Ionizing Radiation Alters the Properties of Sodium Channels in Rat Brain Synaptosomes

The effect of ionizing radiation on neuronal membrane function was assessed by measurement of neurotoxin-stimulated 22Na+ uptake by rat brain synaptosomes. High-energy electrons and gamma photons were equally effective in reducing the maximal uptake of 22Na+ with no significant change in the affinity of veratridine for its binding site in the channel. Ionizing radiation reduced the veratridine-stimulated uptake at the earliest times measured (3 and 5 s), when the rate of uptake was greatest. Batrachotoxin-stimulated 22Na+ uptake was less sensitive to inhibition by radiation. The binding of [3H]saxitoxin to its receptor in the sodium channel was unaffected by exposure to ionizing radiation. The effect of ionizing radiation on the lipid order of rat brain synaptic plasma membranes was measured by the fluorescence polarization of the molecular probes 1,6-diphenyl-1,3,5-hexatriene and 1-[4-(trimethylammonium)phenyl]-6-phenyl-1,3,5-hexatriene. A dose of radiation that reduced the veratridine-stimulated uptake of 22Na+ had no effect on the fluorescence polarization of either probe. These results demonstrate an inhibitory effect of ionizing radiation on the voltage-sensitive sodium channels in rat brain synaptosomes. This effect of radiation is not dependent on changes in the order of membrane lipids.     

Effect of ionizing radiation on the functional state of Helix pomatia neurons. The systems of Na+ and K+ transport

The influence of ionizing radiation (5.16 C/kg) on passive and active Na+ and K+ transfer within the giant neurons of edible snail (Helix pomatia) has been investigated. It has been shown that ionizing radiation increases passive permeability of membranes, inhibits active ion transport, changes the number and the affinity of functionally active Na+,K+-ATPase molecules. The authors discuss the mechanisms of action of ionizing radiation.     

The effect of ionizing radiation on the regulation of the Na,K-ATPase activity of the kidney by univalent cations

The effect of ionizing radiation of 0.206 C/kg on the kinetics of activation of rat kidney Na,K-ATPase preparation by Na and K ions was studied as an index of possible qualitative and quantitative changes in the properties of the enzyme. Ionizing radiation was shown not only to increase the enzyme activity but also to change the optimal rate of ATP hydrolysis by Na,K-ATPase and to induce some differences in the shape of the curve for Na,K-ATPase dependence upon Na-sodium//potassium ion ratio in the incubation medium.     

The effect of low-level x-irradiation on the key enzymes of the active ion transport system in the cell

A study was made of the effect of ionizing radiation (0.013 C/kg) on Na, K- and Ca, Mg-ATPase activity in membranes of rat organs differing in radiosensitivity. It was shown that radiation mainly caused activation of enzymes that was most pronounced in brain membranes.     

Lethal and mutagenic effects of radiation and alkylating agents on two strains of mouse L5178Y cells

In previous studies, the two closely related strains of L5178Y (LY) mouse lymphoma cells, LY-R and LY-S, have been shown to differ in their sensitivity to UV and ionizing radiation. Thus, in comparison to strain LY-R, strain LY-S has been found to be more sensitive to the lethal effects of ionizing radiation, more resistant to the lethal effects of UV radiation, but less mutable at the hypoxanthine-guanine phosphoribosyl transferase (HGPRT) locus by both UV and X-radiation. In the present work, the lethal and mutagenic effects of ethyl methanesulfonate (EMS), methyl nitrosourea (MNU) and UV radiation (254 nm) were compared in the two strains. Mutability at the Na+/K+-ATPase locus as well as the HGPRT locus was determined. As previously reported, we found strain LY-S to be more resistant than strain LY-R to the lethal effects of UV radiation. In contrast, strain LY-S was more sensitive to the cytotoxic effects of the two alkylating agents. In spite of these differences in sensitivity, we found strain LY-S to be less mutable than strain LY-R by all 3 agents at the HGPRT locus. At the Na+/K+-ATPase locus, strain LY-S was also less mutable than strain LY-R by equal concentrations of EMS and UV radiation and by equitoxic concentrations of MNU. However, the difference between the strains was much more pronounced at the HGPRT locus than at the Na+/K+-ATPase locus. We have suggested that the interaction of unrepaired lesions in strain LY-S tends to cause an excess of deletions and multilocus effects, which in turn result in a locus-dependent decrease in the recovery of viable LY-S mutant cells.     

The Inhibitory Effects of Low-Dose Ionizing Radiation in IgE-Mediated Allergic Responses

Ionizing radiation has different biological effects according to dose and dose rate. In particular, the biological effect of low-dose radiation is unclear. Low-dose whole-body gamma irradiation activates immune responses in several ways. However, the effects and mechanism of low-dose radiation on allergic responses remain poorly understood. Previously, we reported that low-dose ionizing radiation inhibits mediator release in IgE-mediated RBL-2H3 mast cell activation. In this study, to have any physiological relevance, we investigated whether low-dose radiation inhibits allergic responses in activated human mast cells (HMC-1(5C6) and LAD2 cells), mouse models of passive cutaneous anaphylaxis and the late-phase cutaneous response. High-dose radiation induced cell death, but low-dose ionizing radiation of <0.5 Gy did not induce mast cell death. Low-dose ionizing radiation that did not induce cell death significantly suppressed mediator release from human mast cells (HMC-1(5C6) and LAD2 cells) that were activated by antigen-antibody reaction. To determine the inhibitory mechanism of mediator released by low-dose ionizing radiation, we examined the phosphorylation of intracellular signaling molecules such as Lyn, Syk, phospholipase Cγ, and protein kinase C, as well as the intracellular free Ca²⁺ concentration ([Ca²⁺]_i). The phosphorylation of signaling molecules and [Ca²⁺]_i following stimulation of FcεRI receptors was inhibited by low dose ionizing radiation. In agreement with its in vitro effect, ionizing radiation also significantly inhibited inflammatory cells infiltration, cytokine mRNA expression (TNF-α, IL-4, IL-13), and symptoms of passive cutaneous anaphylaxis reaction and the late-phase cutaneous response in anti-dinitrophenyl IgE-sensitized mice. These results indicate that ionizing radiation inhibits both mast cell-mediated immediate- and delayed-type allergic reactions in vivo and in vitro.     

Post-radiation changes in active transport in he CNS: Mg2+, Ca2+-ATPase

The effect of ionizing radiation of lethal (0.31 C/kg) and superlethal (9.288 C/kg) doses on Mg2+, Ca2+-ATPase activity in plasma membranes of brain cortex and cerebellum has been studied. As is shown, ionizing radiation has an inhibitory effect on the enzyme activity which is most pronounced and irreversible after exposure to superlethal doses.     

Novel ionizing radiation-sensitive mutants of Deinococcus radiodurans.

Two new loci, irrB and irrI, have been identified in Deinococcus radiodurans. Inactivation of either locus results in a partial loss of resistance to ionizing radiation. The magnitude of this loss is locus specific and differentially affected by inactivation of the uvrA gene product. An irrB uvrA double mutant is more sensitive to ionizing radiation than is an irrB mutant. In contrast, the irrI uvrA double mutant and the irrI mutant are equally sensitive to ionizing radiation. The irrB and irrI mutations also reduce D. radiodurans resistance to UV radiation, this effect being most pronounced in uvrA+ backgrounds. Subclones of each gene have been isolated, and the loci have been mapped relative to each other. The irrB and irrI genes are separated by approximately 20 kb of intervening sequence that encodes the uvrA and pol genes.     

The action of ionizing radiation on neuronal function in the edible snail. Phospholipase A2 and the Na+ and K+ transport systems

A study was made of the influence of A2 phospholipase on 22Na release from cells of nerve ganglia of edible snail. The treatment of nerve ganglia with A2 phospholipase inhibits Na, K-pump of neuronal membranes and does not exert a substantial effects on Na/Ca metabolism. There is a similarity between the effects of ionizing radiation and A2 phospholipase on the release of 22Na from cells.     

Postradiation changes in the system of active transport of ions in the CNS. Analysis of the cation centers of Na,K-ATPase

A study was made of the influence of ionizing radiation of 0.31 C/kg on the kinetic parameters showing the activity of brain Na, K-ATPase preparation to be a function of ion-regulator concentration. The use of the new method for the analysis of the enzyme cation centers permitted to estimate that whole-body irradiation of rats with the above dose did not cause in vitro a substantial change in the pattern of Na, K-ATPase activation by Na and K ions.     

Mechanism of radiation injury in the ion transport systems of nervous tissue

The biological effect of ionizing radiation (IR) in lethal and sublethal doses on the sodium-potassium transport systems in the fractions, enriched of neuron and glial cells and in cortex slices from rat brain was investigated. It was shown that IR leads to marked disturbances in the activity of Na,K-ATPase both in neuron and in glial cells. Some phasic character of alterations may be noted, which is expressed in different degree for various cellular elements of the brain. Using the surviving brain slices we have shown that IR causes essential phasic changes in potassium ion reaccumulation in different times after exposure. The mechanisms of the disturbance of Na,K-pump function in nervous tissue after irradiation are under discussion.     

Cidofovir administered with radiation displays an antiangiogenic effect mediated by E6 inhibition and subsequent TP53-dependent VEGF repression in HPV18+ cell lines

Therapeutic administration of the antiviral agent cidofovir with radiation markedly enhanced the antitumor effect of ionizing radiation in cells of two HPV18+ human cervical carcinoma cell lines. Although this potent radiosensitizing effect was associated with repression of the viral oncoproteins E6/ E7 and restoration of TP53 as shown previously, additional mechanisms may be involved. In the present study, we investigated the antiangiogenic effect of the combination of cidofovir and radiation in cells of two HPV18+ cervical cancer cell lines, HeLa and ME180, and assessed the molecular mechanisms associated with the antiangiogenic effect observed. Cells were exposed to cidofovir (10 microg/ml) and irradiated (1-9 Gy). The angiogenic response was studied in vitro by a matrigel invasion assay. Modulations of E6, TP53 and VEGF mRNA and protein levels were studied by real-time RT-PCR, Western blot analysis and ELISA, respectively. Then a double RNA interference approach was used to analyze the connection between E6/TP53 and VEGF. The combination of cidofovir and radiation had a potent antiangiogenic effect. It induced E6 inhibition, restoration of TP53, and reduction of the proangiogenic phenotype of HPV18+ cells associated with VEGF inhibition. A siRNA strategy showed an anti-VEGF action of the combination mediated directly by E6 inhibition and TP53 restoration, since E6 siRNA inhibited VEGF whereas co-transfection with E6 and TP53 siRNA abrogated the anti-VEGF effect. This study showed that the combination of cidofovir with ionizing radiation has an antiangiogenic effect associated with VEGF inhibition subsequent to E6 inhibition and TP53 restoration.     

The modifying effect of the frequency of pulses of ionizing radiation on its efficiency in biological experiments

In biological experiments the ionizing radiation is usually defined by the type, energy and total absorbed dose, as well as by the dose rate. If radiation generated by a physical particle accelerator is employed, we consider it necessary to supplement these data with the value for frequency of pulses of the ionizing radiation jet. As follows from our experiments, this parameter expressively affects the relative biological effect of ionizing radiation.     

The effect of alpha-phenyl-N-t-butylnitrone on ionizing radiation-induced apoptosis in U937 cells

Ionizing radiation induces the production of reactive oxygen species (ROS), which play an important causative role in apoptotic cell death. alpha-Phenyl-N-t-butylnitrone (PBN) is one of the most widely used spin-trapping compounds for investigating the existence of free radicals in biological systems. We investigated the effects of PBN on ionizing radiation-induced apoptosis in U937 cells. Upon exposure to 2 Gy of gamma-irradiation, there was a distinct difference between the control cells and the cells pre-treated with 2 mM PBN for 2 h in regard to apoptotic parameters, cellular redox status, mitochondria function and oxidative damage to cells. PBN effectively suppressed morphological evidence of apoptosis and DNA fragmentation in U937 cells exposed to ionizing radiation. The [GSSG]/[GSH+GSSG] ratio and the generation of intracellular ROS were higher and the [NADPH]/[NADP+ +NADPH] ratio was lower in control cells compared to PBN-treated cells. The ionizing radiation-induced mitochondrial damage reflected by the altered mitochondrial permeability transition, the increase in the accumulation of ROS, and the reduction of ATP production were significantly higher in control cells compared to PBN-treated cells. PBN pre-treated cells showed significant inhibition of apoptotic features such as activation of caspase-3, up-regulation of Bax and p53, and down-regulation of Bcl-2 compared to control cells upon exposure to ionizing radiation. This study indicates that PBN may play an important role in regulating the apoptosis induced by ionizing radiation presumably through scavenging of ROS.     

Influence of an alternating magnetic field on the bactericidal effect of ionizing radiation

A study was made of the effect of alternating magnetic field (AMF) and ionizing radiation delivered separately or in a combination on the microorganisms differing in radio-resistance. AMF (240 and 750 E) had no pronounced bactericidal action. A synergistic increase in the sterilizing effect of ionizing radiation was demonstrated after incubation of irradiated bacteria in AMF. The radiation-magnetic technique is proposed for sterilization of preparations and articles made of non-thermoresistant materials which permits to decrease by 1.5 times the bactericidal dose of ionizing radiation.     

Effect of the superoxide anion (O2-) on Na+-dependent amino acid transport

Superoxide anion (O2-) generated either by the autoxidation of dihydroxyfumaric acid (DHF) or enzymatically by the xanthine-xanthine oxidase system inhibited the uptake of 2-aminoisobutyric acid (AIB) in thymocytes. The transport of this non-metabolizable amino acid in thymocytes is mediated by a Na+-dependent mechanism. Inhibition of this transport system by O2- was similar to that observed when radiosensitive lymphocytes are subjected to ionizing radiation. As in irradiated thymocytes, O2- generation affected primarily the maximal rate of uptake of the amino acid (i.e. Vmax). No change was observed in the apparent affinity of the amino acid for its carrier (i.e. Km) or the efflux rate of the amino acid. The data suggests that the superoxide anion may be one of the major species responsible for the observed radiation damage to radiosensitive lymphoid cells.     

ON01210.Na (Ex-RAD?) Mitigates Radiation Damage through Activation of the AKT Pathway

Development of radio-protective agents that are non-toxic is critical in light of ever increasing threats associated with proliferation of nuclear materials, terrorism and occupational risks associated with medical and space exploration. In this communication, we describe the discovery, characterization and mechanism of action of ON01210.Na, which effectively protects mouse and human bone marrow cells from radiation-induced damage both in vitro and in vivo. Our results show that treatment of normal fibroblasts with ON01210.Na before and after exposure to ionizing radiation provides dose dependent protection against radiation-induced damage. Treatment of mice with ON01210.Na prior to radiation exposure was found to result in a more rapid recovery of their hematopoietic system. The mechanistic studies described here show that ON01210.Na manifests its protective effects through the up-regulation of PI3-Kinase/AKT pathways in cells exposed to radiation. These results suggest that ON 01210.Na is a safe and effective radioprotectant and could be a novel agent for use in radiobiological disasters.     

Ionizing radiation-induced mutagenesis: radiation studies in Neurospora predictive for results in mammalian cells.

Ionizing radiation was the first mutagen discovered and was used to develop the first mutagenicity assay. In the ensuing 70+ years, ionizing radiation became a fundamental tool in understanding mutagenesis and is still a subject of intensive research. Frederick de Serres et al. developed and used the Neurospora crassa ad-3 system initially to explore the mutagenic effects of ionizing radiation. Using this system, de Serres et al. demonstrated the dependence of the frequency and spectra of mutations induced by ionizing radiation on the dose, dose rate, radiation quality, repair capabilities of the cells, and the target gene employed. This work in Neurospora predicted the subsequent observations of the mutagenic effects of ionizing radiation in mammalian cells. Modeled originally on the mouse specific-locus system developed by William L. Russell, the N. crassa ad-3 system developed by de Serres has itself served as a model for interpreting the results in subsequent systems in mammalian cells. This review describes the primary findings on the nature of ionizing radiation-induced mutagenesis in the N. crassa ad-3 system and the parallel observations made years later in mammalian cells.     

The role of regulatory networks of the cell response to damages in radiation effects

In view of modern knowledge and concepts about components, function and mechanisms of response of cell molecular structures to damaging effects, response which is generating specialized modules of reactions, it is shown that main components of the mechanism of maintenance of genome constancy at ionizing radiation exposure are checkpoints of cell cycle, DNA repair and apoptosis. They operate under the control of a genetic system at participation of Tp53 gene, corresponding protein and of regulatory networks formed by cascades of mitogen-activated protein kinases (MAPK). At ionizing radiation exposure the MAPK special modules participate in formation of radiation effect: ERK 1/2 (extracellular signal-regulated kinase 1 and 2), JNK/SAPK (c-Jun N-terminal kinase/stress activated protein kinase) and p38 MAPK. Executing physiological functions of maintenance of normal life activity of cells, they do not lose this capacity after exposure to ionizing radiation, participating in formation of radiation effect in a wide range of doses, and are inactivated only by exposure to very high doses. It is concluded that in light of the modern data the main problem is not a problem of mechanisms of biological effect of ionizing radiation but a problem of biological mechanisms of radiation exposure.