cytochalasin-b and Asthma

Topics: Anaphylaxis; Animals; Antigen-Antibody Reactions; Asthma; Basophils; Bronchi; Cromolyn Sodium; Cyclic AMP; Cytochalasin B; Cytotoxicity Tests, Immunologic; Diethylcarbamazine; Eosinophils; Histamine; Histamine Release; Humans; Immunization; Immunoglobulin E; Immunoglobulin G; In Vitro Techniques; Kinins; Leukocytes; Mast Cells; Mucous Membrane; Parasympathomimetics; Prostaglandins; Receptors, Adrenergic; Receptors, Cholinergic; SRS-A

We have investigated the exocytotic characteristics of neutrophils from allergic patients and healthy volunteers employing the whole cell membrane capacitance (Cm) measurement. The mean serum IgE level from allergic patients (423.75 +/- 12.75 IU/ml) determined by chemiluminescence immunoassay was much higher than that of healthy volunteers (28.47 +/- 16.68 IU/ml). Intracellular dialysis of buffered Ca2+ and GTPgammaS triggered biphasic exocytosis. The total capacitance increment displayed a steep dependence on pipette free Ca2+ concentration ([Ca2+]p), with maximal stimulation achieved at 10 microM. A significant decrease in the total capacitance increment was observed in the allergic group at [Ca2+]p >10 microM. Moreover, at submaximal stimulatory [Ca2+]p of 1 microM, the maximal rate of exocytosis in allergic patients (Vmax = 20.75 +/- 6.19 fF/s) was much faster than that of the healthy control group (Vmax = 7.97 +/- 2.49 fF/s). On the other hand, the Ca2+-independent exocytosis stimulated by GTPgammaS displayed no significant difference in either the total membrane capacitance increments or the maximal rate of exocytosis. The results suggest that hypersecretion of neutrophils in allergic diseases may involve the development of abnormal Ca2+-dependent exocytosis.

Topics: Adolescent; Adult; Antibody Specificity; Asthma; Biomarkers; Calcium; Chemokines; China; Cytochalasin B; Dose-Response Relationship, Drug; Exocytosis; Female; Guanosine 5'-O-(3-Thiotriphosphate); Humans; Immunoglobulin E; Male; Neutrophil Activation; Neutrophils; Receptors, Chemokine; Receptors, IgE; Rhinitis, Allergic, Perennial; Sensitivity and Specificity

Although proteinases are thought to contribute to the pathogenesis of bronchial asthma and COPD, the mechanism of proteinase release from inflammatory cells has not been thoroughly clarified. We examined matrix metalloproteinase (MMP-9) release from human leukocytes using soluble agonists such as C5a, FMLP, and PAF. Mononuclear cells, neutrophils, and eosinophils isolated from human leukocytes were incubated with C5a, FMLP, or PAF for 20 min. MMP-9 in supernatants was measured by ELISA. Among mononuclear cells, neutrophils, and eosinophils, MMP-9 was released mainly from neutrophils. FMLP was the most effective stimulus of MMP-9 release from neutrophils among three agonists: C5a, FMLP, and PAF. GM-CSF clearly enhanced FMLP-induced MMP-9 release. Pretreatment of neutrophils with pertussis toxin (PTX) resulted in the inhibition of FMLP-induced MMP-9 release, indicating the contribution of PTX-sensitive G-proteins to intracellular signal transduction in FMLP-induced MMP-9 release. These results suggest that neutrophils release large amounts of MMP-9 in response to FMLP, which is a bacterial product analogue. It cannot be excluded that MMP-9 released from neutrophils may be involved in the pathogenesis of bronchial asthma and COPD.

Topics: Asthma; Complement C5a; Cytochalasin B; Dose-Response Relationship, Drug; Eosinophils; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; In Vitro Techniques; Leukocytes; Leukocytes, Mononuclear; Matrix Metalloproteinase 9; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; Pertussis Toxin; Platelet Activating Factor; Pulmonary Disease, Chronic Obstructive

The decrease in prostaglandin E(2) (PGE(2)) release due to aspirin (ASA)-induced cyclooxygenase inhibition and the increment in cysteinyl leukotriene (Cys-LT) release secondary to the removal of the inhibitory effect of PGE(2) on Cys-LT release have been suggested in the pathogenesis of aspirin-induced asthma (AIA).. In this study, we aimed to investigate the in vitro release of Cys-LT and to determine the effect of PGE(2) on Cys-LT release from peripheral blood leucocytes of patients with AIA after stimulation by ASA.. Patients with AIA (n = 13), patients with ASA-tolerant asthma (ATA) (n = 12) and healthy volunteers as controls (n = 13) were included to the study. ASA and PGE2 at three different concentrations were applied to the peripheral blood leucocytes of the study group, and Cys-LT levels following stimulants were assessed by enzyme immunoassay method.. There was no difference in baseline Cys-LT levels between groups (AIA 353.4 +/- 55.5 pg/mL, ATA 354.7 +/- 40.3 pg/mL, and control group 368.5 +/- 30.2 pg/mL; P > 0.05). Though not present in other groups, the Cys-LT level of 453.6 +/- 70.0 pg/mL following ASA stimulation was higher than baseline in patients with AIA (P = 0.04). When PGE(2) was added to the ASA-stimulated samples of patients with AIA, Cys-LT levels were measured as 298.7 +/- 78.6 pg/mL, 279.8 +/- 79.9 pg/mL, and 243.4 +/- 51.3 pg/mL at PGE(2) 10(-7) m, 10(-6) m and 10(-5) m concentrations, respectively. These levels were lower than the ASA-stimulated Cys-LT values (P = 0.03, P = 0.01 and P = 0.01, respectively). The inhibitory effect of different PGE(2) concentrations on Cys-LT release was also present in patients with ATA and in controls.. The increase in Cys-LT levels following ASA stimulation seems to be unique to AIA, which was not present in patients with ATA and in healthy controls. The inhibitory effect of PGE(2) on stimulated Cys-LT levels is another important finding to elucidate the role of PGE(2) in the pathogenesis of AIA.

Topics: Adult; Aged; Anti-Inflammatory Agents, Non-Steroidal; Aspirin; Asthma; Cysteine; Cytochalasin B; Dinoprostone; Female; Humans; Leukocytes; Leukotrienes; Male; N-Formylmethionine Leucyl-Phenylalanine; Stimulation, Chemical

This study was performed to test the hypothesis that activated eosinophils or their secretory products can directly stimulate sensory neurons to release their neuropeptides. Neurons derived from neonatal rat dorsal root ganglia (DRG), which synthesize and store sensory neuropeptides, were placed in primary cell culture and were exposed to eosinophils or their bioactive mediators. The resultant release of substance P (SP) was measured by enzyme-linked immunosorbent assay and was expressed as a percent (mean +/- SE) of total neuronal SP content. Eosinophils were isolated from human volunteers with a history of allergic rhinitis and/or mild asthma and were activated by incubation with cytochalasin B (5 micrograms/ml) and N-formyl-methionyl-leucyl-phenylalanine (FMLP, 1 microM). Activated eosinophils [6 x 10(6)/ml, suspended in Hanks' buffered salt solution (HBSS)] applied to cultured DRG neurons for 30 min increased basal SP release 2.4-fold compared with HBSS-exposed neurons (activated eosinophils 11.10 +/- 2.48% vs. HBSS 4.59 +/- 0.99%; P = 0.002), whereas neither nonactivated eosinophils nor cytochalasin B and FMLP in HBSS influenced SP release. Additional cultured DRG neurons were exposed to soluble products made by eosinophils. Compared with SP release under control conditions (2.37 +/- 0.34%), major basic protein (MBP) increased release in a concentration-related fashion (e.g., 3 microM MBP: 6.23 +/- 0.67%, P = 0.006 vs. control), whereas neither eosinophil cationic protein (3 microM), eosinophil-derived neurotoxin (3 microM), leukotriene D4 (500 nM), platelet-activating factor (100 nM), nor H2O2 (100 microM) affected SP release. These studies demonstrate that activated eosinophils can stimulate cultured DRG neurons directly and suggest that MBP may be the responsible mediator.

Topics: Animals; Animals, Newborn; Asthma; Capsaicin; Cells, Cultured; Coculture Techniques; Cytochalasin B; Enzyme-Linked Immunosorbent Assay; Eosinophils; Ganglia, Spinal; Humans; Myelin Basic Protein; N-Formylmethionine Leucyl-Phenylalanine; Neurons; Platelet Activating Factor; Polylysine; Potassium Chloride; Rats; Rats, Sprague-Dawley; Rhinitis, Allergic, Seasonal; Substance P; Tissue Extracts

Histamine release induced by platelet-activating factor (PAF) from leukocytes of aspirin-sensitive subjects with asthma was higher than that from normal control subjects, despite the similarity of anti-IgE-induced histamine release. Moreover, basophils of some aspirin-sensitive subjects with asthma released histamine by PAF stimulation in the absence of cytochalasin B that affects histamine release and is required in PAF-induced histamine release from leukocytes of atopic subjects with asthma and normal control subjects. In addition to temperature dependency and inhibition by ethylenediaminetetraacetic acid reported previously, PAF-induced histamine release was enhanced by cytochalasin B and indomethacin and inhibited by dexamethasone. These features are common with IgE-mediated histamine release and suggest the existence of the common pathway to PAF-induced histamine release and IgE-mediated histamine release. The results in the present study indicate the pathophysiologic significance of PAF-induced histamine release and that activation of basophils by PAF may be relevant to the pathogenesis in some aspirin-sensitive subjects with asthma.

Topics: Adult; Aged; Aspirin; Asthma; Basophils; Cytochalasin B; Dexamethasone; Female; Histamine Release; Humans; Immunoglobulin E; Indomethacin; Male; Middle Aged; Platelet Activating Factor

Activation of neutrophils (PMN) within the airways results in the secretion of a number of products such as reduced oxygen metabolites that could contribute to the inflammatory response associated with asthma. However, mediators of allergy, such as histamine, prostaglandin E2 (PGE2), isoproterenol, and adenosine, may serve to mitigate this inflammation through feedback inhibition of neutrophil function. To test the hypothesis that PMN activation and feedback inhibition mechanisms may be abnormal in asthmatics, we compared both superoxide production and adenosine-induced suppression of superoxide production in 12 matched pairs of asthmatics and control subjects. PMN obtained from asthmatic patients generated significantly more superoxide in response to f-met-leu-phe (fMLP) than controls (2.94 +/- 55 nmol/5 x 10(5) PMN/5 min versus 1.38 +/- 0.35 at 2 x 10(-8) M fMLP and 3.81 +/- 0.68 nmol versus 2.04 +/- 0.45 nmol at 10(-7) M; p less than 0.01 for both). In contrast, the respiratory burst generated by two receptor-independent stimuli, the calcium ionophore A23187 and phorbol myristate acetate, was equivalent between control and asthmatic subjects. At 10(-6) M, 2-chloroadenosine induced a 19.5 +/- 5.1% inhibition of fMLP-stimulated superoxide production in PMN from patients with asthma as compared to 55.6 +/- 24.6% inhibition in PMN from control subjects (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 2-Chloroadenosine; Adenosine; Asthma; Calcimycin; Cytochalasin B; Dinoprostone; Dose-Response Relationship, Drug; Humans; Isoproterenol; Lipopolysaccharides; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; Superoxides

Platelet-activating factor (PAF) was found to induce histamine release from human basophils in mixed leukocytes in the presence of cytochalasin B. The reaction was rapid and dependent on temperature. The release was lower at a high concentration of PAF and was inhibited by EDTA, suggesting a noncytolytic mechanism. CV-3988, a PAF antagonist, inhibited the reaction dose-dependently. PAF acted as an IgE-independent stimulus, but PAF-induced histamine release from leukocytes of allergic asthmatics, who had elevated serum IgE levels, was significantly higher than that from leukocytes of controls. These results suggest that PAF-induced histamine release is useful for studying the role of PAF in the pathogenesis of allergic disorders including bronchial asthma.

Topics: Adult; Asthma; Basophils; Cytochalasin B; Edetic Acid; Histamine Release; Humans; Hypersensitivity, Immediate; Immunoglobulin E; Leukocytes; Phospholipid Ethers; Platelet Activating Factor; Stimulation, Chemical