Bronchoalveolar lavage cytology and hematology: a comparison between high and low health status pigs at three different ages.

Bronchoalveolar lavages (BALs) were performed with a bronchoscope on 5- and 7.5-week-old, anesthetized, high health status pigs (n = 14). At 10 weeks of age, pigs (n = 28) were necropsied, lungs were removed, and BAL samples were collected from the right diaphragmatic lobe with a modified 12-Fr (4-mm) Foley catheter. Peripheral blood was sampled from all pigs (n = 28) before each BAL procedure. Peripheral blood and BAL samples were collected according to a similar study design at 5, 7.5, and 10 weeks of age from 12 low health status pigs, which were raised according to standard farm procedures (n = 6) or as segregated early weaned pigs (n = 6). Bronchoalveolar lavage cytology and hematologic 95% confidence intervals were determined for 5-, 7.5-, and 10-week-old high (group A) and low health status pigs (groups B and C). The results were compared between the different groups. Repeated BALs were easily performed in all pigs, making this an additional tool for evaluation of respiratory health. Total numbers of cells and neutrophils in peripheral blood and BAL samples were greater in low health status pigs than in high health status pigs. Hematologic results paralleled the findings in BAL fluid. Segregated early weaning of low health status pigs in a less challenging environment mainly reduced the number of neutrophils in BAL samples and peripheral blood.     

Immunophenotyping of chicken peripheral blood lymphocyte subpopulations: individual variability and repeatability

T-cell lymphocyte populations can be delineated into subsets based on expression of cell surface proteins that can be measured in peripheral blood by monoclonal antibodies and flow cytometry percentages of the lymphocyte subpopulations. In order to accurately assess immunocompetence in birds, natural variability in both avian immune function and the methodology must be understood. Our objectives were to (1) further develop flow cytometry for estimating subpopulations of lymphocytes in peripheral blood from poultry, (2) estimate repeatability and variability in the methodology with respect to poultry in a free-range and environmentally diverse situation, and (3) estimate the best antibody and cell marker combination for estimating lymphocyte subpopulations. This work demonstrated the repeatability of using flow cytometry for measurements of peripheral blood in chickens using anti-chicken antibodies for lymphocyte subpopulations. Immunofluorescence staining of cells isolated from peripheral blood revealed that the CD3(+) antibodies reacted with an average of approximately 12-24% of the lymphoid cells in the blood, depending on the fluorescence type. The CD4(+) and CD8(+) molecules were expressed in a range of 4-31% and 1-10% of the lymphoid cells in the blood, respectively. Both fluorescence label and antibody company contribute to the variability of results and should be considered in future flow cytometry studies in poultry.     

Results of cytologic and microbiologic analysis of bronchoalveolar lavage fluid in New Zealand White rabbits

OBJECTIVE: To determine cytologic and microbiologic findings in bronchoalveolar lavage (BAL) fluid and SpO(2) values obtained during BAL in healthy rabbits. ANIMALS: 9 rabbits. PROCEDURES: Bronchoscopic BAL of left and right caudal lobar bronchi (LB2 and RB4) was performed with 3 mL of sterile saline (0.9% NaCl) solution; SpO(2) was measured before, during, and after BAL. Percentage fluid recovered, total leukocyte counts, and differential cell counts were determined. Aerobic and anaerobic bacterial, mycoplasmal, and fungal cultures were performed from combined LB2 and RB4 samples. RESULTS: Mean +/- SD percentage fluid volumes recovered from LB2 and RB4 were 53 +/- 13% and 63 +/- 13%, respectively. Mean +/- SD total leukocyte counts from LB2 and RB4 were 422 +/- 199 cells/microL and 378 +/- 97 cells/microL, respectively. Macrophages were most frequently identified. There were no significant differences in volumes retrieved, total leukocyte counts, or differential cell percentages between LB2 and RB4. Microbial culture results were negative for 3 rabbits and positive for mixed aerobic and anaerobic bacterial growth in 6 and 2 rabbits, respectively. The SpO(2) was > or = 95% in 7 of 9 rabbits after anesthetic induction, < 95% in 5 of 6 rabbits 1 minute after BAL, and > or = 95% in 5 of 9 rabbits and > 90% in 4 of 9 rabbits 3 minutes after BAL. CONCLUSIONS AND CLINICAL RELEVANCE: Bronchoscopic BAL with 3 mL of saline solution provided adequate fluid recovery for microbiologic and cytologic examination from the caudal lung lobes. Transient low SpO(2) was detected immediately after BAL.     

Effects of inhalation of dust and endotoxin on respiratory tracts of pigs.

OBJECTIVE: To assess the effects of inhalation of feed flour dust and dustborne endotoxin on respiratory tracts of pigs. ANIMALS: 29 healthy Belgian Landrace pigs. PROCEDURE: Pigs housed in an environmental chamber were exposed for 6 days to feed flour dust (1 to 15 mg/m3) and dustborne endotoxins (50 to 2,500 ng/m3). Effects were evaluated by measuring albumin concentration, lactate dehydrogenase (LDH) activity, cell composition of nasal lavage (NL) and bronchoalveolar lavage (BAL) fluids and blood, and percentages of CD4+ and CD8+ T lymphocytes in blood and lavage fluids. Dustborne endotoxin was obtained by mixing endotoxins from Escherichia coli (serotype O127:B8) with feed flour before spraying the flour in the environmental chamber. RESULTS: Exposure did not affect cell composition of NL fluid or blood. Total cell counts of BAL fluids were increased in all groups exposed to dust. Macrophage counts were increased in pigs exposed to inhalable dust concentrations as low as 4.4 mg/m3, and lymphocyte counts were increased in groups exposed to high dust concentrations. Percentages of CD4+ and CD8+ T lymphocytes in blood and lavage fluids were unchanged. In all dust-exposed groups, albumin content of BAL fluid was increased, whereas LDH activity was unaffected. Macrophage and lymphocyte infiltration and edema in the bronchi were identified by light microscopy. Effects attributable to E. coli endotoxin exposure were not identified. CONCLUSIONS: Inhalation of feed flour dust did not affect nasal mucosa but did induce bronchial airway inflammation. Dustborne endotoxins did not have effects attributable to endotoxin alone.     

Flow cytometric analysis of canine umbilical cord blood lymphocytes

Lymphocyte subsets in canine umbilical cord blood were flow cytometrically analyzed and compared with those of the dams' peripheral blood. The proportion of CD3+ T lymphocytes, CD21+CD3- B lymphocytes, and CD3-CD21- non-T non-B lymphocytes in umbilical cord blood was 52.9%, 30.4%, and 16.7%, respectively. T lymphocyte/B lymphocyte ratio was significantly lower in the umbilical cord blood than in the dams' peripheral blood (2.1 +/- 1.4 versus 11.0 +/- 8.1, P < 0.001). In contrast, CD4+ lymphocyte/CD8+ lymphocyte ratio was significantly higher in the umbilical cord blood than in the dams' peripheral blood (7.6 +/- 2.2 versus 1.8 +/- 0.6, P<0.001). These findings clarified the phenotypic characters of canine umbilical cord blood lymphocytes.     

Bronchoalveolar lavage cytology in cynomolgus monkeys and identification of cytologic alterations following sequential saline lavage

Total and differential cell counts were determined on cytolytic specimens obtained by fiberoptic bronchoscopy and bronchoalveolar lavage (BAL) of five normal cynomolgus monkeys. Total nucleated cell counts ranged from 100 to 430 cells/microliters. Macrophages were approximately 91% of total nucleated cells, while lymphocytes were 3%, neutrophils 4%, and eosinophils 2% of the initial BAL from each monkey. Less than 1% of the cells were mast cells and ciliated or nonciliated epithelial cells. The effects of repeated saline BAL on pulmonary cell populations were evaluated. Saline lavage of individual lung lobes resulted in a marked rise in circulating blood neutrophils at 4 hr after BAL; there was a similar rise in neutrophils in lavage fluids 24 hr after the initial lavage. Differential and total cell counts of both blood and lavage fluid returned to normal if subsequent lavages were spaced at 48-hr intervals. Lymphocytes were not present in saline-lavaged lung lobes, and protein levels of lavage fluids did not rise significantly. BAL produced a transient, reversible, intra-alveolar influx of neutrophils which was preceded by mobilization of bone marrow-stored neutrophils. Neutrophilia in the lavage fluid and blood was not detectable if lavage and blood sampling procedures were done at 48-hr intervals (which did not alter Ia antigen expression among BAL cells). These observations indicate that BAL is a valid method for sampling and assessing pulmonary cellular and fluid constituents if the procedures are done at intervals of at least 48 hr.     

Comparison of analysis of bovine surface immunoglobulin bearing and peanut agglutinin binding lymphocytes by flow cytometry and fluorescence microscopy

Bovine peripheral blood lymphocytes were examined for their binding to anti-immunoglobulin serum, peanut agglutinin, and mu, alpha, and epsilon heavy chain specific antisera by immunofluorescence. The percentage of total lymphocytes with positive staining was determined independently by flow cytometry and fluorescence microscopy. The correlation of data from both methods was best for analysis of total surface immunoglobulin and IgM bearing cells. The percentage of lymphocytes bearing surface immunoglobulin (B cells) was determined using both whole antiserum and a F(ab')2 reagent. Quantitation by flow cytometry did not show a significant difference when the two reagents were used, whereas fluorescence microscopy revealed a significant difference (p less than .05). The mean percent of total surface immunoglobulin bearing cells was 30 +/- 3% by either method. Flow cytometry gave significantly larger values than fluorescence microscopy for samples stained with fluorescein conjugated peanut agglutinin. Peanut agglutinin binding cells comprised 70 +/- 3% by flow cytometry and 51 +/- 3% by fluorescence microscopy. Similarly, there was a significant difference between both methods when IgA bearing lymphocytes were examined. Percentages of immunoglobulin E, A, and M bearing lymphocytes as well as total B and T cells in spleen and bronchial lymph node were determined by immunofluorescence using the cytofluorograph. Peanut agglutinin binding cells were less numerous in spleen and lymph node than in peripheral blood. Immunoglobulin E bearing lymphocytes increased from 0.07% in peripheral blood to 4% in spleen and 1.9% in lymph node. In this paper we demonstrate how flow cytometry can be used to examine a large number of samples in a rapid and reproducible manner. This is the first report in which bovine lymphocytes bearing surface IgE are quantitated.     

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.     

Evaluation of elutriation and magnetic microbead purification of canine monocytes

An elutriation technique was developed to obtain large quantities of pure canine monocytes. Firstly, peripheral blood mononuclear cells (PBMC) were isolated from whole blood by Ficoll gradient. Then, the PBMC were separated by an elutriation procedure. We demonstrated that these techniques allow the isolation of canine peripheral blood monocytes with a purity of 64% +/- 7.9 when labelled with anti-CD14 antibody. This purity increased to 83% +/- 2.2 after separation by magnetic anti-CD14 microbeads. The cell viability was more than 95% and apoptotic cells were less than 10%. The monocytes purified by these methods were functionally active in a mixed leukocyte reaction (MLR). A lymphocyte fraction was obtained directly only by elutriation with an average of 79.9% +/- 10.7 of CD5+, 7.9% +/- 3.5 of CD21+ and 1.78% +/- 2.53 of CD14+. Our results indicate that this elutriation procedure is a safe method to purify monocytes as well as lymphocytes, useful in MLR.     

Bronchoalveolar lavage cytology from captive badgers

Background: Bronchoalveolar lavage (BAL) fluid is evaluated for the diagnosis and study of lung disease and airway inflammation. Cytologic profiles for BAL fluid have not been reported for badgers and may be useful in understanding the pathogenesis of pulmonary diseases such as Mycobacterium bovis. Objective: The aim of this study was to evaluate cytologic and microbial findings in BAL fluid from captive European badgers (Meles meles) and identify correlates with the results of concurrently collected blood and fecal samples. Methods: BAL fluid (by a nonbronchoscopic method) and jugular venous blood samples (for routine CBC) were obtained from 23 captive tuberculosis‐free anesthetized badgers on 2 occasions 4 weeks apart. Fecal samples were collected for routine parasitology. Morphologic evaluation and 100‐cell differentials were done on cytocentrifuged BAL specimens. Pellets from centrifuged BAL were aerobically cultured for bacteria. Results: With the 2 BAL samples from each of the 23 badgers combined, the median (range) cell percentages were 73.0% (5–95%) neutrophils, 7.5% (2–16%) macrophages, 8.0% (0–27%) lymphocytes, and 9.5% (0–92%) eosinophils. Macrophages frequently contained silica‐like crystals. Other findings included ciliated epithelial cells, goblet cells, mucus, and Aelurostrongylus sp. larvae. A light growth of Streptococcus, Pasteurella, or Escherichia coli was cultured in 6 badgers. Trypanosoma pestanai were identified in blood from 10 badgers and fecal parasites (mainly coccidia) were found in 20 badgers. No correlation was found between BAL and CBC results and the presence of parasites. Conclusions: The predominance of neutrophils in BAL fluid from badgers differs from the predominance of macrophages found in BAL from other species. This difference may reflect the burrowing lifestyle or the unique immune response of badgers.     

Distribution of T and B lymphocytes in mammary dry secretions, colostrum and blood of adult dairy cattle

The distribution of lymphocyte subpopulations from dry secretions, colostrum and blood from 10 healthy adult Hostein-Fresian cows was studied using the TH21A and B26A mouse monoclonal antibodies (MAb) to adult bovine B and T lymphocytes, respectively. The mammary gland lymphocytes (MGL) were isolated from composite sample of all four quarters by density centrifugation over discontinuous gradient of ficoll-diatrizoate. The peripheral blood lymphocytes (PBL) were purified using the ficoll-thrombin method. Isolated PBL and MGL were analyzed using the two fluorochromes method (TFM) and laser flow cytometry (LFC). The mean viability of isolated PBL and MGL from dry secretions and colostrum after the TFM and LFC were 92.4% +/- 3.2%, 91.4% +/- 6.0% and 87.1% +/- 6.1%, respectively. There was a good correlation between the two MAbs and the percentage of surface immunoglobulin (SIg) positive cells in the peripheral blood using the TFM. The PBL yielded a mean percentage of 21.2% B cells, 66.4% T cells and 9.4% "Null cells" (TH21A+; SIg-). The TFM on MGL from dry secretions and colostrum indicated two distinct patterns (group I and II) of SIg and reactivity to MAb markers (p less than 0.001). The MGL data included in group I and group II were gathered from both colostral and dry secretions. In comparison to the distribution of lymphocyte subsets within peripheral blood the mean percentages of B cells, T cells and "Null cells" in the mammary gland were respectively, 2.8%, 88.1% and 5.4% for group I and 3.5%, 89.0% and 15.1% for group II. In the mammary secretions, the use of SIg alone was not considered to be a good marker for B cells; in four animals a mean percentage of 15.6% (13.9/89.0 X 100) of the mammary gland T lymphocytes were also SIg+. Of the TH21A+ MGL, only 18.8% were SIg+ in group II compared with 34.1% for MGL from group I and 69.3% for the PBL. Marked differences in cell size distribution and cell surface antigen density were found when PBL and MGL from dry secretions were compared by LFC using the B26A MAb. The results of this study demonstrate a difference in the percentages of peripheral blood and mammary gland B and T lymphocytes and confirm previous findings in which the T lymphocytes were found to represent the major subpopulation of lymphocytes in bovine mammary secretions. This may represent an essential event in the adoptive transfer of cellular immunity through the colostrum in cattle.     

Alveolar clearance in horses with chronic obstructive pulmonary disease

OBJECTIVE: To assess sensitivity of scintigraphic alveolar clearance rate as an indicator of alveolar epithelium damage in horses. ANIMALS: 5 healthy horses (group A) and 5 with chronic obstructive pulmonary disease (COPD; group B). PROCEDURE: Horses underwent clearance rate (k [%/min]) determination. Clearance rate of group-B horses was determined after remission of the disease following 2 months at pasture (remission 1), stabling in a controlled environment (remission 2), and during crisis induced by exposure to moldy hay and straw. Methacholine challenge test was performed at each investigation period to determine nonspecific pulmonary airway hyperresponsiveness. Pulmonary function tests (PFT) also were performed, and cell populations in bronchoalveolar lavage (BAL) fluid were determined on another occasion. RESULTS: Group-B horses had significantly faster mean clearance rate during crisis (k = 4.30+/-0.95%/min), compared with that for remission 1(k = 1.98+/-0.55%/min), which did not differ from the rate in group-A horses (k = 1.95+/-0.33%/min). Despite lack of clinical signs of COPD during remission when stabled in a controlled environment, an intermediate value was found (k = 3.20+/-0.72%/min). CONCLUSIONS: This technique allowed grading of lung damage induced by COPD, whereas use of PFT and determination of BAL fluid cell populations failed to differentiate between remission 1 and remission 2. CLINICAL RELEVANCE: Determination of alveolar clearance rate by use of scintigraphy is a sensitive indicator of lung damage. A modified clearance rate was found despite the lack of clinical and functional changes.     

Lymphocyte subset distribution in apparently normal and single intradermal test-positive water buffaloes analyzed by flow cytometry

Monoclonal antibodies (mAbs) against bovine lymphocyte cell surface antigens namely, MHC Class I, MHC class II (DP, DQ and DR), CD3, CD4, CD8, gamma delta TCR, WC1N1 and WC1N2, were tested for their reactivity on apparently normal buffalo mononuclear cells prepared from spleen, lymph nodes and peripheral blood. All the mAbs cross-reacted with the buffalo mononuclear cells. The mean (+/-SD) CD4:CD8 cell ratio in the peripheral blood of apparently normal buffaloes was 1.08+/-0.049 while in the spleen and lymph nodes it was 0.90+/-0.080 and 1.81+/-0.430, respectively. The lymphocyte subsets in the buffaloes positive for tuberculosis by the single intra dermal (SID) test was found to be altered; the CD4 cells were reduced while the CD8 and gamma delta cells were increased. The mean CD4:CD8 ratio in the SID positive buffaloes was 0.36+/-0.010.     

Cytokine profiles of peripheral blood and airway CD4 and CD8 T lymphocytes in horses with recurrent airway obstruction

Equine recurrent airway obstruction (RAO) is thought to result from an aberrant immune response to inhaled antigens, modulated by T lymphocytes via the secretion of pro-inflammatory cytokines. However data relating to the phenotypes of the T lymphocytes present in peripheral blood and bronchoalveolar lavage fluid of RAO horses and their cytokine profiles are contradictory. The aim of this study was to further investigate the cytokine (IL-4, IL-5, IL-13 and INF-gamma) mRNA expression profile in peripheral blood lymphocytes and bronchoalveolar lavage lymphocytes from RAO and control horses, before and at 48 h after horses were exposed to hay/straw. In contrast to previous studies, cytokine expression was quantified in populations of CD4 and CD8 T lymphocytes which were purified using magnetic bead antibody cell separation. Hay/straw exposure induced clinical airway obstruction, airway neutrophilia and airway lymphocytosis in RAO horses, and, induced a mild, but significant, airway neutrophilia in controls. However, hay/straw exposure had no significant effect on peripheral blood lymphocyte or bronchoalveolar lavage lymphocyte cytokine expression in either group. In conclusion, RAO was not associated with alterations in lymphocyte cytokine expression that are consistent with Th1 or Th2 responses, but rather with a general down-regulation in expression of the measured cytokines in peripheral blood lymphocytes and bronchoalveolar lavage lymphocytes.     

Characterization of feline T and B cells

Feline peripheral-blood lymphocyte populations (n = 22) were examined for the following markers: rosette formation with guinea pig erythrocytes (GPE-T cells), rosette formation with human RBC (HRBC-T cells), rosette formation with sheep RBC, mixed rosette formation with GPE-T cells and HRBC-T cells (total T cells), erythrocyte antibody-complement rosettes, and surface immunoglobulin. An average of 28% +/- 7% (range, 16% to 39%) of the feline lymphocytes formed rosettes with GPE-T cells, and 27% +/- 7% (range, 11% to 36%), with HRBC-T cells. An average of 57% +/- 9% (range, 33% to 75%) of the lymphocytes formed mixed rosettes. The erythrocyte antibody-complement rosette-forming cells and surface immunoglobulin-bearing cells were found in peripheral blood lymphocytes (10% +/- 6% and 24% +/- 8%, respectively). The murine monoclonal antibodies OKT 11 and HuLy-m1, specific for a framework determinant of human E-rosette receptor antigens, cross-reacted with feline cell membrane molecules recognizing a bimolecular complex (45,000 to 50,000 daltons) similar to that described in persons. We investigated the distribution of these E-rosette receptor-like antigens on feline lymphocytes. By complement-mediated lymphocytotoxicity, about 30% of the feline lymphocytes expressed the antigens. When lymphocytes were treated with HuLy-m1 antibody, spontaneous rosette formation with HRBC-T cells was significantly inhibited.     

Effects of an external nasal strip and frusemide on pulmonary haemorrhage in Thoroughbreds following high-intensity exercise.

The purpose of this study was to examine the effects of an external nasal strip (NS), frusemide (FR) and a combination of the 2 treatments (NS + FR) on exercise-induced pulmonary haemorrhage (EIPH) in Thoroughbred horses. It was hypothesised that both the NS and FR would attenuate EIPH as assessed by red blood cell count in bronchoalveolar lavage fluid. In random order, 8 horses completed each of 4 sprint exercise tests on a treadmill: 1) NS; 2) FR (0.5 mg/kg bwt i.v., 4 h pre-exercise); 3) NS + FR; and 4) control (C; no treatment). After a 5 min warm-up (4.5 m/s), horses completed 2 min running at 120% maximum oxygen consumption (VO2max) with the treadmill set at 3 degrees incline. Mean +/- s.d. running speed was 14.2+/-0.2 m/s. In the FR and NS + FR trials, horses carried weight equal to that lost as a result of frusemide administration. During exercise at 120% Vo2max, oxygen consumption (Vo2) and carbon dioxide production (Vco2) were measured at 15 s intervals. Plasma lactate concentration was measured in samples collected before exercise, at the end of the sprint and after 5 min cool-down at the trot. Thirty minutes after the run, bronchoalveolar lavage (BAL) was performed and the red cell count in the fluid quantified. Vo2 and Vco2 were significantly lower in NS and NS + FR trials than in the C and FR trials at the end of the sprint exercise protocol. However, plasma lactate concentrations did not differ among treatments. Compared with the C trial (61.1+/-30.5 x 10(6) red blood cells/ml BAL fluid), pulmonary haemorrhage was significantly (P<0.05) decreased in both the NS (15.9+/-4.0 x 106 RBC/ml) and FR (12.2+/-5.8 x 10(6) RBC/ml) trials. EIPH in the NS + FR trial (7.9+/-1.0 x 10(6) RBC/ml) was further diminished (P<0.05) compared to the NS trial, but not different from the FR trial. We conclude that both the external nasal strip and frusemide attenuate pulmonary haemorrhage in Thoroughbred horses during high-speed sprint exercise. The external nasal strip appears to lower the metabolic cost of supramaximal exertion in horses. Given the purported ergogenic effects of frusemide, the external nasal strip is a valuable alternative for the attenuation of EIPH.     

Effects of lung site and fluid volume on results of bronchoalveolar lavage fluid analysis in horses.

Bronchoalveolar lavage (BAL) fluid was analyzed in healthy horses, using different lavage fluid volumes and lung sites. The only significant difference in the cellular composition of BAL fluid between the right and left lungs was the mast cell numbers, which were significantly higher in the left lung. Total cell count ranged from 34 to 330 cells/microliter for the right lung and 43 to 330 cells/microliter for the left lung. Percentage of neutrophils ranged from 1 to 7% in the right lung and 1 to 5% in the left lung. The small-volume (50 ml) lavage had a greater percentage of neutrophils and a lesser percentage of mast cells in the large-volume (350 ml) lavage. Statistical difference in the composition of BAL fluid recovered was not detected between the 3 sequential 100-ml lavages and a single 300-ml lavage, except that macrophages were significantly higher in the 3 sequential 100-ml lavages. Values for BAL fluid analysis in healthy horses have varied considerably and this variation is from a failure to adhere to any standard technique for volume of fluid infused.     

Changes in subpopulations of lymphocytes in peripheral blood, and supramammary and prescapular lymph nodes of cows with mastitis and normal cows

Direct immunofluorescence (IF) and indirect IF techniques were employed to analyze the distribution of B and T lymphocyte populations in peripheral blood, and in supramammary (draining), and prescapular (non-draining) lymph nodes of cows with mastitis and normal cows. In the peripheral blood there was a significant decrease in the percent and absolute number of B lymphocytes in mastitic cows (n = 29; 17.1 +/- 10.2%; 3.4 +/- 2.7 X 10(5) cells/ml) as compared to normal cows (n = 38; 25.2 +/- 7.8%; 9.3 +/- 5.4 X 10(5) cells/ml). The percent T lymphocyte count in mastitic cows (71.2 +/- 7.1%) was slightly increased over that of normals (65.8 +/- 7.2%), although the absolute number of T lymphocytes was decreased in mastitic cows (1.49 +/- 0.91 X 10(6) cells/ml vs. 2.47 +/- 1.28 X 10(6) cells/ml). In the prescapular lymph node the percent of B lymphocytes, but not T or "null lymphocytes", decreased significantly in mastitic cows as compared to that of normals. The decrease, i.e. 32%, paralleled the 32.1% decrease found in peripheral blood B lymphocytes. In contrast, in the supramammary lymph node of mastitic cows, the percent B lymphocytes increased over that of normals (35.1 +/- 2.0% vs. 20.4 +/- 9.4%), whereas the percent T lymphocytes decreased to 54.5 +/- 2.8% compared to 70.7 +/- 3.5% in normal cows. There was no significant change in percent "null lymphocytes". The weight of prescapular lymph nodes did not change in mastitic cows when compared to that of normals. As a result, the estimated number of B lymphocytes, but not of T and "null lymphocytes", decreased.(ABSTRACT TRUNCATED AT 250 WORDS)     

Flow cytometric characterization of T lymphocyte subsets in the peripheral blood of Chinese rhesus macaques: normal range, age- and sex-related differences

Available data on the normal levels of white blood cell populations in healthy rhesus macaques (Macaca mulatta) originated and living in China is scanty. To obtain such data, blood samples from 150 Chinese rhesus macaques were collected and the normal range of white blood cells and their subsets were analyzed according to age and sex by flow cytometry. CBC data showed that the count of total white blood cells and lymphocytes decreased with age. Phenotypic analysis of CD4 and CD8 expression on CD3+ T lymphocytes showed that the percentage of CD4+ T cells (51.4+/-9.6%), CD4-CD8- T cells (8.5+/-4.1%) and the ratio of CD4+ T to CD8+ T cells (1.26+/-0.55) decreased with age; and the percentage of CD8+ T cells (42.0+/-9.7%), CD4+CD8+ T cells (1.3+/-0.9%) and CD3+ lymphocytes (55.3+/-13.3%) increased with age. However, no statistically significant difference was observed between the male and female groups in most parameters in these monkeys except for the percentage of CD4+CD8+ T cells. This study provided basic information about blood cell count and T lymphocyte subsets in Chinese rhesus macaques. It may be useful for comparative studies using Indian and Chinese rhesus macaques.     

Flow cytometric detection of bovine viral diarrhea virus in peripheral blood leukocytes of persistently infected cattle.

Flow cytometry was investigated for detection of bovine viral diarrhea virus (BVDV) in peripheral blood mononuclear leukocytes of persistently infected cattle. The mononuclear leukocytes were purified by sedimentation in a gradient of Ficoll-Paque, fixed, permeabilized, and then labelled by indirect immunofluorescence using biotinylated immunoglobulins from a porcine antiserum to BVDV. Flow cytometric analysis of blood samples obtained from persistently infected cattle revealed virus in 3.0-21.0% (mean +/- SD, 11.2% +/- 6.4%) of the mononuclear leukocytes. Fluorescent cells were not observed in controls. Flow cytometric detection of BVDV in blood cells of persistently infected bovines is a rapid and objective technique which does not require cell culture facilities.