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Biochemical and cytological analysis of bronchoalveolar lavage (BAL) fluid and effects on arterial blood gases in dogs with lower respiratory airway disease R. GONUL1*, L. KOENHEMSI1, M.E. OR1, A. UYSAL1, K. SONMEZ2, A. GUREL2, A.F. BAGCIGIL3, N.Y. OZGUR3, H. YARDIBI4, K. ALTUNATMAZ5 Department of Internal Medicine, Veterinary Faculty, Istanbul University, 34 320 Avcilar, Istanbul, TURKEY. Department of Pathology, Veterinary Faculty, Istanbul University, 34 320 Avcilar, Istanbul, TURKEY. Department of Microbiology, Veterinary Faculty, Istanbul University, 34 320 Avcilar, Istanbul, TURKEY. 4 Department of Biochemistry, Veterinary Faculty, Istanbul University, 34 320 Avcilar, Istanbul, TURKEY. 5 Department of Surgery, Veterinary Faculty, Istanbul University, 34 320 Avcilar, Istanbul, TURKEY. 1 2 3 *Corresponding author: gonul@istanbul.edu.tr SUMMARY The aims of this study were to investigate the effects of BAL collection on the respiratory function in healthy dogs and in dogs suffering from lower airway respiratory diseases and to consider the potential diagnostic value of some biochemical and cytological parameters measured in BAL fluids. For that, endoscopy and BAL collection were performed under anaesthesia induced with medetomidin (40 μg/kg IM) and propofol (1 mg/kg IV) in dogs with pulmonary disorders (n = 30) and in healthy dogs (n = 10). The evaluation of the respiratory function was made throughout determination of blood gas and acid-base balance before anaesthesia and directly after BAL fluid sampling. In parallel, the effects of the anaesthesia alone on the respiratory system were also assessed in healthy dogs (n = 10) which were not submitted to the BAL collection. The anaesthesia protocol instead of the BAL collection by itself induced hypoventilation and low O2 exchange between alveoli and arteries as evidencing by significant decreases of PaO2 and O2Sat and significant increases of PaCO2 and arterial-alveolar PO2 gradient (A-aPO2). In diseased dogs, PaO2/O2Sat and the A-aPO2 were initially affected and under anaesthesia, variations of O2Sat, PaCO2 and base deficit were aggravated whereas changes in PaO2 and A-aPO2 were less pronounced compared to healthy controls. Significant increases of LDH, ALT and ALP activities and of urea concentrations in BAL fluids from diseased dogs coupled to a high cellularity (epithelial and inflammatory cells) and positive bacterial isolation in some cases have confirmed the inflammatory and/or infectious origin of the pulmonary diseases. Although biochemical and cytological analysis of the BAL fluids can help to characterize the pulmonary disease, its collection under anaesthesia may require some attention in diseased dogs because of its direct effects on respiratory function. Keywords: Dog, bronchoalveolar lavage, arterial blood gases, anaesthesia, biochemistry, cytology. RÉSUMÉ Analyse biochimique et cytologique du liquide de lavage bronchoalvéolaire (LBA) et effets sur les gaz sanguins chez les chiens atteints d’une maladie des voies aériennes basses Cette étude a eu pour objectifs d’évaluer les effets directs d’un lavage bronchoalvéolaire sur la fonction respiratoire chez des chiens sains ou soufrant de maladie pulmonaire et de considérer la valeur diagnostique potentielle de divers paramètres biochimiques et cytologiques mesurés dans le liquide de lavage broncho-alvéolaire (LLBA). Pour cela, une endoscopie et un lavage broncho-alvéolaire ont été réalisés sous anesthésie [médétomidine (40 μg/kg IM) et propofol (1 mg/kg IV)] chez des chiens malades (n = 30) et sains (n = 10). La fonction respiratoire a été évaluée par détermination des gaz sanguins et de l’équilibre acido-basique avant l’anesthésie et directement après le LBA. Les effets sur le système respiratoire de l’anesthésie seule ont également été appréhendés sur des chiens sains (n = 10) non soumis à un lavage bronchoalvéolaire. Le protocole d’anesthésie, plutôt que le lavage par lui-même, a induit une hypoventilation et une diminution des échanges en O2 entre les alvéoles et les artères (diminutions significatives de PaO2 et O2Sat et augmentations significatives de PaCO2 et du gradient en oxygène artério-alvéolaire (A-aPO2)). Chez les chiens malades, PaO2/O2Sat et A-aPO2 étaient initialement modifiés et sous anesthésie, les variations en O2Sat, PaCO2 et le déficit basique se sont aggravés alors que les modifications de la PaO2 et de l’A-aPO2 ont été moins importantes que chez les sujets contrôles. L’origine inflammatoire et/ou infectieuse des maladies pulmonaires a été confirmée par des augmentations significatives des activités LDH, ALT, PAL et des concentrations en urée dans les liquides de lavage chez les chiens malades associées à une forte cellularité (cellules épithéliales et inflammatoires) et dans quelques cas à une isolation bactérienne positive. Bien que les analyses biochimiques et cytologiques des liquides de lavage puissent aider à la caractérisation de la maladie pulmonaire, son recueil sous anesthésie requiert une attention particulière chez les malades en raison de ses effets directs sur la fonction respiratoire. Mots clés : Chien, lavage bronchoalvéolaire, gaz sanguins artériels, anesthésie, biochimie, cytologie. Introduction Bronchoscopy and bronchoalveolar lavage (BAL) may be used for the diagnosis and treatment of respiratory diseases in humans and small animals [3, 6-8, 13, 14]. BAL is a reliable Revue Méd. Vét., 2010, 161, 5, 233-238 procedure which helps in diagnosing the disorders of the respiratory system, and in the studies performed, researchers have analyzed the BAL fluid with regard to enzyme, cytology, culture and histopathology [2, 5-8, 12-14, 18, 20, 21]. Despite its widespread use, an agreement has not been achieved on 234 the laboratory techniques that are used, the fluid volume and the technical issues such as various lavage applications [17]. For this reason, there are various reference values for dogs and the results of studies with various methods make the comparison difficult [17]. Detection of alterations in enzyme activities and identification of the cellular components in BAL fluid are useful for the diagnosis of pulmonary inflammations and lacerations [9, 12]. Epithelial and inflammatory cells in BAL fluid are the most sensitive indicators of the inflammatory response and they evidence pathological alterations in pulmonary parenchyma. Similarly, detection of cytoplasm and membrane enzymes in the acellular portion of the BAL fluid is an indicator of cell death or membrane damage [9, 12]. Furthermore, despite the fact that studies related to immunoglobulins in the respiratory tract have been performed to study local immune mechanisms in chronic bronchitis, no article on dogs has been available. Moreover, the effects of BAL applications on blood gases in dogs with respiratory tract disorders have not been sufficiently studied [18]. Accurate analysis of blood gases facilitates the clinical evaluation of metabolic acid-base and respiratory system disorders [10, 16]. It is also helpful in making a proper diagnosis and plan of treatment by enabling the distinction of ventilation (carbon dioxide) and oxygenation (oxygen) problems [16]. In this way, the arterial-alveolar PO2 gradient (A-aPO2) can be used to evaluate the degree of pulmonary disorder [16]. The arterial-alveolar PO2 gradient (A-aPO2) increases in right to left shunt, low mixed venous oxygen saturation, ventilation/perfusion non-conformance, and diffusion insufficiencies [19]. However, researchers have reported a decrease of arterial oxygen pressure following BAL in cats, dogs and humans [7, 17, 18]. Besides, RAJAMAKI et al. [18] have reported that although the effects of the BAL procedure on blood gases have been studied in healthy animals, they have not been studied in dogs with respiratory tract disorders, and for this purpose, they have studied dogs in which pulmonary eosinophilia had been detected. On the other hand, the cardiopulmonary effects of anaesthetic drugs that are used are not completely known [4]. The aims of the present study are firstly to compare blood gases analysis and oxygen saturation, acid-base balance and electrolyte concentrations between dogs with lower respiratory tract disorders and apparently clinically healthy dogs, secondly to evaluate the diagnostic values of some biochemical markers (Total protein, urea, creatinine, Ca/P and enzyme LDH, ALP, ALT and GGT activities, immunoglobulins (IgA, IgG and IgM) and cellular constituents (bacteriological and various cell populations) in BAL fluid and thirdly to investigate the effects of anaesthesia with medetomidin and propofol during endoscopy application on blood gases in healthy and sick dogs. Materials and Methods ANIMALS AND PROTOCOL DESIGN The study was conducted in accordance with the ethical committee principles and with the approval of the Ethical GONUL (R.) AND COLLABORATORS Committee of the Veterinary Faculty of Istanbul University (2006-39). A total of 30 sick dogs of various breeds and ages, and of both sexes have been brought to the internal medicine clinic, Veterinary Faculty, Istanbul, Turkey for respiratory tract complaints. Diagnosis of lower respiratory tract disorders such as chronic broncho-pneumonia was based on clinical and radiological examinations. This group constituted the study group (BAL-diseased group). Additionally, twenty dogs without any respiratory tract disorder throughout routine clinical examination were divided into 2 control groups: the BAL characteristics were evaluated in one group, named BAL-control group (n = 10) and in the other, called Anaesthesia-control group (n = 10), blood gas analysis was performed under anaesthesia. After sedation with medetomidin (Domitor, Orion Pharma, Finland, 40 µg/kg IM), and anaesthesia induction with propofol (Fresenius Kabi AB, Sweden, 1 mg/kg IV) eventually re-administered with the same dosage when required, bronchoscopy and BAL were applied to the BAL-diseased and BAL-control dogs accordingly to the previous described technique [2, 6, 17, 18]. Additional oxygen was not administered to the dogs during the procedure and sedation was reversed using atipamezole (Antisedan, Orion Pharma, Finland, 200 µg/kg IM). The anaesthesia reversion with atipamezole was rapid and complete within 5-10 minutes. Furthermore, 2 femoral arterial blood samples, one before sedation, the other, 5 minutes after BAL application but before anti-sedation, were obtained from each animal from BAL-diseased and BAL-control groups in accordance with the described technique [18]. After applying the above-mentioned anaesthesia protocol to the Anaesthesia-control group, laryngeal inspection was performed and dogs were intubated. Arterial blood samples were collected as previously described before anaesthesia and 10 minutes after intubation. BIOCHEMICAL, CYTOLOGICAL AND BACTERIOLOGICAL ANALYSES The BAL fluid samples were separated for biochemical, bacteriological and cytological analyses in accordance with the described technique [2, 6, 12]. The different biochemical markers were measured in the BAL fluids by spectrophotometry using commercial kits (Total proteins, urea, creatinine, Ca, P, LDH, ALP, ALT, and GGT, Spinreact kits, Spinreact SA., SPAIN). Cell populations present in BAL fluids were identified according to the following technique: after a first grossly evaluation and centrifugation (716g, 5 minutes at room temperature), they were left in rack about 1-2 minutes in order to let the precipitate to settle. The fluid supernatant was decanted completely by gently inverting the tube. Eight smears were made from each concentrated sample and for each smear 20 µL supernatant were used. After air-drying them at room temperature about half an hour, smears were stained simultaneously (2 with Gram’s, 2 with Diff-Quick, 2 with Revue Méd. Vét., 2010, 161, 5, 233-238 BRONCHOALVEOLAR LAVAGE AND BLOOD GASES IN DOGS Wright’s and 2 with May-Grünwald Giemsa stains). Each smear was inspected with light microscope and evaluated. Cell population were assessed by counting cell numbers for each types of cell groups in random 10 areas under 40X magnification and calculating their percentage reported to the total number of cells [2, 12-14]. The bacteriological examination of BAL fluids was performed according to the technique of PADRID and Mc KERNAN [15]. The immunoglobulin concentrations were determined using dog IgG, IgM, IgA ELISA quantification kits (Bethyl Lab. Inc., Montgomery, TX, USA) whose the detection limits were 7.8 - 500 μg/L for IgG and 15.6 - 1 000 μg/L for IgA and IgM. Each BAL fluid was diluted to 1:5, 1:10, 1:100 and 1:1000 for IgG and IgM measurements and to 1:5, 1:10 and 1: 100 for IgA determination in order to conform to the kit limits. All samples were analysed twice. Variations in blood gases values, acid-base balance and some electrolyte concentrations were analysed in accordance with the technique using the IRMA TruPoint® (ITC, USA) blood gases analyzer. The arterial-alveolar PO2 gradient (A-aPO2) was formulated according to sea level and room temperature conditions as follows [1, 22]: A-aPO2 (mmHg) = FiO2(BP - pH2O) - 1.25PaCO2 - PaO2 or A-aPO2 (mmHg) = 150 - 1.25PaCO2 - PaO2 where FiO2 was the fraction of inspired oxygen, BP the barometric pressure, pH2O the partial pressure of water at body temperature, PaCO2 the partial pressure of CO2 and PaO2 the partial pressure of O2. Parameters pH Beb (mmol/L) Beecf (mmol/L) HCO3- (mmol/L) TCO2 (mmol/L) PaCO2 (mmHg) PaO2 (mmHg) O2Sat.(%) tHb (g/L) Hct (%) A-aPO2 (mmHg) Na (mmol/L) K (mmol/L) iCa (mmol/L) Anaesthesia-control group Before During anaesthesia anaesthesia 7.43 ± 0.02a 7.37 ± 0.02b a -1.25 ± 1.15 -2.15 ± 1.17b 235 STATISTICAL ANALYSIS The statistical analysis was performed using independent samples t-test for comparison between groups and paired samples t-test for comparison within the groups. The differences were considered as significant when P value was less than 0.05. Results CLINICAL SIGNS Fever, loss of appetite, nasal discharge, coughing and abnormal pulmonary auscultation findings were observed in dogs with respiratory system disorders, whereas no signs of pulmonary or other disorders were detected in healthy dogs. No additional health disorder was detected in dogs throughout the study. Besides, mucosa hyperaemia and increased secretions were evidenced in sick dogs during endoscopy. BLOOD GAS ANALYSIS AND ACID-BASE EQUILIBRIUM As reported in Table I, the anaesthesia protocol in the 2 control groups (Anaesthesia- and BAL-control groups) has induced significant decreases of pH values (P < 0.05), partial pressure of oxygen (PaO2) and oxygen saturation (O2Sat) (P < 0.05 in anaesthesia-control group and P < 0.001 in BALcontrol group). By contrast, significant increases of partial BAL-control group Before During anaesthesia anaesthesia 7.44 ± 0.06a 7.40 ± 0.01b BAL-diseased group Before During anaesthesia anaesthesia 7.40 ± 0.01a 7.30 ± 0.01b a -1.08 ± 0.33 -3.14 ± 0.59b a -1.75 ± 0.34 -3.38 ± 0.60b -2.23 ± 1.13 21.83 ± 0.75 22.80 ± 0.75a 33.23 ± 1.27a 111.41 ± 4.23a 97.86 ± 0.27a 122.8 ± 10.9a 36.11 ± 3.20a -2.94 ± 0.50a -3.01 ± 1.24 21.93 ± 0.98 23.05 ± 0.98b 38.11 ± 1.27b 88.55 ± 7.10b 95.35 ± 1.07b 135.5 ± 7.8b 39.81 ± 2.28b 13.82 ± 1.30b -0.61 ± 0.57 -1.40 ± 0.60 22.43 ± 0.50a 23.12 ± 0.53a 33.20 ± 1.40a 111.60 ± 3.10a,A 97.94 ± 0.32a,A 133.8 ± 6.9a 39.30 ± 2.00a -3.10 ± 0.70a,A -0.65 ± 1.03 -0.92 ± 1.03 23.79 ± 0.74b 24.95 ± 0.74b 38.89 ± 1.30b 84.79 ± 3.83b 93.60 ± 2.79b 150.4 ± 5.7b 44.28 ± 1.68b 26.60 ± 1.16b 22.51 ± 0.29 23.57 ± 0.30 35.23 ± 0.79a 72.83 ± 1.79a,B 95.52 ± 0.18a,B 130.8 ± 4.0a 38.49 ± 1.18a 24.00 ± 2.10a,B 22.13 ± 0.43 23.34 ± 0.42 41.55 ± 0.75b 63.29 ± 2.21b 90.30 ± 1.61b 142.4 ± 3.3b 36.04 ± 0.97v 34.78 ± 3.20b 146.48 ± 0.89 4.21 ± 0.15 1.41 ± 0.07a 146.03 ± 0.64 4.44 ± 0.17 1.52 ± 0.03b 145.80 ± 2.00 4.17 ± 0.17 1.44 ± 0.50a 145.85 ± 1.79 3.96 ± 0.13 1.53 ± 0.05b 147.07 ± 1.17 4.29 ± 0.10 1.34 ± 0.02 146.22 ± 1.03 4.42 ± 0.78 1.38 ± 0.03 Beb: Base excess of blood; Beecf: Base excess of extra cellular fluid; TCO2: Total carbon dioxide; PaCO2: Partial pressure of carbon dioxide; PaO2: Partial pressure of oxygen; O2Sat.: Oxygen saturation; tHb: total hemoglobin; Hct: Haematocrit; A-aPO2: Arterial-alveolar PO2 gradient; iCa: ionized calcium. Different superscripts a,b in the same line indicate significant differences (P < 0.05) within the same group (effects of anaesthesia or BAL collection or Disease) and different superscripts A,B in the same line indicate significant differences (P < 0.05) between the BAL-control and BAL-disease groups. TABLE I : Arterial blood gas and acid-base equilibrium in Anaesthesia-control (n = 10), BAL-control (n = 10) and BAL-diseased (n = 30) dog groups. Anaesthesia was performed with association of medetomidin plus propofol. Results are expressed as mean ± standard error. Revue Méd. Vét., 2010, 161, 5, 233-238 236 pressure of carbon dioxide (PaCO2), Total carbon dioxide (TCO2), haematocrit (Hct), Total hemoglobin (tHb), ionized calcium (iCa) (P < 0.05) and arterial-alveolar PO2 gradient (A-aPO2) (P < 0.01) were observed during anaesthesia in both 2 groups. When the responses of these groups to anaesthesia were compared, the increase of A-aPO2 and the decrease of the PaO2 appeared more marked in the BALcontrol group (A-aPO2 variations: + 29.70 mmHg and PaO2 variations: - 26.81 mmHg) than in the Anaesthesia-control group (+ 16.76 mmHg and - 22.86 mmHg, respectively). Compared to the BAL-controls, the PaO2 and O2Sat values before anaesthesia were significantly depressed (P < 0.001) and PaCO2 and arterial-alveolar PO2 gradient (A-aPO2) values significantly enhanced (P < 0.05) in BAL-diseased dogs. Furthermore, the anaesthesia-induced variations of pH, O2Sat, PaCO2 and Beb (Base excess in blood) were amplified in the diseased animals whereas changes in all other parameters (except of Na and K concentrations which remained roughly constant during anaesthesia in all 3 groups) were on the contrary alleviated (PaO2, Hb, A-aPO2,) or abolished (TCO2, HCO3-, Beecf, Hct, iCa). GONUL (R.) AND COLLABORATORS BAL-control group BAL-diseased group Total Protein (g/L) 1.1 ± 0.4 1.3 ± 0.2 Urea (mmol/L) 1.10 ± 0.03a 1.30 ± 0.03b Creatinine (μmol/L) 7.4 ± 0.5 8.5 ± 0.5 Ca (mmol/L) 1.10 ± 0.04 1.10 ± 0.03 P (mmol/L) 0.100 ± 0.003 7.4 ± 0.6a 0.200 ± 0.010 12.7 ± 0.4b LDH (U/L) 24.5 ± 1.0a 14.0 ± 0.3a 55.1 ± 2.8b 22.7 ± 0.6b GGT (U/L) 7.8 ± 0.2 7.2 ± 0.1 IgG (mg/L) 20 ± 10 100 ± 100 IgM (mg/L) 2±1 7±3 IgA (mg/L) 20 ± 6 8±4 Neutrophil (%) 6.5 ± 5.7a 55.5 ± 13.3b Eosinophil (%) 2±2 5.4 ± 5.2 Macrophages (%) 74.07 ± 6.57a 24.3 ± 16.9b Biochemistry ALT (U/L) ALP (U/L) Cytology BIOCHEMICAL AND CYTOLOGICAL ANALYSIS OF THE Lymphocyte (%) 14.35 ± 4.12 6.34 ± 10.38 BAL FLUID Epithelial cells (%) 1.24 ± 0.08a 4.1 ± 0.4b The Table II presents the biochemical and cytological findings of the BAL fluids collected from dogs with lower respiratory tract disorders and from healthy dogs. Whereas the total protein and calcium concentrations and the GGT activity in the BAL fluids were similar between the 2 groups, the other investigated parameters tended to increase (creatinine and phosphorus concentrations) or significantly increased (urea concentrations and ALT activity (P < 0.01), ALP and LDH activities (P < 0.001)) in diseased dogs. As far as immunoglobulins were concerned, the IgG and IgM concentrations appeared to be augmented in dogs with respiratory diseases compared to controls but not significantly because of the great dispersion of values. The cytological analysis of the BAL fluids revealed significant and marked increases of the counts of inflammatory neutrophil leukocytes and macrophages as well as epithelial cells compared to healthy dogs. Furthermore, the proportion of bacteriological positive BAL fluids was 43.33% and cocci and/or rods (mainly E. coli, Staphylococcus epidermidis, Klebsiella pneumoniae, Mycoplasma spp. and Enterobacter cloaca) were also isolated from BAL fluids from diseased dogs while no micro-organism was found in healthy controls (Table III). Discussion In the present study which entailed a detailed endoscopic examination of dogs with lower respiratory system disorders, changes in blood gases analysis, acid-base balance and in some electrolyte concentrations were detected before and after endoscopy. In addition, extensive biochemical, cytological, bacteriological and immunological evaluation of collected ALT: Alanine aminotransferase; ALP: Alkaline phosphatase; LDH: Lactate deshydrogenase; GGT: Gamma-glutamyl Transferase. Different superscripts a,b in the same line indicate significant differences (P < 0.01). TABLE II: Biochemical and cytological parameters of the BAL fluid collected under anaesthesia (medetomidin plus propofol) from healthy dogs (BAL-control group, n = 10) or from dogs with lower respiratory airway diseases (BAL-diseased group, n = 30). Results are expressed as mean ± standard error. BAL-control BAL-diseased group group Germs E. coli 0 4 Staphylococcus epidermidis 0 2 Klebsiella pneumoniae 0 2 Mycoplasma spp 0 2 Enterobacter cloaca 0 2 0 1 0/10 13/30 Others Total TABLE III: Bacteriological analysis of the BAL fluid collected under anaesthesia (medetomidin plus propofol) from healthy dogs (BALcontrol group, n = 10) or from dogs with lower respiratory airway diseases (BAL-diseased group, n = 30). bronchoalveolar lavage fluid was performed from patients with respiratory tract disorders and the effects on blood gases of anaesthesia induced by the combination medetomidin plus propofol were examined in healthy and diseased dogs. Revue Méd. Vét., 2010, 161, 5, 233-238 BRONCHOALVEOLAR LAVAGE AND BLOOD GASES IN DOGS Dogs with lower respiratory tract disorders exhibited symptoms such as fever, anorexia, depression, nasal discharge, coughing, expectoration, tracheal sensitivity and abnormal pulmonary auscultation findings, similar to those reported by other researchers [11, 12]. BAL applications, similar to those reported by other researchers [13, 17] did not cause any additional difficulties in healthy dogs or in those with respiratory tract disorders. In agreement with previous reports [9, 12], epithelial and inflammatory neutrophils and macrophages in the current study were increased and various bacterial agents such as E. coli, Staphylococcus epidermidis, Klebsiella pneumoniae, Mycoplasma spp. and Enterobacter cloaca have grown in the BAL fluids in dogs with respiratory tract disorders, corroborating the inflammatory and /or the infectious nature of some respiratory diseases. Furthermore, it was reported that cellular enzymes in BAL fluid can be used as important markers of cellular integrity or cellular damage, and total protein and urea concentrations can be used to detect alterations in vascular permeability and permeability of the membranes of the respiratory system [9, 12]. Accordingly, urea concentration and ALT, LDH and ALP activities in the BAL fluid of patients with respiratory tract disorders were found to be statistically significantly increased. PADRID et al. [14] reported low concentrations of IgG (<0.01 mg/ml) in BAL fluid from healthy cats and that they failed to detect IgM while detecting IgA. In the present study, the immunoglobulin concentrations in BAL fluid of healthy dogs were also very low (20 ± 10 mg/L, 2 ± 1 mg/ml and 20 ± 6 mg/ml, for IgG, IgM and IgA respectively). Although not significantly because of great value dispersion, increases of IgG and IgM concentrations were observed in sick animals in the present study, suggesting probably local defence reactions. A PaO2 lower than 80 mmHg, indicates hypoxemia [11, 16, 19]. The difference between alveolar and arterial O2 pressures, called alveolar-arterial gradient (A-aPO2), indicates tissue oxygenation and is usually below 15 mmHg [1, 16, 19]. In this study, dogs with pulmonary disorders exhibited significantly decreased PaO2 and O2Sat and a dramatically increased A-aPO2 compared to healthy dogs, indicating hypoxemia and pulmonary oxygenation deficit, respectively. Moreover, after endoscopy and BAL application, these parameters were deeply depressed and the base deficit increased in the diseased group. These alterations reflect ventilation problems induced by various diseases but also by the BAL procedure itself which includes sedation and decumbency [16-18]. In this way, it was previously observed decrease of PaO2 and increase of PaCO2 induced by medetomidin and propofol anaesthesia [4, 5]. In agreement with that, the combination of medetomidin and propofol used in the present study clearly induced strong diminutions of PaO2, O2Sat and the base deficit coupled to augmentations of PaCO2, TCO2 and A-aPO2. In parallel, the haematocrit values and the haemoglobin concentrations were significantly enhanced in anaesthesia-controls, probably for compensating the oxygenation deficit. Although RAJAMAKI et al. [17] also reported that these changes could be more pronounced in dogs with pulmonary tract diseases in a first study, the same researchers found that variations of PaO2 and A-aPO2 were similar in dogs with pulmonary eosinophilia and in healthy dogs [18]. In this study, variations of O2Sat, PaCO2, pH and base deficit were exacerbated after BAL application in dogs with lower Revue Méd. Vét., 2010, 161, 5, 233-238 237 airway respiratory diseases whereas the intensity of tissue oxygenation evidencing by PaO2 and arterial-alveolar PO2 gradient (A-aPO2), already depressed, and the red blood cells mobilisation reflecting by haematocrit and haemoglobinemia were less affected by endoscopy and BAL collection in the diseased animals than in the healthy controls. Consequently, it would be out of interest to pay more attention and to investigate the oxygen saturation in respiratory disease dogs. As a conclusion, the biochemical, cytological and bacteriological analyses of BAL fluids can be useful for diagnosis and prognostic of the lower airway respiratory disorders in dogs: marked increases of LDH, ALT and ALP activities coupled to increased cellularity (inflammatory and epithelial cells) confirms pulmonary inflammation or damage, variations of protein and urea concentrations indicate alterations in vascular and respiratory permeability, increases of immunoglobulin concentrations evidences local and specific defence reactions and identification of some microorganisms can lead to an accurate diagnosis. Additionally, the detection of blood gases values and namely of PaO2, O2Sat, PaCO2 and arterialalveolar PO2 gradient (A-aPO2), helps in the follow-up of the disease. Nevertheless, despite the comfort of medetomidin and propofol anaesthesia easily reversed with antipamezole, as the anaesthesia protocol and the BAL procedure directly alter gas exchange and the acid-base equilibrium, the blood gas analysis would be helpful in the surveillance of dogs with lower airway pulmonary disorders in order to limit disease aggravation. Acknowledgement The authors wish to acknowledge Research Fund of Istanbul University for supporting this research (Project number: 596/15122006). References 1. - ARGYLE B.: Blood gases. Blood Gases Computer Program Manual. Mad Scientist Software, Alpine UT, 1996. http://www.madsci.com./manu/ gas_aa.htm. Accessed June 22, 2009. 2. - COLLIE D.D., DEBOER D.J., MUGGENBURG B.A., BICE D.E.: Evaluation of association of blood and bronchoalveolar eosinophil numbers and serum total immunoglobulin E concentration with the expression of nonspecific airway reactivity in dogs. Am. J. Vet. 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