Effect of concanavalin A on the level of sulfhydryl groups in thymocytes

Fluorescein mercury acetate (FMA), a fluorescent probe, is used for the investigation of SH-groups of thymocytes' plasma membrane. It is found that mitogenic lectin Con A decreases the amount of membrane SH-groups and increases the fluorescence polarization degree of FMA (PFMA). The value of PFMA increases also during the incubation of cells with potassium ferricyanide and H2O2 but it decreases in the presence of NADH. The analysis of the data permits a conclusion that the thymocyte activation by Con A results in the selective oxidation of certain SH-groups with the formation of disulphide cross-linking between the plasma membrane receptors bound with the lectin molecules.     

Thymoquinone,a biologically active component of Nigella sativa,induces mitochondrial production of reactive oxygen species and programmed death of tumor cells

Biophysics - Mechanisms of tumor-cell responses to 2-isopropyl-5-methyl-1,4-benzoquinone (thymoquinone) and 1,4-benzoquinone were studied using fluorescence and the inhibition assay. It was shown...     

Regulation of morphological and functional properties of astrocytes by hydrogen peroxide

Effects of hydrogen peroxide on morphological characteristics, proliferation index, and menadione-dependent lucigenin-enhanced chemiluminescence of C6 glioma cells were studied. It was established that H₂O₂ at 5 × 10^?7?1 × 10^?8 M concentrations acted as a regulator of morphological and functional properties of astrocytes, inducing their reactivation, which is manifested as cell body hypertrophy and an increase of proliferative activity and menadione-induced production of superoxide anion radicals (O ₂ ^?? ). Cytodestructive action of hydrogen peroxide at a concentration higher than 1 × 10^?6 M on C6 glioma cells shows itself as a decrease of their proliferation index and the ability to generate O ₂ ^?? under the effect of menadione. Use of lipopolysaccharide B as a functional stimulator has shown that H₂O₂ modifies signaling pathways leading to an increase of mitotic activity of C6 glioma cells and decreases the yield of lucigenin-dependent chemiluminescence of astrocytes under the action of menadione to the level of control values.     

Effect of hydrogen peroxide on ability of neutrophils to generate the reactive oxygen and chlorine species and secrete myeloperoxidase in vitro

The influence of H2O2 at concentrations of 10(-8)--10(-2) mol/l on neutrophil ability to generate the reactive oxygen and chlorine species (ROCS) and secrete myeloperoxidase (MPO) was studied, and H202 injurious effect on neutrophils was also investigated in this work. It was revealed that H2O2 at concentrations of 2 x 10(-3)--2 x 10(-2) mol/l induced disturbance of the neutrophil membrane barrier properties and lactate dehydrogenase release. The incubation of the neutrophils with the addition of 10(-4)--10(-7) mol/l H2O2 led to an increase in the cell ability to generate ROCS during phagocytosis and decreased neutrophil ability to secrete MPO and ROCS in extracellular medium during adhesion. The mechanisms of H2O2 effect are coupled with arachidonic acid metabolism. Inhibition of metabolic pathways of 5-lipoxygenase or cyclooxygenase increased the destructive effect of H2O2 on the cells. Five-lipoxygenase way prohibition led to cancellation of H2O2 influence on MPO and ROCS secretion and to enhancement of H2O2 effect on neutrophil ability to generate ROCS during phagocytosis. The data obtained testify to the high neutrophil resistance to destructive effect of H2O2 and confirm the regulatory role of H2O2 with respect to the neutrophil functions.     

Regulation of morphologicaland functional properties of astroglial cells by hydrogen peroxide

Effects of hydrogen peroxide on morphological characteristics, proliferation index, menadione-dependent lucigenin-enhanced chemiluminescence of C6 glioma cells were studied. It was established that H2O2 at 1 x 10(-8) - 5 x 10(-7) M concentrations acts as a regulator of morphological and functional properties of astrocytes by inducing their reactivation that is manifested as a cell body hypertrophy and an increase of proliferative activity and of menadione-dependent production of superoxide (O2- ). Cytodestructive action of hydrogen peroxide at a concentration higher than 1 microM on C6 glioma cells shows itself as a decrease of their proliferation index and the ability to generate O2- under menadione action. Using lipopolysaccharide B as a functional stimulator it has been shown that H2O2 modifies signaling pathways leading to the increase of mitotic activity of C6 glioma cells and decreases the yield of lucigenin-enhanced chemiluminescence of astrocytes under menadione action to the level of control values.     

The role of active forms of oxygen in platelet aggregation and deaggregation

The effect of hydrogen peroxide on ADP-induced platelet aggregation in the presence of active oxygen species scavengers was studied. It was shown that the superoxide radical and singlet oxygen, alongside with hydrogen peroxide, may play a role in platelet interactions.     

The priming effect of halogenated phospholipids on the functional responses of human neutrophils

Halogenated lipids formed in the reactions with myeloperoxidase (MPO)-derived species may contribute to the regulation of the functional activity of cells. In the present study we have investigated the effects of chloro- and bromohydrins formed in the HOCl and HOBr reactions, respectively, with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) on three different functional responses of human neutrophils: H₂O₂ generation, degranulation (MPO exocytosis), and cell aggregation. It was shown that POPC chloro- and bromohydrins (POPC-Cl and POPC-Br) induced the priming of neutrophils, resulting in significant upregulation of cell responses to neutrophil stimulators such as N-formyl-Met-Leu-Phe and lectin from Solanum tuberosum. The stimulating effects of POPC-Cl and POPC-Br were observed at low micromolar concentrations (liposomal concentration of POPC, 0.5–5 μM; the content of POPC-Cl or POPC-Br, 38 ± 3% of total lipids) after a short exposure (about 5 min) of the neutrophils to POPC-Cl or POPC-Br. These results suggest that halogenated lipids formed in vivo via MPO-dependent reactions may be considered as a new class of biologically active substances that are potentially able to contribute to the priming of myeloid cells in the sites of inflammation and serve as inflammatory response modulators.     

Peroxynitrite regulates exocytosis of neutrophil granules

Peroxynitrite is formed in the organism by activated neutrophils as a result of the enhanced production of nitrogen monoxide and superoxide anion radical in the inflammation foci. Since peroxynitrite modifies the structure of macromolecules, including the elements of actin cytoskeleton, it can influence signal transduction pathways that regulate intracellular granule exocytosis. In this paper we explore a dual effect of peroxynitrite on the processes of neutrophil degranulation by the methods of flow cytometry, light microscopy, and atomic force microscopy. We showed that peroxynitrite at concentrations less than 300 μM activated graded exocytosis of neutrophil intracellular granules, which resulted in the enhancement of neutrophil adhesion to the substrate, cell spreading on the substrate, and activation of neutrophil ability to kill microorganisms. Peroxynitrite at higher concentrations inhibited exocytosis of neutrophil granules and hindered cell adhesion to the substrate. The character of influence of the specific agents, such as colchicine and cytochalasin that selectively disrupt cytoskeletal structures, on peroxynitrite-induced changes in neutrophil morphology indicates an important role of actin cytoskeleton in the regulation of intracellular granule exocytosis induced by peroxynitrite. Our results support the hypothesis suggesting that peroxynitrite is a natural regulator of neutrophil effector functions.     

Mechanisms of Redox Regulation of Chemoresistance in Tumor Cells by Phenolic Antioxidants

Effects of water-soluble sulfur-containing phenolic antioxidants sodium 3-(3′-tert-butyl-4′- hydroxyphenyl)propyl thiosulfonate and potassium 3,5-dimethyl-4-hydroxybenzyl thioethanoate on chemoresistance in tumor cells have been studied. The studied phenolic antioxidants cause oppositely directed changes in the redox properties and chemoresistance in tumor cells. Potassium 3,5-dimethyl-4-hydroxybenzyl thioethanoate increases redox buffering capacity and doxorubicin resistance in tumor cells. Sodium 3-(3′- tert-butyl-4′-hydroxyphenyl)propyl thiosulfonate reduces the redox buffering capacity, which leads to a decrease in the chemoresistance of tumor cells. These observations suggest that one of the key mechanisms responsible for the formation of tumor cell resistance to antitumor compounds is the attenuation of apoptosis through increase of redox buffering capacity. The dependence of protein sensor redox state on oxidant concentrations and on redox buffering capacity in cells has been determined based on the proposed biophysical model of redox-dependent mechanism of apoptosis activation.     

Lectin-induced activation of plasma membrane NADPH oxidase in cholesterol-depleted human neutrophils

The gp91phox subunit of flavocytochrome b₅₅₈ is the catalytic core of the phagocyte plasma membrane NADPH oxidase. Its activation occurs within lipid rafts and requires translocation of four subunits to flavocytochrome b₅₅₈. gp91phox is the only glycosylated subunit of NADPH oxidase and no data exist about the structure or function of its glycans. Glycans, however, bind to lectins and this can stimulate NADPH oxidase activity. Given this information, we hypothesized that lectin–gp91phox interactions would facilitate the assembly of a functionally active NADPH oxidase in the absence of lipid rafts. To test this, we used lectins with different carbohydrate-binding specificity to examine the effects on H₂O₂ generation by human neutrophils treated with the lipid raft disrupting agent methyl-β-cyclodextrin (MβCD). MβCD treatment removed membrane cholesterol, caused changes in cell morphology, inhibited lectin-induced cell aggregation, and delayed lectin-induced assembly of the NADPH oxidase complex. More importantly, MβCD treatment either stimulated or inhibited H₂O₂ production in a lectin-dependent manner. Together, these results show selectivity in lectin binding to gp91phox, and provide evidence for the biochemical structures of the gp91phox glycans. Furthermore, the data also indicate that in the absence of lipid rafts, neutrophil NADPH oxidase activity can be altered by these select lectins.