It has been demonstrated that pterostilbene inhibits reactive oxygen species production in neutrophilsin vitro. However, little is known about its effects on neutrophils during inflammationin vivo. In this study, the effect of pterostilbene on neutrophil activity was investigated in experimental arthritis model. Lewis rats were injected by a single intradermal injection of heat-killedMycobacterium butyricumin Freund’s adjuvant to develop arthritis. Another group of arthritic animals received pterostilbene 30 mg/kg, daily, p.o. The number and activity of neutrophils in blood were measured on a weekly basis during the whole experiment. Moreover, the total radical trapping potential in plasma was measured at the end of the experiment. In the pterostilbene treated arthritic group, the treatment significantly lowered the number of neutrophils in blood on days 14 and 21 without significant downregulation of neutrophil oxidative burst. Pterostilbene nonsignificantly increased total radical trapping potential in arthritic animals. These results indicate that the promising effects of pterostilbene on reactive oxygen species operate by different mechanismsin vitroand in the animal model of inflammation. In conclusion, the positive effects of pterostilbene in the model of arthritis may be attributed to regulation of neutrophil number.

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.

Neutrophils are able to release cytotoxic substances and inflammatory mediators, which, along with their delayed apoptosis, have a potential to maintain permanent inflammation. Therefore, treatment of diseases associated with chronic inflammation should be focused on neutrophils; formation of reactive oxygen species and apoptosis of these cells represent two promising targets for pharmacological intervention. Piceatannol, a naturally occurring stilbenoid, has the ability to reduce the toxic action of neutrophils. This substance decreased the amount of oxidants produced by neutrophils both extra- and intracellularly. Radicals formed within neutrophils (fulfilling a regulatory role) were reduced to a lesser extent than extracellular oxidants, potentially dangerous for host tissues. Moreover, piceatannol did not affect the phosphorylation of p40^phox—a component of NADPH oxidase, responsible for the assembly of functional oxidase in intracellular (granular) membranes. The stilbenoid tested elevated the percentage of early apoptotic neutrophils, inhibited the activity of protein kinase C (PKC)—the main regulatory enzyme in neutrophils, and reduced phosphorylation of PKC isoformsα,βII, andδon their catalytic region. The results indicated that piceatannol may be useful as a complementary medicine in states associated with persisting neutrophil activation and with oxidative damage of tissues.

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.

Nitro-oleic acid (NO₂-OA), pluripotent cell-signaling mediator, was recently described as a modulator of the signal transducer and activator of transcription 3 (STAT3) activity. In our study, we discovered new aspects of NO₂-OA involvement in the regulation of stem cell pluripotency and differentiation. Murine embryonic stem cells (mESC) or mESC-derived embryoid bodies (EBs) were exposed to NO₂-OA or oleic acid (OA) for selected time periods. Our results showed that NO₂-OA but not OA caused the loss of pluripotency of mESC cultivated in leukemia inhibitory factor (LIF) rich medium via the decrease of pluripotency markers (NANOG, sex-determining region Y-box 1 transcription factor (SOX2), and octamer-binding transcription factor 4 (OCT4)). The effects of NO₂-OA on mESC correlated with reduced phosphorylation of STAT3. Subsequent differentiation led to an increase of the ectodermal marker orthodenticle homolog 2 (Otx2). Similarly, treatment of mESC-derived EBs by NO₂-OA resulted in the up-regulation of both neural markers Nestin and β-Tubulin class III (Tubb3). Interestingly, the expression of cardiac-specific genes and beating of EBs were significantly decreased. In conclusion, NO₂-OA is able to modulate pluripotency of mESC via the regulation of STAT3 phosphorylation. Further, it attenuates cardiac differentiation on the one hand, and on the other hand, it directs mESC into neural fate.