Antihistamines of the H 1 and H 3 /H 4 groups interfere with oxidative burst of human professional phagocytes in vitro. In the concentration of 10 μM, H 1 antihistamines of the 1 st and 2 nd generation inhibited oxidative burst of human neutrophils in the rank order of potency: dithiaden > loratadine > brompheniramine > chlorpheniramine > pheniramine. Of the H 1 antihistamines, the most effective was dithiaden in suppressing oxidative burst of whole human blood and dosedependently the chemiluminescence of isolated neutrophils at extra-and intracellular level. Inhibition of free oxygen radical generation in isolated neutrophils by dithiaden resulted from the inhibition of protein kinase C activation. The potentiation of recombinant caspase-3 by dithiaden is supportive of the antiinflammatory effect of dithiaden and suggestive of increasing the apoptosis of professional phagocytes. Of the H 3 /H 4 antihistamines, the most effective was JNJ7777120 in decreasing chemiluminescence in whole blood and also at extra-and intracellular sites of isolated neutrophils. JNJ 10191584 and thioperamide were less effective and the latter significantly potentiated free oxygen radical generation intracellularly. The results demonstrated that, compared with the H 3 /H 4 antihistamines investigated, H 1 antihistamines were much more potent in inhibiting free oxygen radical generation in human professional phagocytes. This finding should be taken into account therapeutically.

The effect of three therapeutically used drugs and five polyphenolic compounds on the mechanism of oxidative burst was compared in whole blood and isolated neutrophils at cellular and molecular level. In 10 μM concentration, the compounds investigated decreased the oxidative burst of whole blood in the rank order of potency: N-feruloylserotonin (N-f-5HT) > curcumin (CUR) > quercetin (QUER) > arbutin (ARB) > resveratrol (RES) > dithiaden (DIT) > carvedilol (CARV) > brompheniramine (BPA). The ratio between the percentage inhibition of extracellular versus intracellular chemiluminescence (CL) followed the rank order QUER > N-f-5HT > RES > CUR > DIT and is indicative of the positive effect of the compounds tested against oxidative burst of neutrophils, demonstrating suppression of reactive oxygen species extracellularly with minimal alteration of intracellular reactive oxygen species (ROS). Activation of protein kinase C was significantly decreased by DIT, CUR, QUER and N-f-5HT. CARV, DIT, QUER and ARB reduced activated neutrophil myeloperoxidase release more significantly compared with the effect on superoxide anion generation. All compounds tested increased the activity of caspase-3 in cell-free system. It is suggested that other regulatory mechanisms than protein kinase C might participate in the inhibition of neutrophil activation with the compounds tested. Different mechanisms are concerned in controlling the assembly of NADPH oxidase and the regulatory role of calcium ions is suggested. Compounds decreasing the amount of extracellular ROS generation, yet affecting but minimally intracellular ROS generation, are promising for further investigation in vivo.

IntroductionH 1 -antihistamines, recently defined as inverse agonists or neutral antagonists, exert an antiinflammatory effect, which might result both from receptor interaction and non-receptormediated activities. It has been suggested that the physicochemical nature of H 1 -antihistamines, namely lipophilic molecules which carry a positive charge, allows them to associate with cell membranes and competitively inhibit the binding of second messengers, e. g. Ca 2+ , thereby reducing the activity of calcium-dependent enzymes [1]. In this study, we compared the effect of the H 1 -antihistamine Dithiaden ® (Dit) with histamine (His) on reactive oxygen metabolite (ROM) generation by human polymorphonuclear leukocytes (PMNL) under different mechanisms of stimulation. Materials and methodsThe oxidative burst of PMNL was measured in whole human blood by the luminol-enhanced chemiluminescence (CL) method in luminometer Immunotech LM-01T (Immunotech, Czech Republic [2]). Luminolenhanced CL of isolated PMNL was recorded in Lumi-aggregometer model 500 Harveston, USA [3,4]). Samples were treated with the appropriate stimuli in the final contentrations: N-formylmethionyl-leucyl-phenylalanine (fMLP) 0.1 mmol/L, calcium ionophore A23187 0.5 mmol/L, phorbol-12-myristate-13 acetate (PMA) 0.01 mmol/L, or opsonised zymosan (OZ) 0.1 g/L. Statistical significance of differences between the means was established by Student's ttest and p values below 0.05 were considered significant. Figure 1 demonstrates the effect of His and Dit on CL of whole blood stimulated with OZ. His showed a dose-dependent inhibitory effect on the CL at concentrations above 10 mmol/L. In contrast, Dit significantly decreased CL at concentrations from 10 nmol/L. In isolated PMNL, His decreased CL in the rank order of stimuli: fMLP > A23187 > OZ and had no effect on CL stimulated with PMA. Dit dose-dependently decreased CL in the rank order of stimuli PMA > OZ > A23187 and increased CL induced with fMLP only at a concentration of 10 mmol/L. Figure 2 (panel A) shows the effect of Dit and His on CL of PMNL stimulated with OZ. Dit but not His at a concentration of 10 mmol/L decreased OZ-stimulated CL. Simultaneous administration of Dit and His did not affect the inhibitory effect of Dit. In contrast, Dit (10 mmol/L) increased CL of isolated PMNL stimulated with fMLP ( Fig. 2, panel B). His in equimolar concentration significantly decreased fMLP-stimulated CL and, when applied simultaneously with Dit, it abolished the effect of the latter. Results and discussionIt seems likely that His does not play a regulatory role in aggregation of leukocytes and the inhibitory effect of Dit might be nonspecific [5]. In OZ-stimulated PMNL (both whole blood and isolated cells), His and Dit worked synergistically by a dose-dependent inhibition of CL, with Dit being much more potent. However, in fMLP-stimulated PMNL, His antagonised the potentiation of CL by Dit. The difference in the effect of His and Dit on CL stimulated with OZ and fMLP may be due to the different mechanisms of...