Hidefumi Miyagawa

Effect of cetirizine on human eosinophil superoxide generation, eosinophil chemotaxis and eosinophil peroxidase in vitro

Cetirizine, a potent H1-antagonist, has been reported to inhibit eosinophil migration into human skin. We, therefore, further evaluated the effect of cetirizine on eosinophil function, including superoxide anion generation, chemotaxis, and eosinophil peroxidase (EP) release. In allergic subjects, superoxide anion generation 60 min after platelet-activating factor (PAF) activation was inhibited by concentrations of cetirizine ranging from 0.01 to 1 microgram/ml (2.612 x 10(-8) to 2.612 x 10(-6) M). No significant inhibition was observed in normal subjects. PAF (10(-6) M)-induced eosinophil chemotaxis was also inhibited by cetirizine. In allergic subjects, percent inhibitions were 47.5 +/- 6.1% at 0.01 microgram/ml, 50.8 +/- 5.1% at 0.1 microgram/ml and 58.9 +/- 6.4% at 1 microgram/ml of cetirizine. In allergic subjects, N-formyl-methionyl-lencyl-phenylalanine induced eosinophil chemotaxis was inhibited by cetirizine, although EP release was not. These results suggest cetirizine has effects on eosinophils which can not be explained by H1-blockade alone.

The effect of ketotifen on eosinophils as measured at LTC4 release and by chemotaxis.

We examined the effect of ketotifen on calcium ionophore (A23187)-induced leukotriene C4 (LTC4) release from eosinophils, and platelet activating factor (PAF)-induced eosinophil chemotaxis to estimate the mechanism of the antiasthmatic property of ketotifen. Purified eosinophil suspensions were obtained from eight atopic asthmatic subjects and eight normals. Twenty microM of ketotifen significantly inhibited A23187-induced LTC4 release from eosinophils, and 10 microM of ketotifen significantly inhibited PAF-induced eosinophil chemotaxis. These findings may demonstrate the antiallergic pharmacological properties of ketotifen.

Density distribution and density conversion of neutrophils in allergic subjects

We investigated the density distribution of neutrophils in peripheral blood of allergic subjects. We divided neutrophils into four groups: fraction 1 (density greater than 1.085), fraction 2 (1.081 less than density less than or equal to 1.085), fraction 3 (1.077 less than density less than or equal to 1.081) and fraction 4 (density less than or equal to 1.077). The percentage of neutrophils in fraction 2 in allergic rhinitis (AR) subjects or asthmatics was lower than that in normals (p less than 0.01). The percentage of neutrophils in fraction 3 and fraction 4 from AR or asthmatics was greater than that in normals (fraction 3, p less than 0.01; fraction 4, p less than 0.05). In neutrophils from AR subjects (fraction 3), chemotaxis to N-formyl-methionyl-leucyl-phenylalanine or platelet-activating factor (PAF) was enhanced compared to fraction 2. PAF (10(-7) M) changed the density of neutrophils (p less than 0.01), which were inhibited by WEB 2086 (p less than 0.05). Furthermore, granulocyte-macrophage colony-stimulating factor changed the density of neutrophils (p less than 0.01). These findings suggest that biological agents may activate neutrophils and convert their density resulting in neutrophils with lower density in allergic subjects.

The effect of WEB 2086 on PAF-induced eosinophil chemotaxis and LTC4 production from eosinophils

We investigated the effect of WEB 2086, a selective platelet-activating factor (PAF) receptor antagonist, on PAF-induced eosinophil chemotaxis and LTC4 production. WEB 2086 inhibited PAF-induced eosinophil chemotaxis in normals and asthmatics. To further determine if WEB 2086 is a selective PAF receptor antagonist, we examined the effect of WEB 2086 against formyl-methionyl-leucyl-phenylalanine (fMLP)-induced or eosinophil chemotactic factor of anaphylaxis (ECF-A)-induced eosinophil chemotaxis. WEB 2086 did not have a significant inhibition against fMLP or ECF-A-induced eosinophil chemotaxis. These results suggest that WEB 2086 is a selective and potent inhibitor of PAF-induced eosinophil chemotaxis and LTC4 production from eosinophils, due to its antagonism of PAF-receptors.

Increased Hypodense Eosinophils After Activation with PAF-Acether and Calcium Ionophore in Asthmatic Subjects

Eosinophils play a major role in the pathogenesis of bronchial asthma. In this study, we examined the density characteristics of blood eosinophils from 9 normal healthy individuals and 9 allergic asthmatic patients. Furthermore, the effect of platelet-activating factor, a potent mediator of inflammation, and calcium ionophore, A23187, on the density of normodense eosinophils (density > 1.085 g/ml) has also been examined. Initially, asthmatic patients had 27.0 +/- 1.1% eosinophils of lighter density (density < or = 1.081 g/ml), significantly greater than that in the normal individuals (7.5 +/- 0.5%). After exposure to platelet-activating factor (1 microM) or calcium ionophore (A23187, 1 microgram/ml), the normodense eosinophils switched to hypodense in both groups: 16.7 +/- 2.1% and 54.2 +/- 3.7%, respectively, in normal individuals, and 30.6 +/- 5.7% and 77.4 +/- 2.3%, respectively, in asthmatic patients. These data demonstrated that a certain percentage of normodense eosinophils from asthmatics and normal subjects switched to hypodense after activation with platelet-activating factor or calcium ionophore. Furthermore, eosinophils from asthmatics switched to a greater degree than in normal subjects, suggesting that the normodense eosinophils in asthmatics become primed probably by endogenously released mediators.

Elevation of Neutrophil Chemotactic Activity in the Human Serum and the Decreased Number of Neutrophils after Platelet-Activating Factor Inhalation

We previously reported a significant decrease in neutrophils at 5 min and an increased number of neutrophils at 15 min after PAF-acether inhalation. To investigate the mechanism, we measured the neutrophil chemotactic activity of the sera. Neutrophil chemotactic activity (NCA) of the serum at 5 min was greater than at baseline (p less than 0.01) or at 15 min (p less than 0.05). Furthermore, we investigated the correlation between the change in NCA and decreased number of neutrophils. A significant correlation was found between these two at 5 min (r = 0.751, p less than 0.05). These data suggest that an elevation of NCA may play an important role in the decrease of circulating neutrophils after PAF-acether inhalation.

Effect of Terfenadine on Neutrophil and Eosinophil Chemotactic Activities after Inhalation of Platelet-Activating Factor in vivo and on Neutrophil Chemotaxis in vitro

In this double-blind crossover study we evaluated the effect of terfenadine on the rise in neutrophil chemotactic activity (NCA) and eosinophil chemotactic activity (ECA) in serum induced by platelet-activating factor (PAF) inhalation in 8 asthmatics. Additionally, we examined the direct effect of terfenadine on neutrophil chemotaxis in vitro in 7 allergic subjects. NCA and ECA in serum after PAF inhalation and neutrophil chemotaxis were measured using a modified Boyden chamber method. An initial elevation of NCA after PAF inhalation was inhibited by terfenadine, but the effect was diminished after subsequent PAF inhalations. Terfenadine showed no effect on ECA. In vitro PAF- and fMLP-induced neutrophil chemotaxis were significantly inhibited by terfenadine. These results suggest that terfenadine may have antiallergic properties in addition to its H1 receptor blockade.

Heterogeneity in chemotaxis of neutrophils with different densities.

We investigated N-formyl-Methionyl-Leucyl-Phenylalanine (fMLP)-induced chemotaxis of neutrophils with different densities. FMLP-induced (10(-8)M) chemotaxis of neutrophils with lower density were significantly reduced when compared to neutrophils with higher density (p less than 0.05). These findings imply a relationship between the neutrophil density and chemotaxis.

Fibronectin levels in plasma after platelet-activating factor inhalation

We evaluated fibronectin levels in plasma and blood leukocyte counts after platelet-activating factor-acether (PAF) inhalation to examine whether PAF may cause pulmonary vascular injury. PAF inhalation challenges were performed on 5 normal subjects and 5 atopic asthmatics. Fibronectin levels in plasma were significantly increased at 5 min (342.5 +/- 36.9 micrograms/ml) after PAF challenge as compared with the baseline values of 277.1 +/- 33.0 micrograms/ml (p less than 0.024). Total white blood cell and neutrophil counts were significantly decreased 5 min after PAF inhalation (WBC from 5,360 +/- 460 to 4,260 +/- 380/mm3, p less than 0.028; neutrophils from 3,113 +/- 441 to 2,214 +/- 344/mm3, p less than 0.043). Our study shows that plasma fibronectin levels transiently increase and that peripheral total WBC counts decrease after PAF inhalation. Elevated fibronectin levels may provide evidence of pulmonary vascular injury due to neutrophils which might be primed and recruited to pulmonary circulation by PAF.

The effect of a selective α2-adrenergic receptor antagonist (midaglizole) on the cyclic 3′,5′-adenosine monophosphate production in human mononuclear cells☆

Abstract A selective α 2 -adrenergic antagonist, midaglizole, has been recently reported to have efficacy in patients with asthma. To understand the mechanisms, we investigated the effect of midaglizole on the cyclic 3′,5′-adenosine monophosphate (cAMP) production in human mononuclear cells (MNCs). MNCs were separated by a histopaque gradient from 10 normal subjects and 10 subjects with asthma. cAMP was measured by RIA kits. Midaglizole (10 μmol/L) significantly enhanced isoproterenol-induced cAMP production in both groups, although midaglizole (from 1 to 100 μmol/L) did not increase the cAMP production by itself. The percent increase in cAMP was more in subjects with asthma (183.8%) than in normal subjects (140.4%); however, the absolute increase was not different. Platelet-activating factor has been demonstrated to inhibit β-agonist—induced cAMP production in several mammalian tissues, including human MNCs. Midaglizole also prevented platelet-activating factor—induced desensitization of the cAMP response to isoproterenol in MNCs. These findings suggest that midaglizole may be a useful additional agent for the therapy of bronchial asthma through an enhancement of the cAMP production.