bromochloroacetic-acid and Asthma

The bronchial epithelium is likely to play a vital role in airway diseases in children, such as asthma and viral-associated wheeze. In adults, studies with primary bronchial epithelial cells cultured from samples obtained by fibre-optic bronchoscopy have provided key insights into the role of the epithelial cell. However, it is difficult to justify bronchoscopy in children to obtain epithelial cells for research purposes.. To examine the possibility of retrieving and culturing viable epithelial cells using a blind non-bronchoscopic method from children undergoing elective surgery.. Subjects were children undergoing elective surgery under general anaesthesia. Following intubation, non-bronchoscopic bronchoalveolar lavage and non-bronchoscopic bronchial brushing were performed. A sheathed bronchial cytology brush was advanced through the endotracheal tube, wedged and then withdrawn 2-3 cm before gentle sampling was used to collect bronchial epithelial cells. Initial samples were used to characterize the number, type and viability of epithelial cells recovered compared to a control group of adults undergoing standard bronchoscopic sampling. Subsequent samples were used to establish primary bronchial epithelial cell cultures in children both with and without wheezing illness.. A total of 63 children underwent bronchial brushing [38 male; median age 7.1 years (1.0-14.2 years]. Initial samples (n=30) showed recovery of viable epithelial cells comparable to that from a single brush obtained via a bronchoscope in an adult control group (n=11). In 27 (82%) of the subsequent 33 samples obtained non-bronchoscopically from children, primary bronchial epithelial cell cultures were successfully established. There were no adverse effects attributable to sampling.. We have shown that non-bronchoscopic bronchial brushing is a safe and effective technique for recovering viable bronchial epithelial cells that consistently yield primary cultures. This method will facilitate examination of the role of the epithelium in paediatric disease.

Topics: Adolescent; Asthma; Bronchi; Bronchoalveolar Lavage; Cell Survival; Cells, Cultured; Child; Child, Preschool; Cytodiagnosis; Epithelial Cells; Feasibility Studies; Female; Humans; Immunohistochemistry; Infant; Keratins; Male; Respiratory Sounds; Specimen Handling

Although the airway epithelium participates in inflammation and repair, the circadian expression of epithelial cell markers involved in these processes has not been investigated.. 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.. 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.. 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.. 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

Topics: Adult; Antigens, CD; Asthma; Biomarkers; Bronchi; Bronchoscopy; Circadian Rhythm; Epithelium; Female; Forced Expiratory Volume; HLA-DR Antigens; Humans; Keratins; Male; Staining and Labeling; Trachea; Vital Capacity

This study aimed to confirm the ameliorative effect of ghrelin on asthma and investigate its mechanism.. The murine model of asthma was induced by ovalbumin (OVA) treatment and assessed by histological pathology and airway responsiveness to methacholine. The total and differential leukocytes were counted. Tumor necrosis factor α, interferon γ, interleukin-5 and interleukin-13 levels in bronchoalveolar lavage fluid were quantified by commercial kits. The protein levels in pulmonary tissues were measured by Western blot analysis.. Ghrelin ameliorated the histological pathology and airway hyperresponsiveness in the OVA-induced asthmatic mouse model. Consistently, OVA-increased total and differential leukocytes and levels of tumor necrosis factor α, interferon γ, interleukin-5 and interleukin-13 in bronchoalveolar lavage fluid were significantly attenuated by ghrelin. Ghrelin prevented the increased protein levels of the endoplasmic reticulum stress markers glucose regulated protein 78 and CCAAT/enhancer binding protein homologous protein and reversed the reduced levels of p-Akt in asthmatic mice.. Ghrelin might prevent endoplasmic reticulum stress activation by stimulating the Akt signaling pathway, which attenuated inflammation and ameliorated asthma in mice. Ghrelin might be a new target for asthma therapy.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Blotting, Western; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Endoplasmic Reticulum; Ghrelin; Keratins; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred C57BL; Stress, Physiological

Several studies have suggested the involvement of an autoimmune mechanism in aspirin (ASA)-intolerant asthma. To test this hypothesis, we measured the levels of circulating autoantibodies, such as IgG and IgA to tissue transglutaminase (TGase), IgG to cytokeratins (CKs) 8, 18, and 19, Clq-binding immune complex (CIC), and antinuclear antibody (ANA), in the sera of 79 patients with ASA-intolerant asthma (Group I) and those of two control groups, consisting of 61 patients with ASA-tolerant asthma (Group II) and 88 healthy control subjects (Group III) by means of ELISA. Significantly higher prevalences of IgG antibodies to CK18 (13.9%) and CK19 (17.7%) were noted in Group I, as compared with Group III (p<0.05 for all) not with Group II. Regarding the prevalences of other autoantibodies, the levels of ANA (1.3%), IgG to TGase (3.8%), and CIC (24.7%) in Group I were not significantly different from those in Groups II and III. Significant correlations were found between positivities for the anti-CK18 and anti-CK19 autoantibodies and the PC(20) methacholine values in the analysis of asthma Groups I and II vs. normal controls, (p=0.001 and p=0.003, respectively). Further studies are needed to explore the potential involvement of an autoantibody-mediated mechanism in the clinical manifestation of bronchial asthma.

Topics: Aged; Anti-Inflammatory Agents, Non-Steroidal; Aspirin; Asthma; Autoantibodies; Bronchi; Case-Control Studies; Child; Drug Resistance; Female; Humans; Inflammation; Keratins; Male; Middle Aged

Cytokeratin 18 (CK18) protein was identified as an airway epithelial cell autoantigen associated with nonallergic asthma. Cleavage of CK18 protein by caspase-3 is a marker of early apoptosis in epithelial cells. It has been shown that the expression of active caspase-3 was increased in bronchial epithelial cells of asthmatic patients, when compared with healthy controls. To investigate the antigen-binding characteristics of IgG autoantibodies to CK18 protein in nonallergic asthma, the bindings of IgG autoantibodies to the fragments of CK18 protein cleaved by caspase-3 were analyzed by Western blot using serum samples from three patients with nonallergic asthma. Recombinant human CK18 protein was treated by caspase-3 and cleaved into N-terminal fragment (1-397 amino acids) and C-terminal fragment (398-430 amino acids). The binding capacity of IgG autoantibodies to N-terminal fragment of CK18 was maintained in one patient and reduced in other two patients. IgG autoantibodies from all three patients did not bind to C-terminal fragment of CK 18. In conclusion, IgG autoantibodies to CK18 protein from patients with nonallergic asthma seems to preferentially bind to the whole molecule of CK18 protein and their antigen-binding characteristics were heterogeneous among the patients with nonallergic asthma.

Topics: Adult; Aged; Antibodies, Monoclonal; Antigen-Antibody Reactions; Asthma; Autoantibodies; Blotting, Western; Caspase 3; Caspases; Epitopes; Female; Humans; Hydrolysis; Immunoglobulin G; Keratins; Male; Peptide Fragments; Protein Binding

To evaluate the clinical significance of autoantibodies to three major epithelial cytokeratins (CK)--CK8, CK18, and CK19--we compared 66 patients with toluene diisocyanate (TDI)-induced asthma (group I) with three control groups: 169 asymptomatic exposed subjects (group II), 64 patients with allergic asthma (group III), and 123 unexposed healthy subjects (group IV). Serum IgG, specific for human recombinant CKs, were measured by ELISA (enzyme linked immunosorbent assay), and ELISA inhibition tests were performed. The existence of these antibodies was confirmed by IgG immunoblot analysis. Anti-TDI-HSA (human serum albumin) IgE and IgG antibodies were measured by ELISA in the same set of the patients. The prevalence of CK8, CK18, and CK19 auotantibodies in group I was significantly higher than in the other three groups. Results of the ELISA inhibition test showed significant inhibition with the addition of three CKs in a dose-dependent manner. No significant association was found between CK autoantibodies and the prevalence of anti- TDI-HSA IgG and IgE antibodies. These results suggest that autoantibodies to CK18 and CK19 can be used as serologic markers for identifying patients with TDI-induced asthma among exposed workers.

Topics: Adult; Asthma; Autoantibodies; Biomarkers; Enzyme-Linked Immunosorbent Assay; Female; Humans; Immunoblotting; Keratin-18; Keratin-19; Keratin-8; Keratins; Male; Middle Aged; Occupational Diseases; Sensitivity and Specificity; Toluene 2,4-Diisocyanate

To explore the immunological mechanism of exfoliative tongue fur in children with asthma.. Thirty-nine children with asthma, twenty-eight children with repetitive respiratory tract infection (non-asthma) and eleven healthy children were divided into five groups, which were asthma with exfoliative fur or with non-exfoliative fur groups, non-asthma with exfoliative fur or with non-exfoliative fur groups and normal control group. The concentrations of keratin 13 and bcl-2 in cells exfoliated from tongue fur were detected by immunohistochemical method. The expression levels of blood cell chemokine receptor-3 (CCR-3) and CD4(+) were examined by flow cytometry, and the levels of serum cortisol and IgE were detected by radioimmunoassay.. The levels of blood CD4(+) and CCR-3 of children with asthma and exfoliative fur were higher than those in the asthma with non-exfoliative fur group and the normal control group (P<0.05). The serum level of cortisol in the groups of asthma with exfoliative fur and non-asthma with exfoliative fur were lower than that in the other groups (P<0.05). The serum levels of IgE in asthma with exfoliative fur or with non-exfoliative fur groups were higher than that in the other groups (P<0.05). Concentrations of keratin 13 in the cells exfoliated from tongue fur in the groups of asthma or non-asthma with exfoliative fur were lower than that of the other groups (P<0.05). There was no significant difference of expression level of bcl-2 in the cells exfoliated from tongue fur among these five groups.. There is a reasonably close relationship between the formation of exfoliative tongue fur and the immune system such as low level of serum cortisol and high levels of blood CD4(+) and CCR-3, which may all promote the formation of exfoliative fur. The disability of keratinization and apoptosis of epithelial cells of tongue may also be one cause for its formation.

Topics: Asthma; CD4 Antigens; Child; Child, Preschool; Diagnosis, Differential; Female; Glossitis, Benign Migratory; Humans; Hydrocortisone; Infant; Keratin-13; Keratins; Male; Medicine, Chinese Traditional; Receptors, CCR3; Receptors, Chemokine; Tongue

Inhaled isocyanate binds with cytokeratin (CK) of the epithelial cells, which could induce immune responses.. To elucidate the possible existence of an isocyanate-induced, asthma-associated autoantigen from the bronchial epithelial cells, which may be associated with toluene diisocyanate (TDI)-induced asthma development.. We cultured bronchial epithelial cells with incubation of TDI-human serum albumin (HSA) conjugate. Gene expression profiles of cultured epithelial cells were analyzed using a microarray technique. CK19 protein expression within the epithelial cells was confirmed by IgG immunoblot using monoclonal antibody to CK19. Serum IgG to CK19 and specific IgG and IgE antibodies to TDI-HSA conjugate were detected by enzyme-linked immunosorbent assay in 68 TDI asthma patients (group 1) and compared with 40 allergic asthma patients (group 2) and 80 unexposed healthy controls (group 3).. After TDI exposure, increased expression of CK19 and CK14 genes from the culture bronchial epithelial cells was noted using microarray analysis. IgG immunoblot analysis confirmed increased expression of CK19 after the TDI exposure. The levels of serum IgG to CK19 were significantly higher in the TDI asthma group than in groups 2 and 3 (P=.008). The prevalence of IgG to CK19 was significantly higher in group 1 (38.2%) than group 2 (22.5%) or group 3 (1.3%) (P=.008). Significant associations were noted between IgG to CK19 and specific IgG to TDI-HSA conjugate and transglutaminase (P=.02) but not with specific IgE to TDI-HSA conjugate.. We suggest that TDI exposure can augment CK19 expression from the bronchial epithelial cell, which may involve immune responses as an autoantigen to induce airway inflammation in TDI-induced asthma.

Topics: Adult; Allergens; Antibody Specificity; Asthma; Autoantibodies; Autoantigens; Bronchi; Bronchial Provocation Tests; Cell Line, Transformed; Cells, Cultured; Coculture Techniques; Enzyme-Linked Immunosorbent Assay; Epithelial Cells; Female; Gene Expression Profiling; Humans; Immunoconjugates; Immunoglobulin E; Immunoglobulin G; Keratins; Male; Methacholine Chloride; Middle Aged; Oligonucleotide Array Sequence Analysis; RNA, Messenger; Serum Albumin; Toluene 2,4-Diisocyanate

The allergic response to common environmental agents (allergens) has been regarded as an important mechanism in the development of airway inflammation of patients with asthma. However, allergic sensitization cannot be detected in a significant number of adult patients with asthma. The etiologic mechanism responsible for nonallergic asthma has not yet been identified. The idea of a possible involvement of autoimmunity in the pathogenesis of nonallergic asthma has been proposed by earlier studies. To test for the possible presence of an autoimmune response to bronchial epithelial cell antigens in nonallergic asthma, we examined circulating autoantibodies to cultured human bronchial epithelial cells (BEAS-2B) in sera from patients with nonallergic asthma by immunoblot analysis. IgG autoantibodies to the 49-kD bronchial epithelial cell antigen were detected in 10 of 23 patients with nonallergic asthma (43%), 3 of 27 patients with allergic asthma (11%), 2 of 20 patients with systemic lupus erythematosus (10%), and 3 of 34 healthy volunteers (9%) (p < 0.005). The 49-kD auto-antigen was purified and identified as cytokeratin 18 by amino acid sequencing. In this study, we identified cytokeratin 18 as a bronchial epithelial autoantigen associated with nonallergic asthma. Further studies are needed to determine the significance of autoimmunity in nonallergic asthma.

Topics: Adult; Analysis of Variance; Asthma; Autoantigens; Biomarkers; Bronchi; Case-Control Studies; Cells, Cultured; Chi-Square Distribution; Cohort Studies; Epithelial Cells; Female; Humans; Immunoblotting; Keratins; Male; Middle Aged; Probability; Reference Values; Sensitivity and Specificity; Severity of Illness Index

Induced sputum from asthmatic patients has been recently used to assess inflammatory cells. We have previously reported an increased expression of Th-2-type cytokines in induced sputum of asthmatic patients. C-C chemokines, particularly eotaxin and monocyte chemotactic protein (MCP)-4, are associated with eosinophilic infiltration. Interleukin (IL)-16 is associated with chemotactic activity for CD4+ cells. Chemokine expression in BAL and bronchial biopsy specimens has been demonstrated in asthmatic airways, but not in induced sputum.. We examined whether eotaxin, MCP-4, and IL-16 expression could be detected in induced sputum of asthmatic patients (n = 10), and whether the expression was increased compared to normal control subjects (n = 9). Eotaxin, MCP-4, and IL-16 immunoreactivity were determined by immunocytochemistry. In addition, inflammatory cells were investigated using markers for T cells (CD3), eosinophils (major basic protein [MBP]), macrophages (CD68), neutrophils (elastase), and epithelial cells (cytokeratin).. Our results showed that there was a significant difference in the percentages of MBP-positive and epithelial cells between asthmatic patients and normal control subjects (p < 0.05). However, there was no difference between these two groups in the percentage of CD3-, elastase-, and CD68-positive cells. Immunoreactivity for eotaxin, MCP-4, and IL-16 was expressed in the induced sputum of all asthmatic patients, and expression of these chemotactic cytokines was significantly greater than in control subjects (p < 0.001, p < 0.005, and p < 0.001, respectively).. This study showed that induced sputum could be used to detect chemokines in patients with bronchial asthma, and that the upregulation of chemotactic cytokines in the airways can be seen using noninvasive techniques.

Topics: Adult; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Asthma; CD3 Complex; Chemokine CCL11; Chemokines; Chemokines, CC; Cytokines; Eosinophils; Epithelial Cells; Female; Humans; Immunohistochemistry; Interleukin-16; Keratins; Macrophages; Male; Middle Aged; Monocyte Chemoattractant Proteins; Neutrophils; Pancreatic Elastase; Sputum; T-Lymphocytes