Autoreactive eosinophils associated with spontaneous autocytotoxicity in bronchial asthma

Allergic autocytotoxicity (ACT) assay is extensively investigated as anin vitro equivalent of oral ingestion challenge with food antigens among patients with food hypersensitivity. In direct ACT, food antigenic determinants interact directly with plasma cell membranes of sensitive individuals. Antibody-dependent ACT is also known as antibody-dependent-cell-mediated-cytotoxicity (ADCC) phenomenon, when food antigens combine with specific antibody through cell membranes Fc receptors among normal and sensitive individuals. Spontaneous ACT is a separate mechanism of white blood cells disintegration which does not requirein vitro priming of the cells, neither by antigen or antibody. Spontaneous ACT occurs in some individuals as background noise in addition to direct and antibody-dependent ACT. The exact cellular nature of ACT phenomena are unknown at the present time with the exception that the common identifying factor for each of them is the disintegration and death of human white blood cells.Electron microscopy studies among four bronchial asthma patients with spontaneous ACT demonstrated eosinophils with atypical crystalloid cores and diffuse autolytic pattern of granular membranes. These ultrastructural characteristics are associated with new functional profiles of eosinophils expressed morphologically as natural killer and/or suicidal potency. At least two subpopulations of eosinophils are mediating ACT. The first subpopulation has normal ultrastructure observed in direct ACT and the second subpopulations has altered morphology of eosinophils granules described in spontaneous ACT. The natural killer-suicidal eosinophils presented in patient with spontaneous ACT illustrate a new pathway of cytodestructive mechanism in anaphylactic injury.These results were presented in part at the XII International Congress of Allergology and Clinical Immunology, Washington, D.C., October 20–25, 1985.     

Immunopathology of the bronchial mucosa in 'late onset' asthma

A clinical, pathological and immunological study was made of fifty cases of asthma developing in adult life—twenty-seven females and twenty-three males. The mean age of appearance of asthma was 51 years. In spite of the paroxysmal onset observed in thirty-nine cases, the course was always severe, leading to dependence upon corticosteroid treatment in thirty-six cases. Two-thirds of the subjects had either a personal or familial atopic history, one-third had positive skin tests to routine respiratory allergens, and one-third had a peripheral blood eosinophilia higher than 400 mm³. Total serum IgE was not raised in any case and attempts at hyposensitization were not effective. The titre of serum complement was normal and investigation for autoantibodies was negative in thirty-one subjects. Study of the HLA system in twenty-eight cases showed a significantly raised frequency of antigen HLA9. The bronchial biopsies were examined histologically in thirty-one cases and by fluorescence examination in fifteen. The histological appearance was sterotyped and non-specific. There was some modification of the lining consisting of thickening and irregularity of the basal membrane with evidence of hyperfunction of adjacent mesenchymal tissues. The basal membrane showed thickening, estimated in two cases by electron microscopy to be of 20–30 μm. This was due largely to collagen, which concealed the true basal membrane, the ultrastructure of which was well preserved. Immunofluorescence tests made it possible to show the presence of IgA, IgG and IgM constantly associated with the C3 component of complement. This was localized mainly in the superficial part of the thickened basal membrane and in the cytoplasm of the epithelioid cells. Fibrin showed a different appearance, with irregular fluorescence situated essentially along the base of the superficial epithelial membrane. These immunofluorescence findings are compatible with transudation and also of a local immunological reaction.     

Clinical and socio-professional fate of isocyanate-induced asthma

Thirty-one patients with isocyanate-induced asthma were studied 6–54 months after diagnosis. Four had the same work conditions and unchanged or worse respiratory symptoms; seven had an alternative job or safer work conditions at the same workplace and suffered from mild to severe symptoms. The remaining twenty subjects were definitely removed from exposure; of these, ten (50%) remained symptomatic after being removed from exposure for an average of 19 months. Asymptomatic patients appeared to be younger and to have shorter durations of total and symptomatic exposures, while symptomatic patients were more reactive to acetylcholine at diagnosis. For patients removed from isocyanate exposure and for those re-employed at the same work-place, quality of the new work site seems to play a role in the evolution of isocyanate-induced asthma.     

T cells and eosinophils in bronchial smooth muscle cell death in asthma

Background Bronchial smooth muscle cells (SMC) proliferate, express adhesion molecules, secrete cytokines and thus efficiently contribute to the pathogenesis of asthma.
Objective The aim of the study was to investigate whether, and by which mechanism, T cells and eosinophils can cause death of airway SMC.
Methods The T cell- and eosinophil-induced cell death was analysed in primary human bronchial SMC cultures as well as in bronchial biopsy specimens from non-asthmatic and asthmatic individuals.
Results Bronchial SMC death showed characteristic morphological features of apoptosis in 3–6 days cultures with inflammatory cytokines (IFN-γ, TNF-α), soluble death ligands [sFasL, TNF-related apoptosis-inducing ligand (TRAIL)] and activated T-helper type 1 (Th1) and Th2 cell supernatants. The recombinant eosinophil cationic protein induced SMC necrosis within 1 h. Resting SMC expressed the death receptors TNFR1, TNFR2, Fas, TRAILR1, TRAILR2 and membrane FasL as a death-inducing ligand. IFN-γ and TNF-α up-regulated TNFR1, TNFR2, Fas and membrane FasL on SMC. TNF-α up-regulated TRAILR1 and TRAILR2; sFasL up-regulated TNFR2. The intracellular caspase-3 activation in SMC was significantly increased by IFN-γ, sFasL, TRAIL, Th1 and Th2 cell supernatants. Increased expression of TRAIL in asthmatics, but not in non-asthmatic individuals was demonstrated in situ . The apoptosis receptors TRAILR1 and TRAILR2 were expressed in SMC and epithelial cells both in healthy and asthmatic biopsies. Prominent apoptosis of SMC was observed in fatal asthma, but not intermittent asthma biopses.
Conclusion The demonstration of bronchial SMC death both by apoptosis and necrosis indicates the essential role of T cells and eosinophils in the bronchial tissue injury particularly in the severe asthma.     

T cells and eosinophils cooperate in the induction of bronchial epithelial cell apoptosis in asthma

BACKGROUND: Asthma is an inflammatory airway disease associated with an infiltration of T cells and eosinophils, increased levels of pro-inflammatory cytokines, and shedding of bronchial epithelial cells (ECs). OBJECTIVE: Shedding of bronchial ECs is characterized by loss of the normal bronchial pseudostratified epithelium and the maintenance of a few basal cells on a thickened basement membrane. The aim of this study was to investigate whether, and by which mechanism, T cells and eosinophils can cause damage to airway ECs. METHODS: Bronchial ECs, cultured and exposed to cytokines, eosinophil cationic protein, activated T cells, and eosinophils were studied for the expression of apoptosis receptors (flow cytometry, immunoblotting, and RNA expression) and for the susceptibility for undergoing apoptosis. In addition, bronchial biopsy specimens from patients with asthma were evaluated for EC apoptosis. RESULTS: We demonstrate herein that the respiratory epithelium is an essential target of the inflammatory attack by T cells and eosinophils. Bronchial ECs underwent cytokine-induced cell death with DNA fragmentation and morphologic characteristics of apoptosis mediated by activated T cells and eosinophils. T cell- and eosinophil-induced EC apoptosis was blocked by inhibition of IFN-alpha and TNF-alpha; the Fas ligand-Fas pathway appears to be less important. Recombinant eosinophil cationic protein induced mainly necrosis of ECs. Furthermore, we demonstrated in situ apoptotic features of ECs in bronchial biopsy specimens of asthmatic patients. CONCLUSION: T cell- and eosinophil-induced apoptosis represents a key pathogenic event leading to EC shedding in asthma.     

Leukotrienes and isocyanate-induced asthma: a pilot study

BACKGROUND: The role of leukotrienes (LTs) in the pathophysiology of isocyanate-induced asthma is not well known. OBJECTIVE: We sought to characterize the type of airway inflammation induced by exposure to isocyanates and to investigate whether exposure to isocyanates induced an increase in LT receptor cysteinyl leukotriene ((CysLT)(1), CysLT(2) and leukotriene B(4) receptor (BLT(1))) expression, as well as a release of LT (LTC(4) and leukotriene B(4) (LTB(4))) and IL-8 in both asthmatics with isocyanate-induced asthma and healthy subjects. METHODS: We investigated eight subjects with isocyanate-induced asthma and eight healthy subjects. Both groups underwent specific inhalation challenges to isocyanates in the laboratory. Induced sputum was collected before and after exposure to isocyanates. CysLT(1), CysLT(2) and BLT(1) expression was assessed by flow cytometry, whereas LTC(4), LTB(4) and IL-8 were measured in the sputum supernatants by enzyme immunoassay. RESULTS: Exposure to isocyanates induced an increase in sputum neutrophils only in subjects with occupational asthma. There was a significant increase in CysLT(1) and BLT(1) receptor expression, as well as a release of LTB(4) and IL-8 after exposure to isocyanates compared with the baseline, only in subjects with isocyanate-induced asthma, whereas there was no increase in LTC(4). Exposure to isocyanates did not induce any change in LT receptor expression nor in the levels of LTC(4), LTB(4) and IL-8, in healthy subjects. CONCLUSION: The neutrophilia observed after exposure to isocyanates is likely to be related to the release of LTB(4), probably enhanced by the increased expression of BLT(1) on neutrophils as well as by the release of IL-8. The significance of the increase of CysLT1 receptor expression on neutrophils is unknown and needs further investigation.     

Comparative morphological characteristics of bronchial mucosa in patients with chronic obstructive bronchitis and bronchial asthma

The material for histoultrastructural study was biopsies obtained by fibrobronchoscopy from 12 patients with endogenic bronchial asthma (BA) and 4 patients with chronic obstructive bronchitis (COB). The principal periods of affection in these diseases, the role of bronchial epithelium, effector cells of inflammation, nervous elements, smooth muscle cells, microcirculatory disturbances in patho- and morphogenesis of chronic obstructive pulmonary lesions are determined. The results indicate that BA develops due to disturbances of adaptive mechanisms being in some cases the issue of COB progression.     

Inhaled corticosteroids decrease vascularity of the bronchial mucosa in patients with asthma

BACKGROUND: Increased vascularity in airway mucosa is a distinctive feature of airway remodelling in asthma. While corticosteroids have proved most effective in modifying airway inflammation, the effect of inhaled corticosteroids on increased airway mucosal vascularity in asthmatics has been little studied. OBJECTIVE: We examined the effect of inhaled corticosteroid on airway vascularity in bronchial biopsy specimens taken from asthmatic patients. SUBJECTS AND METHODS: We studied bronchial biopsies from 28 asthmatic patients before and after treatment with inhaled beclomethasone dipropionate (BDP) 800 microg/daily, or placebo, for 6 months in a double-blind manner. Biopsy specimens were evaluated for number of vessels and percentage of area occupied by vessels, using computerized image analysis after staining for type IV collagen in vessel walls. Specimens were also examined for extent of collagen III in the subepithelial basement membrane. In addition, we compared asthmatic specimens with biopsy specimens taken from non-asthmatic control subjects. RESULTS: There was a significant increase in number of vessels (P < 0.01) and percent vascularity (P < 0.001) in the submucosa of asthmatic patients compared with control subjects. After 6 months of treatment, we observed significant improvements in forced expiratory volume in 1 s (FEV1), FEV1% and airway responsiveness (P < 0.05, each) in the BDP treatment group compared with the placebo group. This was accompanied by significant decreases in both vessel number and percent vascularity in the airways of BDP-treated patients (P < 0.05, each). We also observed a significant correlation between change in percent vascularity and change in collagen III thickness in the BDP-treated patients (rs = 0.90, P < 0.001). Furthermore, the change in percent vascularity was inversely correlated with both FEV1 (rs = -0.49, P < 0.05) and airway responsiveness (rs = -0.36, P < 0.05). CONCLUSION: These findings suggest that inhaled corticosteroid treatment of asthma reduced airway wall vascularity during airway remodelling.     

T cells and eosinophils in the pathogenesis of asthma.

Persistent asthma is characterized by chronic inflammation of the bronchial mucosa, where T cells and eosinophils are prominent. This article summarizes the evidence that asthmatic bronchial inflammation is initiated and propagated by cytokines secreted by activated T cells and other cells, and describes how the release of specific cytokines could result in local preferential accumulation and activation of eosinophils.     

Soluble human leucocyte antigen-G and interleukin-10 levels in isocyanate-induced asthma

Background We previously reported that in moderate-to-severe asthma there is a deficit of IL-10 secretion that could prevent the production of soluble HLA-G (sHLA-G), a non-classical human leucocyte antigen class I molecule with tissue-protective properties in inflammatory responses.
Objective Our objective was to investigate the production of sHLA-G and the secretion of IL-10 by peripheral blood mononuclear cells (PBMCs) in asthma induced by isocyanates and to compare the results with those obtained in non-occupational allergic asthma.
Method sHLA-G and IL-10 were measured by ELISA in the culture supernatants of unstimulated or lipopolysaccharide (LPS)-stimulated PBMCs obtained from 20 subjects with isocyanate asthma, 16 asymptomatic subjects exposed to isocyanates, 18 subjects with non-occupational allergic asthma, and 26 healthy control subjects.
Results Occupational exposure to isocyanates was associated with high baseline levels of secretion of IL-10 by PBMCs, whether or not the exposed subjects had asthmatic symptoms. However, spontaneous production of sHLA-G by PBMC was significantly higher in subjects with isocyanate asthma compared with asymptomatic-exposed controls. In contrast, PBMCs from subjects with non-occupational allergic asthma produced sHLA-G only after LPS stimulation.
Conclusions sHLA-G production and IL-10 secretion are influenced by workplace exposure to isocyanates and by development of asthma. The different behaviour of both sHLA-G and IL-10 in asthma induced by isocyanates compared with non-occupational allergic asthma suggests a heterogeneous biological role for HLA-G molecules and for IL-10, a key cytokine of immune and inflammatory responses.     

T-lymphocytes of the gastric mucosa in peptic ulcer

Chronic ulcer edges and pylorus of 6 stomachs removed because of ulcer and 10 biopsies of the stomach pylorus from patients with duodenal ulcer were studied. T-helpers and T-suppressors (cytotoxic) were revealed by the indirect three-step immunoperoxidase method with the use of monoclonal antibodies OKT-4 and OKT-8, respectively. T-suppressors (cytotoxic) prevailed among interepithelial gastric lymphocytes in both normal conditions and ulcer. T-helpers dominated in the mucosa tunica propria. They ensure immunoglobulin synthesis after the antigen stimulation and favour the activation of other immunocompetent cells. Both cell types are found in the lymphoid nodules and their centres. Helicobacter pylori seems to take part in the activation of all components of local immune system of the stomach.     

Flunisolide in chronic bronchial asthma.

Sixteen steroid dependent and 13 steroid independent patients with bronchial asthma were treated for three months with flunisolide by aerosol. Asthma improved in these patients, other medication usage decreased and adverse side effects were minimal. A significant increase in morning plasma cortisol levels occurred in steroid dependent patients, whereas cortisol levels in steroid independent patients remained normal. An unexpected decline in response to metyrapone occurred in both groups of patients and suggests that this test is affected by flunisolide usage.     

Endogenous nitric oxide downregulates the Bcl-2 expression of eosinophils through mitogen-activated protein kinase in bronchial asthma

BACKGROUND: Eosinophil apoptosis might play a crucial role in the maintenance of persistent airway inflammation in asthma. Nitric oxide (NO) synthase activity is upregulated in eosinophils of asthmatic patients. Mitogen-activated protein kinase (MAPK) is implicated in regulating eosinophil apoptosis by modulating Bcl-2 expression. NO-induced cell apoptosis is associated with an inhibition of Bcl-2 expression. OBJECTIVE: We sought to study whether NO might induce eosinophil apoptosis through extracellular signal-regulated protein kinase (ERK) or p38 MAPK pathways and Bcl-2 expression. METHODS: Eosinophils were freshly isolated from peripheral blood of 16 asthmatic patients and 12 healthy subjects and then cultured with or without the NO synthase inhibitor N-nitro-l-arginine methyl ester (L-NAME) at 1 and 10 mmol/L for 16 hours. The expression of Bcl-2 and induced NO synthase on eosinophils was analyzed by means of flow cytometry. Apoptosis was assessed by means of propidium iodide and DNA ladder. The activity of ERK and p38 MAPK was determined by means of Western blotting. RESULTS: The induced NO synthase immunoreactivities and the spontaneous release of nitrite from the eosinophils of asthmatic patients were higher compared with those of healthy subjects. Eosinophils of asthmatic patients were found to express more highly constitutive Bcl-2 than those of healthy subjects. After incubation for 16 hours, the expression of Bcl-2 on eosinophils from patients with asthma was significantly enhanced by L-NAME. The percentage of apoptosis was decreased by the addition of 1 mmol/L L-NAME in patients with asthma. The activity of p38 MAPK and ERK in eosinophils from patients with asthma was enhanced in the presence of L-NAME. An inhibition of MAPK reduced the Bcl-2 expression and increased eosinophil apoptosis in patients with asthma. CONCLUSION: We concluded that inhibition of endogenous NO might increase the expression of Bcl-2 in eosinophils from patients with asthma through the signaling pathway of ERK or p38 MAPK, which in turn decrease the apoptosis.     

Tryptophane metabolism in bronchial asthma.

Seventeen out of 21 children with severe asthma had an abnormal increase in urinary Kynurenic (KA) and Xanthurenic (XA) acid following oral tryptophane administration. In a second study 11 asthmatic children had significantly greater increases in XA and KA than nine normal healthy children. A micro-assay of KA and XA by column chromatography was performed. The data suggest the presence of a partial block in the metabolism of tryptophane in some children with bronchial asthma.     

Effects of ketotifen on symptoms and on bronchial mucosa in patients with atopic asthma

Ketotifen is marketed throughout the world as an antiallergy drug, but whether it affects infiltration of inflammatory cells into airway mucosa is not known. We studied the effects of ketotifen on symptoms, pulmonary function, and airway inflammation in 25 patients with atopic asthma. Patients took ketotifen (1 mg twice daily) or a matching placebo for 8 weeks in a double-blind, parallel-group study. Data recorded on diary cards were used for 2 weeks before treatment began, and they were used for the last 2 weeks of treatment to study asthma symptoms, use of β2–agonists, and peak expiratory flow (PEF). Pulmonary function tests, bronchial responsiveness to methacholine, and fiberoptic bronchoscopy were performed before and after treatment. Biopsy specimens were obtained by bronchoscopy. Specimens were stained immunohistochemically with monoclonal antibodies against stored eosinophil cationic protein (EG1), the secreted form of eosinophil cationic protein (EG2), mast-cell tryptase (AA1), neutrophil elastase (NP57), CD3, CD4, CD8, and CD25. The numbers of positively stained cells in the lamina propria were counted. Compared with the placebo, the ketotifen-treated group exhibited significant improvement of asthma symptoms ( P <0.05) and bronchial responsiveness (P<0.05). This was accompanied by a reduction of EG2+ eosinophils ( P <0.05), CD3+ T cells ( P <0.001), CD4+ T cells (P<0.01), and CD25+ activated T cells ( P <0.01) in the bronchial mucosa. These results suggested that the beneficial effects of ketotifen in bronchial asthma may result from consequent inhibition of activated eosinophils and T-cell recruitment into the airway. Moreover, ketotifen may relieve allergic inflammation in bronchial asthma.     

Hypnotherapy in the treatment of bronchial asthma.

The efficacy of hypnotherapy in aborting acute asthmatic attacks was studied in 17 children ranging in age from six to 17. All had as their primary diagnosis bronchial asthma. Prior to hypnotic induction pulmonary function was assessed, then monitored in the immediate post hypnotic period and at two intervals thereafter. The average improvement for all subjects was greater than 50% above the baseline measurement as documented by spirometry, monitored dyspnea, wheezing and subjective ratings by the subjects. It is suggested that hypnotherapy may be an important tool in ameliorating asthma, improving ventilatory capacity and promoting relaxation without recourse to pharmacologic agents. One explanation offered is that hypnosis affects an automic response, thereby diminishing bronchospasm.