used to study effects of repeated lavages on pulmonary health. RESULTS: Mean (+/- SD) cell count was 104 +/- 69 cells/microl, comprising 75 +/- 7% alveolar macrophages, 13 +/- 6% lymphocytes, 5 +/- 4% neutrophils, 4 +/- 5% eosinophils, 2 +/- 2% mast cells, 0.6 +/- 0.7% epithelial cells, and 0.3 +/- 0.4% plasma cells. Centrifugation of samples and washing of cells caused significant cell loss (59 +/- 13%). Repeated lavages did not cause significant variations in cell counts of BALF or results of arterial blood gas analysis, thoracic radiography, or histologic examination of pulmonary specimens. Only a moderate, although significant, decrease in arterial oxygen content was observed after bronchoalveolar lavage. CONCLUSIONS AND CLINICAL RELEVANCE: Analysis indicated that several lavages performed at 5- to 7-week intervals can safely and reliably be used to study the kinetics of pathologic processes in pulmonary tissues or for evaluation of therapeutic efficacy.     

Abnormalities in lung surfactant in horses clinically affected with recurrent airway obstruction (RAO)

Background: Abnormalities in lung surfactant are well described in human respiratory diseases including asthma, but are poorly described in horses. Hypothesis: Lung surfactant is abnormal in horses with clinical signs of recurrent airway obstruction (RAO). Animals: Six healthy horses and 5 horses with RAO. Methods: Bronchoalveolar lavage fluid (BALF) was obtained from all horses by standard procedures. Cell‐free BALF was separated into crude surfactant pellets (CSP) and supernatant via ultracentrifugation. Phospholipid and protein content was analyzed from both of these fractions. Phospholipid composition of CSP was determined using high‐performance liquid chromatography with an evaporative light scatter detector. Surface tension of CSP was measured with a pulsating bubble surfactometer. Results: Compared with healthy horses, surfactant from RAO‐affected horses was characterized by significantly decreased phospholipid content in total surfactant (median; range: 23.2; 14.7–62.2 μg/mL BALF versus 172; 111–267 μg/mL BALF, P= .0062) and CSP (20.2; 6.4–48.9 μL/mL BALF versus 155; 94.4–248 μg/mL BALF, P= .0062), and a significantly lower percentage of phosphatidylglycerol (PG) (4.5; 3.6–5.6% versus 6.6; 4.1–7.6%, P= .028). Furthermore, the ratio between the percentages of phosphatidylcholine and PG was significantly higher in RAO‐affected horses than in healthy horses (20.9; 16.6: 25.9 versus 13.9; 11.8–22.8, P= .045). Conclusions and Clinical Importance: This study demonstrates that surfactant from RAO‐affected horses is abnormal. Further studies are needed to determine if these abnormalities are related to an increased tendency for bronchoconstriction and to a decreased ability to clear airway mucus in RAO‐affected horses.     

Efficacy of three corticosteroids for the treatment of heaves

This study used a cross-over design to compare the efficacy of 3 corticosteroids for the relief of airway obstruction and inflammation in 9 heaves-affected horses. The severity of airway obstruction and inflammation was quantified by measurement of lung function and by bronchoalveolar lavage fluid (BALF) cytology, respectively. Airway obstruction was induced by stabling the horses and they remained stabled during the 10 day treatment period. Lung function was measured before treatment (baseline), at Days 3, 7, and 10 of treatment, and after 30 days at pasture. BALF cytology was investigated at baseline, Day 10, and at pasture. All 9 horses received the following 4 treatments in random order: no treatment, daily oral prednisone tablets (1 mg/kg), daily i.v. dexamethasone solution (0.1 mg/kg), and i.m. dexamethasone-21-isonicotinate (0.04 mg/kg) every 3 days. When horses received no treatment, lung function did not change significantly during stabling but improved at pasture. In all horses, daily i.v. administration of dexamethasone solution improved lung function within 3 days to levels as good as or better than those measured at pasture. Dexamethasone-21-isonicotinate was rapidly effective in 8 of 9 horses. The other horse did not respond to this drug. Prednisone tablets were without effect on Days 3 and 7 of treatment, but by Day 10, 5 of 9 horses showed some improvement in lung function. Dexamethasone i.v. solution decreased the percent neutrophils in BALF at Day 10. Other treatments had no effect on BALF cytology. These results demonstrate that dexamethasone rapidly relieved airway obstruction in heaves-affected horses. Oral prednisone had inconsistent effects but may be beneficial in some horses after more than a week of treatment.     

Cytologic, microbiologic, and biochemical analysis of bronchoalveolar lavage fluid obtained from 24 healthy cats

Twenty-four healthy cats underwent bronchoscopy and bronchoalveolar lavage to determine the normal cytologic environment of the lower respiratory tract of cats. Initial screening to ensure the health of the study population included complete histories, physical examinations, thoracic radiography, CBC, serologic tests for feline leukemia virus, feline immunodeficiency virus, and occult heartworm, and sugar and Baermann fecal flotation. In 18 cats, protected catheter brush samples of airway secretions from the lavaged lung segment were taken for culture of aerobic and anaerobic bacteria and mycoplasma. Bronchial lavage fluid (5 sequential 10-ml aliquots of normal saline solution) was pooled and filtered with cotton gauze. The unspun sample was used for determination of a total nucleated cell count. Lavage fluid was cytocentrifuged and 500 cells/slide were scored for determination of the cellular differential. Activity of lactate dehydrogenase and concentrations of total protein and IgG within the supernatant were measured, and assays were performed to detect the presence of IgA and IgM. Complete histologic evaluation of the lavaged lung of each of 6 random-source cats was performed after differential cell counting revealed 18% eosinophils within bronchoalveolar lavage fluid recovered from this group. Alveolar macrophages were the predominant cells encountered; however, a quarter of all cells recovered were eosinophils. A significant relationship was not found between the abundance of eosinophils in the lavage fluid, and either isolation of aerobic bacteria, high total nucleated cell counts, total protein concentrations, or activity of lactate dehydrogenase. Histologic evaluation of the lungs of 5 of 6 random-source cats revealed normal lungs in 2 cats, and minimal abnormal change in 3 others. Evaluation of the lungs from 1 random source cat revealed acute, mild eosinophilic bronchiolitis. We conclude that large numbers of eosinophils may be retrieved from the bronchoalveolar lavage fluid of healthy cats.     

Influence of Age on Surfactant Isolated from Healthy Horses Maintained on Pasture

Background: Surfactant alterations are described in horses after exercise, anesthesia, and prolonged transport, in horses with recurrent airway obstruction, and in neonatal foals. The effect of horse age or bronchoalveolar lavage fluid (BALF) sample characteristics on surfactant is unknown.
Objectives: To evaluate surfactant phospholipid composition and function in healthy horses, and to investigate the influence of age and BALF sample characteristics on surfactant.
Animals: Seventeen healthy horses 6–25 years of age maintained on pasture year-round.
Methods: BALF was collected by standard procedures and was assessed for recovery volume, nucleated cell count (NCC), and cytology. Cell-free BALF was separated into crude surfactant pellet (CSP) and surfactant supernatant (Supe) by ultracentrifugation. Phospholipid and protein content were determined from both fractions. CSP phospholipid composition was analyzed by high-performance liquid chromatography with an evaporative light scatter detector. Surface tension of CSP was evaluated with a pulsating bubble surfactometer. Regression analysis was used to evaluate associations between age, BALF sample characteristics, and surfactant variables.
Results: Results and conclusions were derived from 15 horses. Increasing age was associated with decreased phospholipid content in CSP but not Supe. Age did not affect protein content of CSP or Supe, or surfactant phospholipid composition or function. Age-related surfactant changes were unaffected by BALF recovery percentage, NCC, and cytological profile.
Conclusions and Clinical Importance: Older horses have decreased surfactant phospholipid content, which might be because of age-related pulmonary changes. Surfactant composition is unaffected by BALF sample characteristics at a BALF recovery percentage of at least 50%.     

IgG antibody responses to an inhaled antigen in horses with "heaves" (recurrent airway obstruction).

A controlled experimental system for the evaluation of pulmonary immune responses in horses with "heaves" (recurrent airway obstruction) has been developed. We hypothesized that the humoral immune response to an inhaled antigen in diseased horses would be different from that of healthy horses and that chronic pulmonary inflammation would bias the production of IgG isotypes in diseased horses as compared to healthy horses. Healthy and affected horses were housed in a natural challenge environment (stabled, fed dusty hay) and exposed by inhalation, to a nebulized solution of keyhole limpet hemocyanin (KLH). Sera and bronchoalveolar lavage fluids (BALFs) were collected from horses prior to and following their inhalation exposure to the antigen. Differential cell counts were performed on the cells in the BALF. An enzyme-linked immunosorbent assay (ELISA) was used to determine the concentrations of IgGa, IgGb, IgG(T) and combined IgG specific for KLH in the sera and BALF. The percentages of neutrophils in the BALF of diseased horses were increased 4-6-fold over healthy horses. Combined IgG specific for KLH was significantly greater in BALF and serum from healthy compared to diseased horses. Differences in isotypes were also evident; however, only IgGb specific for KLH in the BALF was significantly increased in healthy versus diseased horses. Possible explanations for this difference include: (1) increased destruction of antigen before it could interact with lymphocytes, (2) down-regulation of IgGb production by inhibitory cytokines in diseased horses, or (3) binding of IgGb to Fc receptors on the large numbers of neutrophils in the lungs of diseased horses. In contrast to the prevailing notion that horses with heaves have exaggerated immune responses, our data suggest that diseased horses exposed to an aerosolized protein mount weaker IgG responses compared to healthy horses.     

Storage alters feline bronchoalveolar lavage fluid cytological analysis

Bronchoalveolar lavage fluid (BALF) collection is a valuable respiratory diagnostic procedure in cats. This study evaluated effects of BALF storage on total nucleated cell counts (TNCCs) and differential cell counts (DCC), cell morphology, and cytological diagnosis. Forty-five research cats with neutrophilic, eosinophilic, and mixed inflammation, and healthy controls were enrolled. BALF samples were processed within 1h (baseline) or stored at 4°C (4C24) or room temperature (RT24) for 24h, or 4°C (4C48) or room temperature (RT48) for 48h before processing. Stored BALF at RT48 had decreased TNCC compared to baseline. The RT24 and RT48 samples had greater eosinophil % and the RT24, 4C48, and RT48 samples had decreased neutrophil % compared with baseline. Cellular morphology deteriorated in all stored samples. Storage resulted in a change in cytological diagnosis in up to 57% of stored samples. We conclude that cytological analysis of BALF in cats should be performed promptly for optimal results.     

Influence of subclinical inflammatory airway disease on equine respiratory function evaluated by impulse oscillometry

Reasons for performing study: Inflammatory airway disease (IAD) is a nonseptic condition of the lower respiratory tract. Its negative impact on respiratory function has previously been described using either forced expiration or forced oscillations techniques. However, sedation or drug‐induced bronchoconstriction were usually required. The impulse oscillometry system (IOS) is a noninvasive and sensitive respiratory function test validated in horses, which could be useful to evaluate IAD‐affected horses without further procedures. Objectives: To determine the sensitivity of IOS in detecting alterations of the respiratory function in subclinically IAD‐affected horses without inducing bronchoprovocation and to characterise their respiratory impedance according to frequency for each respiratory phase. Methods: Pulmonary function was evaluated at rest by IOS in 34 Standardbred trotters. According to the cytology of bronchoalveolar lavage fluid (BALF), 19 horses were defined as IAD‐affected and 15 horses were used as control (CTL). Total respiratory resistance (Rrs) and reactance (Xrs) from 1–20 Hz as well as their inspiratory and expiratory components were compared between groups. Results: A significant increase of Rrs at the lower frequencies (R_{1–10 Hz}) as well as a significant decrease of Xrs beyond 5 Hz (X_{5–20 Hz}) was observed in IAD compared to CTL horses. IOS‐data was also significantly different between inspiration and expiration in IAD‐affected horses. In the whole population, both BALF eosinophil and mast cell counts were significantly correlated with IOS measurements. Conclusions: Functional respiratory impairment may be measured, even in the absence of clinical signs of disease. In IAD‐affected horses, the different parameters of respiratory function (Rrs or Xrs) may vary depending on the inflammatory cell profiles represented in BALF. Potential relevance: Impulse oscillometry could be used in a routine clinical setting as a noninvasive method for early detection of subclinical respiratory disease and of the results of treatment in horses.