Protective and adaptogenic role of peroxiredoxin 2 (Prx2) in neutralization of oxidative stress induced by ionizing radiation.

A radioprotective effect of exogenous recombinant peroxiredoxin 2 (Prx2) was revealed and characterized using an animal model of whole body X-ray irradiation at sublethal and lethal doses. Prx2 belongs to an evolutionarily ancient family of peroxidases that are involved in enzymatic degradation of a wide variety of organic and inorganic hydroperoxides. Apart from that, the oxidized form of Prx2 also exhibits chaperone activity, thereby preventing protein misfolding and aggregation under oxidative stress. Intravenous administration of Prx2 in animals at a concentration of 20 µg/g 15 min before exposure to ionizing radiation contributes to a significantly higher survival rate, suppresses the development of leucopenia and thrombocytopenia, as well as protects the bone marrow cells from genome DNA damage. Moreover, injection of Prx2 leads to suppression of apoptosis, stimulates cell proliferation and results in a more rapid recovery of the cell redox state. Exogenous Prx2 neutralizes the effect of the priming dose on the second irradiation of the cells. The radioprotective properties of exogenous Prx2 are stipulated by its broad substrate peroxidase activity, chaperone activity in the oxidized state, and are also due to the signal-regulatory function of Prx2 mediated by the regulation of the level of hydroperoxides as well as via interaction with redox-sensitive regulatory proteins.

The role of peroxiredoxin 6 in neutralization of X-ray mediated oxidative stress: effects on gene expression, preservation of radiosensitive tissues and postradiation survival of animals.

Peroxiredoxins are redox-sensing multifunctional enzymes, among them peroxiredoxin 6 (Prx6) can neutralize the most broadest range of hydroperoxides and play an important role in maintaining the redox homeostasis of the cell. In the present study, radioprotective and signaling regulatory effects of Prx6 were demonstrated and characterized. Intravenously administered exogenous Prx6 protects the organism of mice from the destructive action of ionizing radiation in the lethal dose range of 5-10 Gy. Dose reduction factor of 1.4 Prx6 injection reduces the severity of radiation-induced leuko- and thrombopenia in irradiated animals, also preventing the destruction of epithelial cells in the small intestine. Injecting exogenous Prx6 also as its mutated form of Prx6-C47S lacking peroxidase activity affects the expression of genes involved in antioxidant response, DNA reparation, apoptosis and inflammatory processes, in bone marrow cells both in intact animals and in those subjected to ionizing radiation. The radioprotective properties of Prx6 are based, on the one hand, on the capability for ROS neutralization, and on the other hand - on the potentiality for activation of reparation processes of the cell under oxidative stress conditions. Prx6 can be considered as a potentially perspective radioprotective agent for the reduction of risks from the damaging action of ionizing radiation on the mammalian organism.

Exogenous 8-Oxo-7,8-dihydro-2'-deoxyguanosine: Biomedical Properties, Mechanisms of Action, and Therapeutic Potential.

8-Oxo-7,8-dihydroguanine (8-oxo-G) is a key biomarker of oxidative damage to DNA in cells, and its genotoxicity is well-studied. In recent years, it has been confirmed experimentally that free 8-oxo-G and molecules containing it are not merely inert products of DNA repair or degradation, but they are actively involved in intracellular signaling. In this review, data are systematized indicating that free 8-oxo-G and oxidized (containing 8-oxo-G) extracellular DNA function in the body as mediators of stress signaling and initiate inflammatory and immune responses to maintain homeostasis under the action of external pathogens, whereas exogenous 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dGuo) exhibits pronounced antiinflammatory and antioxidant properties. This review describes known action mechanisms of oxidized guanine and 8-oxo-G-containing molecules. Prospects for their use as a therapeutic target are considered, as well as a pharmaceutical agent for treatment of a wide range of diseases whose pathogenesis is significantly contributed to by inflammation and oxidative stress.

Peroxiredoxin 6 is a natural radioprotector.

After injection of 20 mg/kg peroxiredoxin 6 to male Kv:SHK mice 15 min before X-ray irradiation in the range of lethal doses (7-10 Gy), the mice remained alive for 30 days, whereas the mortality of the control animals was 100%. In the irradiated animals, peroxiredoxin 6 decreased the severity of radiation-induced leucopenia, granulocytopenia, and thrombocytopenia, increased the number of blood corpuscles, and prevented the mass death of epithelial cells and the destruction of the small intestine. Thus, peroxiredoxin 6 can be regarded as a prophylactic radioprotective agent.

[Continuous Generation of Hydrogen Peroxide in Water Containing Very Low Concentrations of Unsymmetrical Dimethylhydrazine].

Continuous generation of hydrogen peroxide catalyzed by low concentrations of 1,1-dimethylhydrazine (heptyl)--a rocket fuel component--in air saturated water was shown by the method of enhanced chemiluminescence in the system of luminol-p-iodophenol-peroxidase. The concentration dependence and the influence of heat and light on the formation of hydrogen peroxide in the water under the influence of dimethylhydrazine at concentrations considerably lower than maximum allowable concentrations were studied, and the physical-chemical mechanism of this process was considered. It is supposed that dimethylhydrazine at ultra-low concentrations is associated with air nanobubbles and represents a long-lived complex performing catalysis of hydrogen peroxide formation under the influence of heat and light. We put forward the new concept of.toxicity of dimethylhydrazine at very low concentrations due to violation of homeostasis of reactive oxygen species formation in aqueous solutions entering the body of humans and animals.

[Radioprotectors: History, Trends and Prospects].

The search for ideal protective agents for use in a variety of radiation scenarios has continued for more than six decades. This review describes the history of the major discoveries, shows the chronology of the changes in attitudes, trends and paradigms. The readers are invited to meet with various classes of chemical compounds that have the potential to protect against acute and late effects of ionizing radiation when administered either before or after radiation exposure. The work represents characteristics of radioprotective agents such as a dose reduction factor, time of administration, tissue specificity, toxicity; the mechanisms of their action and practical applications are also described. A separate chapter considers the further development prospects and directions in this field of research.

[Impact of biologically important anions on reactive oxygen species formation in water under the effect of non-ionizing physical agents].

The influence of biologically relevant anions (succinate, acetate, citrate, chloride, bicarbonate, hydroorthophosphate, dihydroorthophosphate, nitrite, nitrate) on the formation of hydrogen peroxide and hydroxyl radicals in water was studied under the effect of non-ionizing radiation: heat, laser light with a wavelength of 632.8 nm, corresponding to the maximum absorption of molecular oxygen, and electromagnetic radiation of extremely high frequencies. It has been established that various anions may both inhibit the formation of reactive oxygen species and increase it. Bicarbonate and sulfate anions included in the biological fluids' and medicinal mineral waters have significant, but opposite effects on reactive oxygen species production. Different molecular mechanisms of reactive oxygen species formation are considered under the action of the investigated physical factors involving these anions, which may influence the biological processes by signal-regulatory manner and provide a healing effect in physical therapy.

Pro-oxidative, genotoxic and cytotoxic properties of uranyl ions.

It is demonstrated that hydroxyl radicals and hydrogen peroxide are formed under the action of uranyl ions in aqueous solutions containing no reducing agents. In the presence of uranyl ions, formation of 8-oxoguanine in DNA and long-lived protein radicals are observed in vitro. It is shown that the pro-oxidant properties of uranyl at micromolar concentrations mostly result from the physico-chemical nature of the compound rather than its radioactive decay. Uranyl ions lead to damage in DNA and proteins causing death of HEp-2 cells by necrotic pathway. It is revealed that the uranyl ions enhance radiation-induced oxidative stress and significantly increase a death rate of mice exposed to sublethal doses of X-rays.

[Natural purine compounds as radioprotective agents].

Purine compounds xanthosine, caffeine, inosine-5'-monophosphate and guanosine-5'-monophosphate in the concentration range of 0.02-1 mmol/L exhibit antioxidant properties in vitro, significantly reducing the formation of hydrogen peroxide and hydroxyl radicals induced by X-rays in aqueous solutions and preventing the formation of 8-oxoguanine in DNA solutions. These compounds neutralize the long-lived protein radicals in vitro induced by radiation. In vivo they exhibit pronounced radiotherapeutic properties, increasing the survival rate of mice up to 50% by intraperitoneal injection (45 mg/kg) after the exposure to a lethal dose of 7 Gy. The tested compounds stimulate hemopoiesis, increasing the number of white blood cells and platelets in the peripheral blood of animals in postradiation period, as well as radiation recovery of DNA damage when administered both before and after irradiation. These purine compounds can be considered as potentially promising preventive and therapeutic agents to reduce the risk of the pathological effects of ionizing radiation on the body of mammals.

[Generation of reactive oxygen species in water under exposure of visible or infrared irradiation at absorption band of molecular oxygen].

It is found that in bidistilled water saturated with oxygen hydrogen peroxide and hydroxyl radicals are formed under the influence of visible and infrared radiation in the absorption bands of molecular oxygen. Formation of reactive oxygen species (ROS) occurs under the influence of both solar and artificial light sourses, including the coherent laser irradiation. The oxygen effect, i.e. the impact of dissolved oxygen concentration on production of hydrogen peroxide induced by light, is detected. It is shown that the visible and infrared radiation in the absorption bands of molecular oxygen leads to the formation of 8-oxoguanine in DNA in vitro. Physicochemical mechanisms of ROS formation in water when exposed to visible and infrared light are studied, and the involvement of singlet oxygen and superoxide anion radicals in this process is shown.

[Long-lived radicals of amino acids induced by X-ray-radiation are the source of hydrogen peroxide in aqueous medium].

The formation of long-lived radicals in the solutions of casein and its hydrolysate with an equimolar mixture of amino acids was compared by measuring the X-ray-induced chemiluminescence. It was shown that free amino acids constituting the protein produce long-lived radicals. It was demonstrated that some amino acids (Leu, Ile, Val, Ser, Trp, Met, Pro, Arg, Gly, Phe) emit light of visible spectrum over a long period of time after the irradiation, which indicates the generation of long-lived radicals of these amino acids. The half-life times of these radicals are several hours. Dissolving irradiated dry amino acids capable of luminescing over a long time gives rise to the formation of hydrogen peroxide in aqueous medium.

[Formation of hydrogen peroxide and hydroxyl radicals in aqueous solutions of L-amino acids by the action of X-rays and heat].

The action of 1 mM solutions of L-amino acids in 5 mM phosphate buffer, pH 7.4, on the production of hydrogen peroxide and hydroxyl radicals under the action of X-rays and heating has been studied. Hydrogen peroxide was estimated by the method of enhanced luminescence in a system luminol-paraiodophenol-peroxidase and hydroxyl radicals were determined by using the fluorescence probe coumarin-3-carboxylic acid. It was shown that amino acids can be divided by their influence on H202 formation into three groups: those that reduce the yield of H202, that do not influence it, and that increase it. A similar action of amino acids was observed upon heating, but the composition of the groups was different. All amino acids lowered the formation of hydroxyl radicals under the action of X-rays, and the most effective among them were Cys > His > Phe = Met = Trp > Tyr. Met, His and Phe lowered the amount of hydroxyl radicals by heating, Ser raised it, whereas Tyr and Pro did not change it. Thus, amino acids differently influence the formation of reactive oxygen species by the action of X-rays and heat, and some of amino acids reveal themselves as effective natural antioxidants.

Effect of amino acids on X-ray-induced hydrogen peroxide and hydroxyl radical formation in water and 8-oxoguanine in DNA.

Generation of hydrogen peroxide and hydroxyl radicals in L-amino acid solutions in phosphate buffer, pH 7.4, under X-ray irradiation was determined by enhanced chemiluminescence in the luminol-p-iodophenol-peroxidase system and using the fluorescent probe coumarin-3-carboxylic acid, respectively. Amino acids are divided into three groups according to their effect on the hydrogen peroxide formation under irradiation: those decreasing yield of H2O2, having no effect, and increasing its yield. All studied amino acids at 1 mM concentration decrease the yield of hydroxyl radicals in solution under X-ray irradiation. However, the highest effect is observed in the order: Cys > His > Phe = Met = Trp > Tyr. At Cys, Tyr, and His concentrations close to physiological, the yield of hydroxyl radicals decreases significantly. Immunoenzyme analysis using monoclonal antibodies to 8-oxoguanine (8-oxo-7,8-dihydroguanine) was applied to study the effect of amino acids with the most pronounced antioxidant properties (Cys, Met, Tyr, Trp, Phe, His, Lys, Arg, Pro) on 8-oxoguanine formation in vitro under X-ray irradiation. It is shown that amino acids decrease the content of 8-oxoguanine in DNA. These amino acids within DNA-binding proteins may protect intracellular DNA against oxidative damage caused by formation of reactive oxygen species in conditions of moderate oxidative stress.

[Oxygen effect in heat-mediated damage to DNA].

The effects of gassing conditions in DNA solution on the major types of heat-mediated DNA damage (depurination of DNA, generation of 8-oxoguanine, cytosine deamination with the formation of uracil) have been studied by ELISA, column liquid chromatography, and spectrophotometry. It was found that the number of DNA lesions depends on oxygen concentration in solution; i.e., the oxygen effect takes place. The heat-induced generation of hydrogen peroxide in water increased after the addition of D20 and decreased by the action of various 1O2 quenchers, suggesting that singlet oxygen is involved in the heat-induced production of reactive oxygen species (ROS) in water. The data obtained favor the hypothesis that all the types of heat-induced damage to DNA are due to a common mechanism associated with the heat-mediated generation of reactive oxygen species in solution.

[Guanosine and inosine as natural geneprotectors for mice blood cells exposed to X-rays].

The radioprotective effects of guanosine and of inosine on bone marrow cells of mice exposed to acute X-rays (1.5 Gy) were studied by using the micronuclear test. The guanosine and inosine (riboxine) decrease the frequency of micronucleated polychromatic erythrocytes and significantly recover erythropoiesis. Also, radioprotective effects of the guanosine and of the inosine on the irradiated leucocytes of mice were tested by the alkaline comet assay. Was shown that purine ribonucleosides diminish quantity of DNA damage and activates repair processes in leucocytes under irradiation of blood and animals. The reactive oxygen species induced by ionizing radiation perform essential role in DNA damaging. Using a sensitive method of enhanced chemiluminescence in a peroxidase-luminol-p-iodophenol system for quantitative measurement of hydrogen peroxide and coumarin-3-carboxylic acid for quantitative measurement of hydroxyl radicals we have shown that guanosine and inosine essentially decrease the yield of hydrogen peroxide and hydroxyl radicals in X-ray-irradiated water. The results obtained indicate that radioprotective properties of guanosine and inosine (riboxine) in the blood cells are operative at the genome level.