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  • Author: Tomáš Perečko x
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Rami B. Kassab, Ondrej Vasicek, Milan Ciz, Antonin Lojek and Tomas Perecko

Abstract

The health benefits of berberine have been recognized for years. Even so, its effects on human neutrophils, the first line of immune defense, have not been reported. The purpose of this study was to investigate the effects of berberine on the human neutrophil oxidative burst. Reactive oxygen species production was analyzed by luminol-enhanced chemiluminescence. The analysis was performed in spontaneous and stimulated (phorbol myristate acetate (PMA) or opsonized zymosan particles (OZP)) whole blood and isolated neutrophils in the presence or absence of berberine. The effects of berberine on oxidant production in cell-free assays were evaluated using luminescence (H2O2-peroxidase-luminol) and fluorescence (Oxygen Radical Absorbance Capacity – ORAC) techniques. Berberine decreased the production of reactive oxygen species in human whole blood and isolated neutrophils stimulated with either PMA or OZP with a different efficiency (EC50 was 69 μM and 197 μM for PMA and OZP, respectively). The effect was more pronounced in isolated neutrophils. Cell-free assays showed the antioxidant activity of berberine against peroxyl radicals and hydrogen peroxide. Based on our results, we suggest that the effects of berberine on reactive oxygen species production in human neutrophils are due to its antioxidant activity.

Open access

Tomáš Perečko, Katarína Drábiková, Radomír Nosáľ, Juraj Harmatha and Viera Jančinová

Abstract

Chronic inflammatory diseases, e.g. rheumatoid arthritis or cystic fibrosis, are characterised by neutrophil infiltration in inflamed tissues. Dysregulated neutrophil death may contribute to the pathogenesis of diseases where neutrophils play a role. Stilbene derivatives are reported to activate apoptosis in different cell lines. Neutrophils from healthy volunteers were incubated in vitro with resveratrol, pterostilbene, pinosylvin or piceatannol (1-100 μmol/l), and cytotoxicity and apoptosis were measured by luminometry and flow cytometry, respectively. Enhancement and/or inhibition of human recombinant caspase-3 enzyme activity were measured by luminometry. None of the stilbene derivatives tested increased ATP liberation from human neutrophils, thus showing no direct cytotoxicity effect. Resveratrol and piceatannol (100 μmol/l) treated neutrophils had a higher rate of apoptosis compared to non-treated cells. Pterostilbene and pinosylvin (1 μmol/l), yet not resveratrol or piceatannol, increased the activity of caspase-3. However in the concentration of 100 μmol/l, all stilbene derivatives tested inhibited caspase-3 activity. Their effects on human neutrophil apoptosis differed according to the structure of the molecule. Additional studies are required to get insight into the mechanisms involved in the effects of the substances tested on neutrophil viability.

Open access

Katarína Drábiková, Tomáš Perečko, Radomír Nosáľ, Juraj Harmatha, Jan Šmidrkal and Viera Jančinová

Abstract

The study provides new information on the effect of natural polyphenols (derivatives of stilbene - resveratrol, pterostilbene, pinosylvin and piceatannol and derivatives of ferulic acid - curcumin, N-feruloylserotonin) on the activity of human neutrophils in influencing oxidative burst. All the polyphenols tested were found to reduce markedly the production of reactive oxygen species released by human neutrophils on extra-and intracellular levels as well as in cell free system. Moreover, pinosylvin, curcumin, N-feruloylserotonin and resveratrol decreased protein kinase C activity involved in neutrophil signalling and reactive oxygen species production. Our results suggest that due to their anti-neutrophil activity, the polyphenols tested might be attractive candidates in therapeutic development.

Open access

Viera Jančinová, Tomáš Perečko, Juraj Harmatha, Radomír Nosáľ and Katarína Drábiková

Abstract

Prolonged or excessive formation and liberation of cytotoxic substances from neutrophils intensifies inflammation and the risk of tissue damage. From this perspective, administration of substances which are able to reduce activity of neutrophils and to enhance apoptosis of these cells may improve the therapy of pathological states connected with persistent inflammation. In this short review, neutrophil oxidative burst and apoptosis are presented as potential targets for pharmacological intervention. Effects of natural polyphenols (resveratrol, pterostilbene, pinosylvin, piceatannol, curcumin, N-feruloylserotonin) are summarised, considering the ability of these compounds to affect inflammation and particularly neutrophil activity. The intended neutrophil inhibition is introduced as a part of a new strategy for pharmacological modulation of chronic inflammatory processes, focused on supporting innate antiinflammatory mechanisms and enhancing resolution of inflammation.

Open access

Viera Jančinová, Tomáš Perečko, Radomir Nosáľ, Danica Mihalová, Katarína Bauerová and Katarína Drabiková

Pharmacological regulation of neutrophil activity and apoptosis

Novel strategies of antiinflammatory therapy are based upon pharmacological agents capable to enhance the resolution - i.e. the termination of the beneficial inflammation before it may turn into an adverse chronic stage. In contrast to the current therapy, which antagonises the formation of proinflammatory mediators, the "proresolving" therapy promotes natural antiinflammatory processes. It is likely that several drugs and phytochemicals would act in this way, but this point has not been investigated and thus might be totally overlooked. In this paper, effects of curcumin (diferuloylmethane) were analysed, considering the ability of this natural compound to affect resolution of inflammation through modulation of its important inputs - activity and apoptosis of neutrophils. The presented data indicate that, besides its well-known ability to suppress mechanisms engaged at the onset and progression of inflammation, curcumin could support resolution of inflammation through decreased activity and enhanced apoptosis of neutrophils. This substance decreased the formation of oxidants in neutrophils, both under in vitro conditions and after oral administration to arthritic rats. Moreover, curcumin accelerated spontaneous apoptosis of neutrophils, as indicated by increased externalisation of phosphatidylserine, by intercalation of propidium iodide and by enhanced activity of the executioner caspase-3.

Open access

Rado Nosáľ, Katarína Drábiková, Viera Jančinová, Tatiana Mačičková, Jana Pečivová, Tomáš Perečko and Juraj Harmatha

Abstract

In this study we investigated the effect of five therapeutically used drugs and four natural polyphenolic compounds on the mechanism of oxidative burst of human neutrophils concerning their participation in the generation of reactive oxygen species (ROS). The compounds investigated decreased the oxidative burst of whole blood in the rank order of potency: N-feruloylserotonin > quercetin > curcumin > arbutin > dithiaden > carvedilol. The generation of intracellular reactive oxygen species in isolated neutrophils decreased in the same rank order, while carvedilol was ineffective. Scavenging of extracellular oxygen radicals followed the rank order of potency: N-feruloylserotonin > curcumin > quercetin > dithiaden. Arbutin and carvedilol had no effect. All compounds tested increased the activity of caspase-3 in cell-free system indicating a positive effect on apoptosis of neutrophils. Activation of protein kinase C was significantly decreased by dithiaden, curcumin, quercetin and N-feruloylserotonin. Carvedilol, dithiaden, quercetin and arbutin reduced activated neutrophil myeloperoxidase release more significantly compared with their less pronounced effect on superoxide generation The presented results are indicative of pharmacological intervention with neutrophils in pathological processes. Of particular interest was the effect of natural compounds. Intracellular inhibition of oxidative burst in isolated neutrophils by the drugs tested and natural antioxidants has to be further analysed since ROS play an important role in immunological responses of neutrophils.

Open access

Antonin Lojek, Milan Číž, Michaela Pekarová, Gabriela Ambrožová, Ondřej Vašíček, Jana Moravcová, Lukáš Kubala, Katarína Drábiková, Viera Jančinová, Tomáš Perečko, Jana Pečivová, Tatiana Mačičková and Radomír Nosáľ

Modulation of metabolic activity of phagocytes by antihistamines

The purpose of the study was to investigate the effects of H1-antihistamines of the 1st generation (antazoline, bromadryl, brompheniramine, dithiaden, cyclizine, chlorcyclizine, chlorpheniramine, clemastine) and the 2nd generation (acrivastine, ketotifen, and loratadine) on the respiratory burst of phagocytes. Reactive oxygen species generation in neutrophils isolated from rat blood was measured using luminol-enhanced chemiluminescence. Changes in nitrite formation and iNOS protein expression by RAW 264.7 macrophages were analysed using Griess reaction and Western blotting. The antioxidative properties of drugs in cell-free systems were detected spectrophotometrically, luminometrically, fluorimetrically, and amperometrically. The majority of the H1-antihistamines tested (bromadryl, brompheniramine, chlorcyclizine, chlorpheniramine, clemastine, dithiaden, and ketotifen) exhibited a significant inhibitory effect on the chemiluminescence activity of phagocytes. H1-antihistamines did not show significant scavenging properties against superoxide anion and hydroxyl radical, thus this could not contribute to the inhibition of chemiluminescence. H1-antihistamines had a different ability to modulate nitric oxide production by LPS-stimulated macrophages. Bromadryl, clemastine, and dithiaden were the most effective since they inhibited iNOS expression, which was followed by a significant reduction in nitrite levels. H1-antihistamines had no scavenging activity against nitric oxide. It can be concluded that the effects observed in the H1-antihistamines tested are not mediated exclusively via H1-receptor pathway or by direct antioxidative properties. Based on our results, antihistamines not interfering with the microbicidal mechanisms of leukocytes (antazoline, acrivastine and cyclizine) could be used preferentially in infections. Other antihistamines should be used, under pathological conditions accompanied by the overproduction of reactive oxygen species.