G. G. Martinovich

Intracellular redox state: towards quantitative description

Redox state is a widely used term for the description of redox phenomena in biological systems. The regulating mechanisms responsible for maintaining the redox state are not yet fully known. But it was shown that changes in the redox state might lead to a cascade of intracellular events, beneficial or deleterious to the cell. There are several methods for the description of the intracellular redox state. These methods are based on using measured intracellular concentrations of reduced and oxidized glutathione in the Nernst equation. However, glutathione is not always a basic redox component in biological fluids, organelles, cells or tissues. As a result, changes in the intracellular redox state are not always accompanied by considerable changes of glutathione concentration. In this work it was proposed to use the concept of effective reduction potential for the quantitative characteristic of the intracellular redox state. The effective reduction potential was substantiated on the basis of a thermodynamic description. A new equation for the calculation of the effective reduction potential was derived. This equation summarizes the contribution of different oxidizing and reducing agents in the formation of an effective redox potential. The theoretical estimation of the effective reduction potential values for the different biological fluids and cells was carried out with the use of a method developed.

Redox Regulation of Calcium Signaling in Cancer Cells by Ascorbic Acid Involving the Mitochondrial Electron Transport Chain

Previously, we have reported that ascorbic acid regulates calcium signaling in human larynx carcinoma HEp-2 cells. To evaluate the precise mechanism of Ca2+ release by ascorbic acid, the effects of specific inhibitors of the electron transport chain components on mitochondrial reactive oxygen species (ROS) production and Ca2+ mobilization in HEp-2 cells were investigated. It was revealed that the mitochondrial complex III inhibitor (antimycin A) amplifies ascorbate-induced Ca2+ release from intracellular stores. The mitochondrial complex I inhibitor (rotenone) decreases Ca2+ release from intracellular stores in HEp-2 cells caused by ascorbic acid and antimycin A. In the presence of rotenone, antimycin A stimulates ROS production by mitochondria. Ascorbate-induced Ca2+ release in HEp-2 cells is shown to be unaffected by catalase. The results obtained suggest that Ca2+ release in HEp-2 cells caused by ascorbic acid is associated with induced mitochondrial ROS production. The data obtained are in line with the concept of redox signaling that explains oxidant action by compartmentalization of ROS production and oxidant targets.

Plant Phenols and Autophagy.

Many plant phenols (stilbenes, curcumins, catechins, flavonoids, etc.) are effective antioxidants and protect cells during oxidative stress. Extensive clinical studies on the potential of phenolic compounds for treatment of cardiovascular, neurodegenerative, oncological, and inflammatory diseases are now being conducted. In addition to direct antioxidant effect, plant phenols may provide a protective effect via activation of the Keap1/Nrf2/ARE redox-sensitive signaling system and regulation of autophagy. In this review, mechanisms of effects of the most common plant phenols on autophagy are presented.

Mazes of Nrf2 regulation

Nrf2 transcription factor plays a key role in maintaining cellular redox balance under stress and is a perspective target for oxidative stress-associated diseases. Under normal conditions, Nrf2 transcriptional activity is low due to its rapid ubiquitination and degradation in the 26S proteasome, as well as through various modifications of amino acid residues of this transcription factor that regulate its transport to the nucleus and binding to DNA. Continuous activation of Nrf2 is possible due to autophagy and epigenetic regulation that may underlie the increased resistance of tumor cells to radiotherapy and chemotherapy. This review deals with the mechanisms of regulation of Nrf2 transcriptional activity and its main elements, and pharmacological approaches to activation of the Keap1/Nrf2/ARE system.

Redox regulation of cellular processes: A biophysical model and experiment

A model for the redox regulation of the functional state of the cell has been constructed on the basis of representation of electron transfer processes by equivalent electric circuits. The mechanism of action of redox-active molecules on biosystems has been discussed in terms of circuit theory. A method for determining the parameters of cellular redox sensors has been proposed. It has been established that the concentration and redox potential of compounds entering the cell are the main regulatory parameters of redox signals for the cell. It has been experimentally shown that the calcium response to hydrogen peroxide in rat C6 glioma cells and human FL amnion cells depends on the redox-buffer capacity of cells.

Phenolic antioxidant TS-13 regulating ARE-driven genes induces tumor cell death by a mitochondria-dependent pathway

Effects of water-soluble phenolic antioxidant sodium 3-(3′-tert-butyl-4′-hydroxyphenyl)-propyl thiosulfonate (TS-13), potassium 3,5-dimethyl-4-hydroxybenzyl thioethanoate (BEP-11-K), and potassium 3-(3′,5′-di-tert-butyl-4′-hydroxyphenyl)-propionate (potassium phenosan) on the proliferative activity of tumor cells and the role of redox-dependent and calcium-dependent signaling mechanisms in realization of tumor cell response to antioxidants were studied. Potassium phenosan and BEP-11-K were found to stimulate proliferation, whereas the ARE-inducing phenolic antioxidant TS-13 inhibited tumor cell growth in culture. The rate of tumor cell growth depended on the rate of intracellular reactive oxygen species production and was suppressed by apocynin (a NADPH oxidase inhibitor) and antimycin A (an ubiquinol-cytochrome c oxidoreductase inhibitor). The action of TS-13 on tumor cells was accompanied by a transient increase in the intracellular production of reactive oxygen species and the intracellular calcium concentration, whereas cell incubation with potassium phenosan and BEP-11-K did not influence the level of reactive oxygen species and intracellular calcium ions. Cyclosporin A blocked the inhibitory effect of TS-13. Thus, it can be reasonably speculated that phenolic antioxidant TS-13 triggers mitochondria-dependent apoptosis in tumor cells by opening mitochondrial permeability transition pores.

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

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 that quinones enhanced the intracellular production of reactive oxygen species, reduced the mitochondrial membrane potential, and induced tumor-cell death through different pathways. It was found that thymoquinone, which induced lower production of reactive oxygen species than 1,4-benzoquinone, was more toxic to tumor cells. It was established that reactive oxygen species produced due to exposure to thymoquinone are involved in redox signaling processes that lead to the formation of mitochondrial permeability transition pores and activation of programmed cell death. These results suggest that the functioning of the established redox signaling mechanism is enabled by the colocalization of mitochondrial oxidoreductases involved in the production of reactive oxygen species and of protein targets of reactive oxygen species involved in the activation of apoptosis.

Effects of Ascorbic Acid on Calcium Signaling in Tumor Cells

Effects of ascorbic acid on calcium homeostasis of human laryngeal carcinoma cells were studied. Intracellular concentration of free calcium and intracellular pH were measured by fluorescent analysis. Ascorbic acid in concentrations of 3–10 mM caused pH drop and sharply increased concentrations of free Ca ions in HEp-2 cells. Intracellular concentration of free Ca ions resulted from Ca ion release from the thapsigargin-sensitive Ca depots.

REGULATION OF THE PROLIFERATIVE ACTIVITY AND CHEMORESISTANCE IN TUMOR CELLS BY SODIUM ASCORBATE

The effect of ascorbate in physiological concentrations on the proliferative activity and chemoresistance in human larynx carcinoma HEp-2 cells was studied. Ascorbate in a concentration of 60 μM was found to increase the cancer cells proliferation rate 1.5 times. Ascorbate changes the functional state of the cancer cells, thereby increasing their resistance to doxorubicin and thymoquinone. It was shown that apocynin (NADPH oxidase inhibitor) blocks the stimulating effect of the antioxidant. The results obtained suggest that reactive oxygen species produced by NADPH oxidase participate in the mechanism of cell adaptive response induced by ascorbate.

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.