Protection against adverse biological effects induced by space radiation by the Bowman-Birk inhibitor and antioxidants.

This study was undertaken to evaluate the protective effects of the soybean-derived Bowman-Birk inhibitor (BBI), BBI concentrate (BBIC) and/or antioxidants against the adverse biological effects induced by space radiation in cultured human epithelial cells. The effects of BBI, BBIC and a combination of ascorbic acid, co-enzyme Q10, L-selenomethionine (SeM) and vitamin E succinate on proton and HZE-particle [high-energy (high E) nuclei of heavier (high atomic number, Z) elements] radiation-induced cytotoxicity in MCF10 human breast epithelial cells and a phenotypic change associated with transformation in HTori-3 human thyroid epithelial cells were assessed with a clonogenic survival assay and a soft agar colony formation assay. The results demonstrate that BBIC and antioxidants are effective in protecting against space radiation-induced cytotoxicity in MCF10 cells and BBI, BBIC and antioxidants are effective in protecting against a space radiation-induced phenotypic change associated with transformation of HTori-3 cells.

Effects of dietary supplements on the space radiation-induced reduction in total antioxidant status in CBA mice.

In the present study, the total antioxidant status was used as a biomarker to evaluate oxidative stress induced by proton, HZE-particle and gamma radiation in CBA mice. The results demonstrated that the plasma level of TAS was significantly decreased (P < 0.05) in CBA mice after exposure to a 50-cGy dose of radiation from HZE particles or a 3-Gy dose of radiation from protons or gamma rays. Diet supplementation with Bowman-Birk Inhibitor Concentrate (BBIC), L-selenomethionine (L-SeM), or a combination of N-acetyl cysteine, sodium ascorbate, co-enzyme Q10 (CoQ10), alpha-lipoic acid, L-SeM and vitamin E succinate could partially or completely prevent the reduction in the plasma level of TAS in CBA mice exposed to proton or HZE-particle radiation. The selected antioxidant combination with or without CoQ10 has a comparable protective effect on the gamma-radiation-induced drop in TAS in CBA mice. These results indicate that BBIC, L-SeM and the selected antioxidant combinations may serve as countermeasures for space radiation-induced adverse biological effects.

Protection against radiation-induced oxidative stress in cultured human epithelial cells by treatment with antioxidant agents.

PURPOSE: To evaluate the protective effects of antioxidant agents against space radiation-induced oxidative stress in cultured human epithelial cells. METHODS AND MATERIALS: The effects of selected concentrations of N-acetylcysteine, ascorbic acid, sodium ascorbate, co-enzyme Q10, alpha-lipoic acid, l-selenomethionine, and vitamin E succinate on radiation-induced oxidative stress were evaluated in MCF10 human breast epithelial cells exposed to radiation with X-rays, gamma-rays, protons, or high mass, high atomic number, and high energy particles using a dichlorofluorescein assay. RESULTS: The results demonstrated that these antioxidants are effective in protecting against radiation-induced oxidative stress and complete or nearly complete protection was achieved by treating the cells with a combination of these agents before and during the radiation exposure. CONCLUSION: The combination of antioxidants evaluated in this study is likely be a promising countermeasure for protection against space radiation-induced adverse biologic effects.

Role of vicinal protein thiols in radiation and cytotoxic responses.

Glutathione (GSH) and more recently protein thiols (P-SH) have been found to play a major role in cellular radiation response. However, the effects of protein vicinal thiols, which are important for the functions of several major enzymes, on cellular responses to radiation have not been clearly delineated. Here we investigated the effects of depleting GSH and protein vicinal thiols (HS-P-SH) and P-SH on cell toxicity and radiation response. We used hydroxyethyldisulfide (HEDS, beta-mercaptoethanol-disulfide) alone and in combination with phenylarsine oxide (PAO) to alter P-SH, HS-P-SH and GSH. HEDS, a direct substrate for thioredoxin reductase and an indirect substrate for glutaredoxin (thioltransferase), did not alter protein vicinal thiols in cells. However, PAO, which specifically forms a covalent adduct with vicinal thiols, blocked bioreduction of HEDS; there was a concomitant and yet unexplained decrease in K1 cell GSH in the presence of HEDS and PAO. G6PD+ (K1) and G6PD- (E89) cells treated with L-buthionine sulfoximine (L-BSO) for 72 h to deplete GSH followed by PAO showed an increased cytotoxic response. However, the surviving E89 cells showed a 10,000-fold greater radiation lethality than the K1 cells. The effects of rapid depletion of GSH by a combination of L-BSO and dimethyfumarate (DMF), a glutathione-S-transferase substrate, were also investigated. Under these conditions, PAO radiosensitized the E89 cells more than 1000-fold over the K1 cells. The potential mechanisms for the altered response may be related to the inhibition of thioredoxin reductase and glutaredoxin. Both are key enzymes involved in DNA synthesis, protein homeostasis and cell survival. With GSH removed, vicinal thiols appear to play a critical role in determining cell survival and radiosensitivity. Decreasing P-SH and removing GSH and vicinal thiols is extremely toxic to K1 and E89 cells. We conclude that radiation sensitivity and cell survival are dependent on vicinal thiol and GSH. In the former and latter cases, the protein thiols are also important.

Detection of oxidative stress induced by low- and high-linear energy transfer radiation in cultured human epithelial cells.

A standardized dichlorofluorescin (DCF) fluorometric assay capable of measuring radiation-induced oxidative stress was used to determine the effectiveness of protons and high-mass, high-atomic number (Z) and high-energy (HZE) particles to produce oxidative stress in vitro. Protons were found to be about equally as effective as X rays in the generation of oxidative stress in cultured cells. However, 56Fe-ion beams with energies of 1 GeV/nucleon and 5 GeV/nucleon were less effective than X rays or gamma rays in inducing dichlorofluorescin (DCFH) oxidation. The relatively lower slope values for the dose responses of HZE-particle radiation-induced DCFH oxidation indicate that the sensitivity of the DCF fluorometric assay is probably dependent on the linear energy transfer (LET) of the radiation beam.

Effects of dietary supplements on space radiation-induced oxidative stress in Sprague-Dawley rats.

Of particular concern for the health of astronauts during space travel is radiation from protons and high-mass, high-atomic-number (Z), and high-energy particles (HZE particles). Space radiation is known to induce oxidative stress in astronauts after extended space flight. In the present study, the total antioxidant status was used as a biomarker to evaluate oxidative stress induced by gamma rays, protons and HZE-particle radiation. The results demonstrate that the plasma level of total antioxidants in Sprague-Dawley rats was significantly decreased (P < 0.01) in a dose-dependent manner within 4 h after exposure to gamma rays. Exposure to protons and HZE-particle radiation also significantly decreased the serum or plasma level of total antioxidants in the irradiated animals. Diet supplementation with L-selenomethionine alone or a combination of selected antioxidant agents was shown to partially or completely prevent the decrease in the serum or plasma levels of total antioxidants in animals exposed to gamma rays, protons or HZE particles. These findings suggest that exposure to space radiation may compromise the capacity of the host antioxidant defense and that this adverse biological effect can be prevented at least partially by dietary supplementation with L-selenomethionine and antioxidants.

Selenomethionine protects against adverse biological effects induced by space radiation.

Ionizing radiation-induced adverse biological effects impose serious challenges to astronauts during extended space travel. Of particular concern is the radiation from highly energetic, heavy, charged particles known as HZE particles. The objective of the present study was to characterize HZE particle radiation-induced adverse biological effects and evaluate the effect of D-selenomethionine (SeM) on the HZE particle radiation-induced adverse biological effects. The results showed that HZE particle radiation can increase oxidative stress, cytotoxicity, and cell transformation in vitro, and decrease the total antioxidant status in irradiated Sprague-Dawley rats. These adverse biological effects were all preventable by treatment with SeM, suggesting that SeM is potentially useful as a countermeasure against space radiation-induced adverse effects. Treatment with SeM was shown to enhance ATR and CHK2 gene expression in cultured human thyroid epithelial cells. As ionizing radiation is known to result in DNA damage and both ATR and CHK2 gene products are involved in DNA damage, it is possible that SeM may prevent HZE particle radiation-induced adverse biological effects by enhancing the DNA repair machinery in irradiated cells.

Radiation response of cells during altered protein thiol redox.

The major focus of this work was to investigate how altered protein thiol redox homeostasis affects radiation-induced cell death. We used the cells of wild-type CHO cell line K1, the CHO cell line E89, which is null for G6PD activity, and a radiation-sensitive CHO cell line, XRS5. The protein-thiol redox status of cells was altered with cell-permeable disulfides, hydroxyethyldisulfide (HEDS) or lipoate. HEDS is primarily reduced by thioltransferase (glutaredoxin), with GSH as the electron donor. In contrast, lipoate is reduced by thioredoxin reductase. HEDS was reduced at a greater rate than lipoate by G6PD-containing K1 (wild-type) cells. Reduction of disulfides by G6PD-deficient cells was significantly slower with HEDS as substrate and was nearly absent with lipoate. The rate of reduction of HEDS by E89 cells decelerated to near zero by 30 min, whereas the reduction continued at nearly the same rate during the entire measurement period for K1 cells. HEDS treatment decreased the GSH and protein thiol (PSH) content more in G6PD-deficient cells than in G6PD-containing cells. On the other hand, lipoate did not significantly alter the protein thiol, but it increased the GSH in K1 cells. Acute depletion of GSH by l-buthionine-sulfoximine (l-BSO) in combination with dimethylfumarate significantly decreased the rate of reduction of HEDS by K1 cells close to that of G6PD-deficient cells. Prior GSH depletion by l-BSO alone significantly decreased the PSH in glucose-depleted E89 cells exposed to HEDS, but this did not occur with K1 cells. The radiation response of G6PD-deficient cells was significantly sensitized by HEDS, but HEDS did not have this effect on K1 cells. The DNA repair-deficient XRS5 CHO cells displayed the same capacity as K1 cells for HEDS reduction, and like K1 cells the XRS5 cells were not sensitized to radiation by HEDS treatment. Deprivation of glucose, which provides the substrate for G6PD in the oxidative pentose phosphate cycle, decreased the rate of bioreduction of HEDS and lipoate in G6PD-containing cells to the level in G6PD-deficient cells. In the absence of glucose, HEDS treatment diminished non-protein thiol and protein thiol to the same level as those in G6PD-deficient cells and sensitized the K1 cells to HEDS treatment. However, depletion of glucose did not alter the sensitivity of XRS5 cells in either the presence or absence of HEDS. Overall the results suggest a major role for pentose cycle control of protein redox state coupled to the activities of the thioltransferase and thioredoxin systems. The results also show that protein thiol status is a critical factor in cell survival after irradiation.