Effect of theophylline on CD11b and L-selectin expression and density of eosinophils and neutrophils in vitro

The nonspecific phosphodiesterase inhibitor theophylline, widely used in asthma therapy, may cause a decrease in inflammatory responses of airways. In asthma, eosinophils migrate to the airway wall and become activated. Activated eosinophils are characterized by low cell density, as well as increased expression of CD11b and reduced expression of L-selectin, two adhesion molecules involved in transendothelial migration. To study the anti-inflammatory effect of theophylline on granulocyte adhesion molecules in vitro, the platelet-activating factor (PAF)-induced density shift was determined by density centrifugation and the modulation of CD11b and L-selectin expression by flow cytometry on eosinophils and neutrophils in human whole blood. A relatively high concentration of theophylline (10(-3) M) inhibited the increase in the percentage of hypodense eosinophils and neutrophils in whole-blood samples after PAF stimulation in vitro. A more pharmacological concentration (10(-4) M) inhibited the CD11b upregulation and L-selectin shedding induced by PAF (10(-7) M) on both eosinophils and neutrophils. The effect of isoproterenol on the inhibitory effect of theophylline was mainly additive, but a small synergistic effect could not be excluded. In conclusion theophylline can attenuate eosinophil and neutrophil activation in vitro at the level of adhesion molecule expression and changes in cell density. This may have implications for transendothelial migration of these cells in asthma.     

Our expanding South Carolina Medicaid managed care program

Eosinophil differentiation is thought to occur by the action of interleukin (IL)-5 on CD34(+) progenitor cells. The allergen-induced increase in eosinophil numbers in isolated airway preparations in vitro, and detection of increased numbers of circulating CD34(+) cells in atopic subjects, led us to the hypothesis that the eosinophil infiltration of the airway in asthma may result from local mucosal differentiation, in addition to recruitment from the bone marrow. We examined CD34(+) cell numbers by immunohistochemistry and IL-5 receptor alpha (IL-5Ralpha) messenger RNA (mRNA) expression by in situ hybridization in bronchial biopsies from atopic asthmatic patients, and from atopic and nonatopic control subjects. CD34(+) cell numbers were increased in the airway in atopic asthmatic and atopic nonasthmatic subjects. In contrast, CD34(+)/ IL-5Ralpha mRNA+ cells were increased in asthmatic subjects when compared with both atopic and nonatopic control subjects. Airway numbers of CD34(+)/IL-5Ralpha mRNA+ cells were correlated to airway caliber in asthmatic subjects and to eosinophil numbers. These findings support the concept that eosinophils may differentiate locally in the airway in asthma.     

Upregulation of alpha GM-CSF-receptor in nonatopic asthma but not in atopic asthma

BACKGROUND: Intrinsic asthma is characterized by an increased number of activated eosinophils and macrophages and an increased expression of the hematopoietic growth factor granulocyte-macrophage colony-stimulating factor (GM-CSF) in the bronchial mucosa. OBJECTIVE: This study was carried out to investigate the expression of alpha GM-CSF receptor (alpha GM-CSFr) messenger RNA and protein in the bronchial mucosa of patients with intrinsic or atopic asthma and of control subjects and to correlate the expression of alpha GM-CSFr to the number of EG2+ cells (eosinophils) and CD68+ cells (macrophages) and pulmonary function. METHODS: Nineteen patients with stable asthma (9 with atopic and 10 with intrinsic asthma) and 22 normal control subjects (12 atopic and 10 nonatopic subjects) were recruited, and FEV1 (percent predicted) and PC20 were measured before bronchoscopy. Endobronchial biopsy specimens were obtained and examined for membrane-bound alpha GM-CSFr by using in situ hybridization and immunocytochemistry. RESULTS: alpha GM-CSFr mRNA- and protein-positive cells were identified in biopsy specimens from all four groups studied. There was no significant difference in the number of cells expressing alpha GM-CSFr mRNA and protein in patients with atopic asthma compared with atopic and nonatopic control subjects. However, the numbers of alpha GM-CSFr mRNA- and protein-positive cells were significantly higher in nonatopic patients with asthma compared with atopic patients with asthma and atopic and nonatopic control subjects (p < 0.001). In the patients with intrinsic asthma, the number of alpha GM-CSFr mRNA-positive cells per millimeter of basement membrane correlated with numbers of CD68+ cells (r2 = 0.87, p < 0.001) but not with EG2+ cells, and colocalization studies demonstrated that 80% of the cells expressing alpha GMCSFr mRNA were CD68+. The expression of GM-CSF was also significantly increased in patients with intrinsic asthma compared with those with atopic asthma and control subjects (p < 0.05). In addition, in intrinsic asthma, there was a correlation between alpha GM-CSFr mRNA and FEV1 (r2 = 0.61, p < 0.05). CONCLUSION: These results demonstrate that elevated numbers of cells expressing alpha GM-CSFr can be detected in nonatopic asthma but not in atopic asthma and suggest that this increased expression is predominantly macrophage-associated and may play an important pathophysiologic role in intrinsic asthma.     

Comparison between eosinophilic markers in induced sputum and blood in asthmatic patients

BACKGROUND: The usefulness and safety of the analysis of blood inflammatory markers in asthma are widely recognized. Recently, the analysis of induced sputum has been proposed as a safe, non-invasive tool in the study of airway inflammation in asthma. OBJECTIVE: Our aim was to test whether sputum analysis is more useful than blood analysis in the evaluation of airway inflammation in untreated and treated asthmatic patients. METHODS: Twelve untreated patients with mild to moderate asthma underwent a methacholine challenge test, sputum induction and blood sampling. A group of 14 normal subjects was also evaluated for baseline comparison. The same evaluation was repeated after 3 months of budesonide treatment. Before and after treatment, we tested the relationship of eosinophilic markers in induced sputum and blood with clinical and functional data. We also compared eosinophilic markers in induced sputum with the same markers in blood. RESULTS: Untreated patients showed a significant relationship between sputum eosinophils and symptom score, and between sputum eosinophilic cationic protein and symptom score, FEV1 and PD20FEV1. No relationship between blood eosinophilic markers and clinical or functional data was observed. In budesonide-treated patients, both sputum and blood eosinophils were significantly lower than in untreated patients, but eosinophil decrease was greater in sputum than in blood. Sputum eosinophilic proteins were also significantly lower in treated patients, whereas serum eosinophilic proteins were low at baseline and remained unchanged after treatment. Sputum eosinophilic markers were lower in normal subjects than in both untreated and treated patients, while blood eosinophils, but not serum eosinophilic cationic protein, were lower in normals than in untreated patients. CONCLUSIONS: The analysis of induced sputum is more useful than the analysis of blood in the evaluation of asthma severity and of the effect of glucocorticoid treatment in patients with mild to moderate asthma.     

The advantages of the lateral decubitus position after spinal anesthesia with hyperbaric tetracaine

Exercise-induced bronchoconstriction (EIB) is widely prevalent in asthmatic patients. Eosinophilic airway inflammation is considered to be a major factor in the pathogenesis of asthma. However, the effects of eosinophilic airway inflammation on EIB have been elucidated insufficiently. To examine the relationship between the severity of EIB and eosinophilic inflammation, sputum induction and exercise challenge were performed in 21 asthmatic patients. Significantly higher percentages of eosinophils and levels of eosinophil cationic protein (ECP) were found in induced sputum in EIB-positive asthmatics (median (range), eosinophils: 23.5 (11.0-61.0)%; ECP: 1,475 (74.8-17,701) ng x mL(-1)) than in EIB-negative asthmatics (eosinophils: 6.0 (1.0-41.5)% (p=0.006); ECP: 270.6 (10.8-7,700) ng x mL(-1) (p=0.049)). There was a significant correlation between the severity of EIB and the sputum eosinophil percentage (r=0.59, p=0.009) and the level of ECP (r=0.47, p=0.037). The area under the curve of the forced expiratory volume in one second for 30 min after exercise correlated with the percentage of eosinophils (r=0.60, p=0.008) and the level of ECP (r=0.45, p=0.04). There was no correlation between airway responsiveness to methacholine on the one hand and EIB, sputum eosinophils or ECP on the other. In conclusion, these results provide evidence that the severity of bronchoconstriction evoked by exercise is more closely related to eosinophilic airway inflammation than airway hyperresponsiveness to methacholine in asthmatic patients.     

Effects of trauma and sepsis on soluble L-selectin and cell surface expression of L-selectin and CD11b

OBJECTIVES: To examine (1) the effects of trauma on changes in neutrophil L-selectin and CD11b expression and on the levels of soluble L-selectin and (2) whether these alterations are different on leukocyte subpopulations in those patients who develop multiple organ dysfunction syndrome. MATERIALS AND METHODS: Twenty patients with Injury Severity Score (ISS) > or = 16 and 15 patients with ISS score < 16 were studied. Arterial blood were collected serially after injury. The staining of leukocyte surface adhesion molecules was performed with antibodies against L-selectin and CD11b. Positive cell count and mean fluorescence intensity were determined by flow cytometry. Soluble L-selectin was measured using enzyme-linked immunosorbent assay. RESULTS: In patients with ISS > or = 16, neutrophil L-selectin expression showed an immediate increase, reaching peak levels between 3 to 4 hours after injury (p < 0.05 vs. patients with ISS < 16), followed by a gradual decrease. Plasma levels of soluble L-selectin reached peak levels at 6 hours after injury. However, in patients with ISS < 16, minimal changes in L-selectin expression and soluble L-selectin were observed. Neutrophil CD11b expression showed an immediate increase for the first 3 hours followed by a gradual increase up to 24 hours after injury. In patients who developed multiple organ dysfunction syndrome, CD11b both on neutrophils and lymphocytes remained elevated for 120 hours. CONCLUSIONS: These findings suggest that acute neutrophil activation is an early event after trauma and may be implicated as "a vulnerable window" for leukocyte-mediated end organ injury.     

Anisotropic expansion in gypsum-bonded cristobalite investment mold

OBJECTIVE: To determine the response of white blood cells in endovascular aortic aneurysm repair. MATERIALS AND METHODS: Seven patients treated with an endoluminal procedure (AAA-E) and seven patients undergoing conventional surgery (AAA-C) were included (all males, aged 52-80 years). A panel of monoclonal antibodies against CD11a, CD11b, CD11c, CD18 and L-selectin was used. To determine the surface receptors on both circulating and sequestered white blood cells, plasma from the patients and cells from healthy donors were combined for flow-cytometry. RESULTS: The expression of CD11a adhesion molecules only showed slight variations regarding granulocytes, but was more pronounced on monocytes, however, without significant differences between the two patient groups, CD11b, CD11c and CD18 molecules on both granulocytes and monocytes were significantly upregulated 60 min after the endovascular procedure compared to conventional aneurysm repair, and L-selectin molecules were by this time correspondingly cleaved off. CONCLUSION: Endovascular aneurysm repair differed significantly from conventional aneurysm surgery with peak adhesion molecule expression 60 min after balloon deflation, probably caused by release of tumour necrosis factor-alpha (TNF-alpha).     

BAL neutrophilia in asthmatic patients. A by-product of eosinophil recruitment?

Although neutrophil number may be increased in the airways of patients with asthma, its pathogenetic role in this disorder remains unclear. We evaluated BAL of 8 normal control subjects, 30 +/- 2 years of age, and 24 patients with mild asthma: 17 patients with allergic asthma, 24 +/- 1 years of age, and 7 patients with nonallergic asthma, 30 +/- 1 years of age. The BAL of asthmatic patients showed increased numbers of neutrophils (p < 0.01), eosinophils (p < 0.01), and ciliated epithelial cells (p < 0.05) and increased concentrations of myeloperoxidase (MPO) (p < 0.01) compared with control subjects. Positive correlations were observed between the number of BAL neutrophils and eosinophils (Rs = 0.780, p < 0.0001) and between BAL neutrophil numbers and BAL MPO levels (Rs = 0.40, p < 0.05). No correlations were found between the following: (1) BAL eosinophils or neutrophils and BAL epithelial cells (p > 0.05, each comparison); (2) BAL neutrophils or eosinophils and log Pd15 methacholine (MCh) (p > 0.05, each comparison); or (3) BAL epithelial cells or log Pd15 MCh and BAL MPO (p > 0.05, each comparison). Dividing the patient population into two groups, allergic asthmatics and nonallergic asthmatics, similar BAL neutrophil, eosinophil, and epithelial cell numbers and similar MPO levels were found (p > 0.05, each comparison). In addition, the correlations between BAL neutrophils and eosinophils showed similar significance in the two patient subgroups (p > 0.05, each comparison). These results suggest that, both in allergic and nonallergic asthma, airway recruitment and activation of neutrophils occur as does parallel eosinophil migration. However, airway neutrophils do not seem to contribute significantly to epithelial cell injury or to airway hyperresponsiveness in the steady state.     

Granulocyte macrophage colony-stimulating factor is the main cytokine enhancing survival of eosinophils in asthmatic airways

Interleukin (IL)-3, IL-5 and granulocyte macrophage colony-stimulating factor (GM-CSF) prolong the survival of eosinophils, which are conspicuous in asthmatic airways, but it is still controversial which one plays a major role in enhancing the survival of eosinophils in asthmatic airways. The role of these cytokines in airway eosinophilia was investigated using bronchoalveolar lavage (BAL) fluids from 11 symptomatic and nine asymptomatic patients with asthma and eight normal subjects. Eosinophil survival-enhancing activity (ESEA) was measured by a numerical change in viable eosinophils isolated from the peripheral blood of atopic patients and cultured with BAL fluids. ESEA was characterized by neutralization with antibodies to IL-3, IL-5 and/or GM-CSF. The differential count of BAL cells was achieved using Diff-Quik stain. T-cell subsets and activated T-cells were analysed by flow cytometry with dual stain using monoclonal antibodies to CD3, CD4, CD8 and CD25. ESEA was detected in eight of 11 BAL fluids of symptomatic asthma, but not in those of normal controls or asymptomatic asthmatics. In six symptomatic asthmatics, the mean percentage of inhibition in ESEA by anti-GM-CSF was higher than that of anti-IL-5 as well as anti-IL-3 (p<0.05). A mixture of antibodies to IL-3, IL-5 and GM-CSF totally inhibited the ESEA in four cases. The ESEA correlated with the percentage of eosinophils (p<0.05) and that of CD25(+)CD4 lymphocytes (p<0.05) of BAL cells. In conclusion, granulocyte macrophage colony-stimulating factor, rather than interleukin-3 or -5, is associated with eosinophil survival-enhancing activity inside the airways of symptomatic asthmatics. The activation of CD4 lymphocytes is related to the elevation of such activity.     

Repeatability of cellular and soluble markers of inflammation in induced sputum from patients with asthma

Sputum induced by inhalation of nebulized hypertonic saline is increasingly used to monitor airways inflammation in asthma. The aim of this study was to assess the repeatability of measuring cellular and soluble markers of inflammation in whole sputum samples as obtained by sputum induction in patients both with mild and moderate-to-severe asthma. Twelve patients with mild, atopic asthma without inhaled steroid treatment and nine patients with moderate-to-severe, atopic asthma treated with inhaled steroids were studied on two separate days at least 2 days apart. Whole sputum samples, induced by inhalation of hypertonic (4.5%) saline, were homogenized, and analysed for differential cell counts and for concentrations of albumin, fibrinogen, interleukin-8 (IL-8), and eosinophil cationic protein (ECP). Repeatability was expressed as intraclass correlation coefficient (Ri), and as coefficient of repeatability (CR) in percentage cells or in doubling concentration. Samples from two patients with mild asthma contained more than 80% squamous cells and were excluded from analysis. The repeatability for cell differential counts in both groups combined was: for neutrophils, Ri = 0.57 and CR = 31.0; for eosinophils, Ri = 0.85 and CR = 12.4; and for lymphocytes, Ri = 0.76 and CR = 6.9. The repeatability of the fluid phase measurements was: for albumin, Ri = 0.71 and CR = 3.2; for fibrinogen, Ri = 0.88 and CR = 2.8; for IL-8, Ri = 0.66 and CR = 2.2; and for ECP, Ri = 0.82 and CR = 1.1. We conclude that the repeatability of cellular and soluble markers of inflammation in induced sputum from patients with mild and moderate-to-severe asthma is satisfactory. Hence, induced sputum, processed by using the whole expectorated sample, seems to be a valuable method to monitor airway inflammation in asthma.     

Neutrophil and platelet activation at balloon-injured coronary artery plaque in patients undergoing angioplasty

OBJECTIVES: This study sought to investigate changes in the expression of activation-dependent adhesion receptors on neutrophils and platelets after exposure to the balloon-injured coronary artery plaque. BACKGROUND: Activation of blood cells at the balloon-injured coronary artery plaque may contribute to abrupt vessel closure and late restenosis after percutaneous transluminal coronary angioplasty. METHODS: In 30 patients undergoing elective coronary angioplasty, blood specimens were obtained through the balloon catheter proximal to the plaque before dilation and distal to the plaque after dilation. Simultaneous blood samples obtained through the guiding catheter served as control samples. Total surface expression of the inducible fibrinogen receptor (CD41) and surface expression of the activated fibrinogen receptor (LIBS1) on platelets as well as Mac-1 (CD11b) and L-selectin (CD62L) surface expression on neutrophils were assessed by flow cytometry. RESULTS: After exposure to the dilated coronary artery plaque, surface expression of LIBS1 on platelets increased by 40.5 +/- 11.0 mean (+/-SE) fluorescence (p=0.001) and that of CD11b on neutrophils increased by 20.1 +/- 4.4 mean fluorescence (p=0.018). Concomitantly, anti-CD62L binding on neutrophils decreased by 6.6 +/- 2.4 mean fluorescence (p=0.022). In contrast, surface expression of the adhesion receptors did not change significantly between the coronary ostium and the prestenotic coronary segment. CONCLUSIONS: The results of this study demonstrate neutrophil and platelet activation at the balloon-injured coronary artery plaque. This cellular activation may serve as a target for pharmacologic interventions to improve the outcome of coronary angioplasty.     

Chemokine-dependent upregulation of CD11b on specific leukocyte subpopulations in human whole blood: effect of anticoagulant on rantes and MIP-1 beta stimulation

The effect of anticoagulant (heparin vs EDTA) on chemokine induced CD11b upregulation on neutrophils, eosinophils, and monocytes in human whole blood was determined. For most of the chemokines (IL-8, GRO-alpha, MCP-1, MIP-1 alpha) the difference in the response of leukocytes in EDTA anticoagulated blood vs those in heparinized blood was the degree of their maximal response, with a slightly higher maximal increase in CD11b expression usually seen in cells from EDTA anticoagulated blood. Two chemokines were exceptions to this: RANTES and MIP-1 beta. RANTES is considered to be a stimulator of monocytes and eosinophils and not of neutrophils. As expected, neutrophils in heparinized whole blood did not respond to RANTES; however, neutrophils in EDTA anticoagulated blood had a significant increase in CD11b when exposed to high concentrations (1 microM) of RANTES. RANTES-induced CD11b expression on monocytes and eosinophils in these samples were the same in either heparin or EDTA. In EDTA anticoagulated blood, MIP-1 beta did not elicit a response in either monocytes, eosinophils or neutrophils; however, in heparinized blood, all three cell types increased CD11b expression upon exposure to 1 microM MIP-1 beta.     

A new enzyme immunoassay to detect antibodies to arboviruses in the blood of wild birds

A selective recruitment of eosinophils to sites of inflammation is claimed to be controlled by regulation of cytokines, chemokines and adhesion molecules. In animal models, eotaxin has been suggested to be a potent chemokine since it in cooperation with interleukin-5 induce selective chemotaxis and infiltration of eosinophils to lung tissue after an allergen provocation. We have investigated the in vitro effect of eotaxin on human peripheral blood eosinophils with respect to CD11b/CD18 expression and adhesion properties to the matrix protein fibronectin. We did not find any effect of eotaxin per se on CD11b/CD18 expression, neither on eosinophils from healthy subjects nor from patients with asymptomatic pollen related asthma. However, eotaxin significantly upregulated the quantitative level of CD11b/CD18 and increased the adhesion to fibronectin in eosinophils from healthy subjects preincubated in vitro with interleukin-5, but not in eosinophils preincubated with fMLP. Moreover, eosinophils harvested 24 hours after an in vivo allergen inhalation provocation in asthmatics, upregulated CD11b/CD18 after in vitro incubation with eotaxin alone.     

Combined use of exhaled hydrogen peroxide and nitric oxide in monitoring asthma

Oxidative stress contributes to airway inflammation and exhaled hydrogen peroxide (H2O2) and nitric oxide (NO) are elevated in asthmatic patients. We determined the concentrations of expired H2O2 and NO in 116 asthmatic (72 stable steroid-naive, 30 stable steroid-treated, and 14 severe steroid-treated unstable patients) and in 35 healthy subjects, and studied the relation between exhaled H2O2, NO, FEV1, airway responsiveness, and eosinophils in induced sputum. Both exhaled H2O2 and NO levels were elevated in steroid-naive asthmatic patients compared with normal subjects (0.72 +/- 0.06 versus 0.27 +/- 0.04 microM and 29 +/- 1.9 versus 6.5 +/- 0. 32 ppb, respectively; p < 0.001) and were reduced in stable steroid-treated patients (0.43 +/- 0.08 microM, p < 0.05, and 9.9 +/- 0.97 ppb, p < 0.001). In unstable steroid-treated asthmatics, however, H2O2 levels were increased, but exhaled NO levels were low (0.78 +/- 0.16 microM and 6.7 +/- 1.0 ppb, respectively). There was a correlation between expired H2O2, sputum eosinophils and airway hyperresponsiveness (methacholine PC20). Exhaled NO also correlated with sputum eosinophils, but not with airway hyperresponsiveness. Our findings indicate that measurement of expired H2O2 and NO in asthmatic patients provides complementary data for monitoring of disease activity.     

Gross periostitis ossificans in mandibular osteomyelitis. Review of the English literature and radiographic variation

BACKGROUND: Although the airway epithelium participates in inflammation and repair, the circadian expression of epithelial cell markers involved in these processes has not been investigated. OBJECTIVE: We sought to determine whether expression of CD51 (vitronectin and fibronectin receptor), CD54 (intercellular adhesion molecule-1), HLA-DR (activation marker), CD29 (beta1 integrin), CD49b (collagen receptor), and CD11b (complement receptor) exhibit a circadian rhythm in asthma. METHODS: Eleven patients with nocturnal asthma (NA), 9 subjects with nonnocturnal asthma (NNA), and 10 control subjects underwent bronchoscopy at 4 PM and 4 AM in a random order 1 week apart, with brushing of the proximal and distal airways. The percentage of cells staining for a particular marker was determined. RESULTS: At 4 PM, HLA-DR in the proximal airways and CD54 in the distal airways was significantly greater in control subjects as compared with asthmatic subjects (HLA-DR, control subjects: 10.0% [range, 5.0% to 21.0%]; NNA: 8.0% [range, 4.0% to 14.5%] NA: 3.5% [range, 2.0% to 6.0%], P = .01; CD54, control subjects: 17.0% [range, 8.0% to 25.0%], NNA: 8.0% [range, 5.3% to 11.5%], NA: 7.0% [range, 4.0% to 15.0%], P = .O;). At 4 AM, CD51 in the distal airways was significantly greater in patients with NA as compared with patients with NNA and control subjects (control subjects, 23.0% [range, 13.8% to 30.5%]; NNA, 32.0% [range, 13.0% to 35.0%]; NA, 40.0% [range, 23.0% to 50.0%], P = .05). Expression of CD51 in the distal airways correlated with the degree of airway obstruction (r = -0.57, P = .001). Control subjects exhibited significant circadian variation in the expression of HLA-DR in the proximal airways and CD54 in the distal airways. CONCLUSION: The increased CD51 at night in patients with NA may be related to increased airway inflammation and repair processes in response to injury. The circadian changes in CD54 and HLA-DR in control subjects require further study to determine their significance. (J Allergy Clin     

Expression of L-selectin on peripheral memory CD4+ T cells in atopic diseases

L-selectin is one of the adhesion molecules, which is known as a leukocyte homing receptor. L-selectin contributes labile rolling adhesion of leukocytes on the blood vessel wall. To elucidate characteristics of L-selectin expression on peripheral CD4+ lymphocytes in atopic diseases, flow cytometric analysis was performed. Subjects were 7 patients with atopic asthma (BA), 10 with atopic dermatitis (AD), 9 with allergic rhinitis (AR) and 10 normal adults (control). Peripheral lymphocytes in all subjects were three-color-analyzed using monoclonal antibodies. As a result, compared with the control group. BA and AD showed significantly higher proportions of L-selectin-positive memory CD4+ T cells (BA 64.0%; AD 62.0%; AR 50.9%; Control 38.0%), and significantly lower proportions of L-selectin-negative memory CD4+ T cells (BA 18.3%; AD 11.6%; AR 21.1%; Control 29.2%), respectively (reported percentages represent average values for each subject group). As previously reported. L-selectin-positive memory CD4+ T cells produce mainly Th2-type cytokines (Kanegane et al., 1996); Given these results, the present study suggests that increased Th2-like cells express L-selectin for extravasation in the process of local infiltration in atopic diseases.     

Crown-root fracture of lower molar--restorative procedures

An understanding of the mechanisms of post-injury leukocyte trafficking is essential to the development of future therapeutic interventions aimed at preventing infection and multiple organ failure in trauma patients, yet very little is known about the cellular and molecular events resulting in mobilization of members of the leukocyte family following trauma. We have studied the post-injury expression of the lymphocyte, monocyte and neutrophil adhesion molecules CD11a (LFA-1), CD11b, CD11c, CD29 (beta-1 integrin) and CD62L (L-selectin) in a group of 36 trauma patients, 13 of whom had suffered major trauma (ISS > or = 16), 15 moderate trauma (ISS = 9-15) and eight minor trauma (ISS < 9). Three ml blood samples were taken within 2.5 h of injury (mean sample time = 1.2 h, median = 1 h) into EDTA anticoagulant. Fifty-three normal control subjects were also studied for comparison. Leukocytes were stained using fluorescent-labelled monoclonal antibodies specific for each adhesion molecule, and the mean receptor density per cell measured using flow cytometry. Monocytes, neutrophils and lymphocytes in the trauma patients showed significantly increased mean-receptor density of L-selectin (p < 0.0001, 0.0001 and 0.004 respectively). Neutrophils and monocytes showed a significantly decreased level of expression of CD11a, and neutrophils showed a significant decrease in expression of CD11c. Our results indicate that there is a reduction in CD11a expression after trauma which may play an important role in the demargination of neutrophils and monocytes. The strong increase in L-selectin expression in all cell populations was unexpected, and is potentially important because this molecule supports rolling behaviour in all members of the leukocyte family, and would promote close contact between leukocytes and the endothelium at the site of injury without firm adhesion taking place. These events may be of significance in planning future strategies to combat post-trauma complications.     

Frequencies of T cells expressing interleukin-4 and interleukin-5 in atopic asthmatic children. Comparison with atopic asthmatic adults

T-cell-derived cytokines have been implicated in the pathogenesis of asthma and it has been suggested that Th2-type cytokines (interleukin-4 [IL-4], interleukin-5 [IL-5]) are pivotal in the allergic inflammation. However, there are little data on human cytokine production by individual T cells at the protein level, in particular in asthmatic children. In this study we analyzed the cytokine production at the single cell level in peripheral blood from mild atopic asthmatic (AA) children and adults and age-matched atopic nonasthmatic (AN) and nonatopic nonasthmatic (NN) control subjects (n = 9 in each group) using the technique of intracellular cytokine detection by flow cytometry. Comparing asthmatic children with atopic and nonatopic control subjects, an increased percentage of IL-5-producing T cells (AA: median 4.9% [range 1.1 to 8.9%]; AN: 0.3% [0.2 to 0.9%], p = 0.003; NN: 0.4% [0.1 to 3.8%], p = 0.001) was detectable, with a positive correlation to the number of peripheral eosinophils and to bronchial hyperresponsiveness. The frequency of IL-4-producing T cells was increased in both atopic groups compared with nonatopic controls (AA: 1.2% [0.2 to 2.6%], p = 0.011; AN: 0.8% [0.4 to 3.7%], p = 0.007; NN: 0.4% [0.2 to 0.9%]) with a positive correlation to total IgE concentration. In adults there were no differences in IL-5- or IL-4-producing T cells between all three groups. A substantial proportion of T cells coproducing IL-4 and IL-5 was not detectable in children and adults. These findings indicate that in asthmatic children the frequencies of Th2-type-producing T cells are increased and that expression of IL-4 and IL-5 is regulated independently.