Content of iron and copper in the nuclei and induction of pH 9-labile lesions in L5178Y sublines inversely cross-sensitive to H2O2 and x-rays

Abstract

Cells from the L5178Y murine lymphoma subline LY-R are twice as resistant to killing by ionizing radiation than the subline LY-S. In contrast, LY-R cells are more sensitive to killing by H2O2, the effect being more pronounced at 37 degrees C than 0 degree C. Initial DNA damage after H2O2 treatment (both temperatures, 5 min) has been estimated by the 'comet' assay (single-cell gel electrophoresis) and fluorescent halo technique. According to both methods, the initial damage is significantly higher in LY-R cells, particularly that inflicted at 0 degree C. Differences between DNA unwinding and rewinding abilities at pH 9 and 6.9 (estimated by the fluorescent halo technique) point to a considerable difference in pH-9-labile damage between the sublines, as observed previously for x-irradiated cells (Kapiszewska et al. 1992). In contrast to findings with x-irradiated cells, however, after H2O2 treatment this damage is more extensive in LY-R cells than in LY-S cells. Thus, the initial pH-9-labile damage corresponds to the pattern of sensitivity to H2O2 and x-rays. We suggest that this is caused by different proportions of cuprous and ferric ions found in the nuclei of LY sublines and by the different ability of these ions to react with H2O2 and water radiolysis products. The copper/iron ratio in the nucleus is 1.31 in LY-R cells and 4.84 in LY-S cells.

Labile iron pool correlates with iron content in the nucleus and the formation of oxidative DNA damage in mouse lymphoma L5178Y cell lines.

“…The LIP level in LY-R cells is 3-times higher than in LY-S cells (Lipinski et al, 2000) (Table 1). Interestingly, it is well correlated with the content of iron in L5178Y cell nuclei, previously measured by flame atomic absorption spectrometry (Szumiel et al, 1995) (Table 1).…”

“…Interestingly, Petrat et al (2001) found that hepatocyte nucleus contained 6.6 ± 2.9 mM of Total Fe content in the nucleus (ng/10 6 cells) (Szumiel et al, 1995) 7.7 ± 1.8 3.1 ± 0.9 2.48 chelatable iron. Although the amount of total concentration iron in hepatocytes was estimated to be similar to that of the cytoplasm (Lai et al, 1996), a surprisingly high content of redox-active iron in the nucleus presents a hazard of DNA damage.…”

“…A spontaneous loss of radiation resistance of LY-R cells and its conversion into radiation-sensitive LY-S cells was accompanied by an unexpected increase in H 2 O 2 resistance of the latter cells (Beer et al, 1983;Bouzyk et al, 1991;Kruszewski & Szumiel, 1994). The lower H 2 O 2 sensitivity of LY-S cells than that of LY-R cells is related to their better antioxidant defence system (Bouzyk et al, 1997) and lower content of transition metal ions in the nucleus (Szumiel et al, 1995). In this study, we used a modified comet assay to assess the oxidative DNA damage inflicted in both cell lines by H 2 O 2 treatment.…”

Nuclear translocation of the p65 subunit of NF-κB in L5178Y sublines differing in antioxidant defense

“…We found that the amount of initial DNA damage [1,3] and iron content [3] contribute to the difference in sensitivity to hydrogen peroxide. Further experimental data [4] pointed to a lower efficiency of the antioxidant defense system in the hydrogen peroxide-sensitive LY-R cells than in the hydrogen peroxide-resistant LY-S cells.…”

<title>Biological dosimetry after H<formula><inf><roman>2</roman></inf></formula>O<formula><inf><roman>2</roman></inf></formula>/L-histidine treatment</title>

“…H202 is known to induce single strand DNA breaks in mammalian cells 17,18 In combination with L-histidine also double strand breaks are induced for 11202 concentrations above 20 jiM 19 The reason and the mechanism for this interaction is not completely understood 18 However, as for ionizing radiation, intracellular misrepair of the double strand breaks leads to a complex pattern of chromosome aberrations including dicentric chromosomes20. H202 is known to induce single strand DNA breaks in mammalian cells 17,18 In combination with L-histidine also double strand breaks are induced for 11202 concentrations above 20 jiM 19 The reason and the mechanism for this interaction is not completely understood 18 However, as for ionizing radiation, intracellular misrepair of the double strand breaks leads to a complex pattern of chromosome aberrations including dicentric chromosomes20.…”