| With the rapid increasing use of electric power for domestic and industrial appliances, the growing of extremely low frequency electromagnetic fields (ELF EMF) exposure emitted by these appliances in the range of 50 Hz or 60 Hz, has raised public concerns about its potential health hazard. Evaluation the health effects of ELF EMF has become one of important public health issues.A number of epidemiological studies have reported the possible association between ELF EMF exposure and the incidence of certain cancers, such as leukaemia and breast cancer. The IARC classified ELF EMF as the 2B category (possible human carcinogen) in 2001, but it is still lack of evidence to make a final conclusion. DNA is the important hereditary material,.and easy to be caused damage and repair bug by environmental factors. Due to DNA damage related to mutagenic and carcinogenic risk, the effects of ELF EMF on DNA damage has became a key issue in bioelectromagnetics field. But after 30 years studies, these is controversial.on the effects of ELF EMF on DNA damage,Recently, the relationship between histone H2AX and DNA double-strand breaks (DSBs) has gradually drawed researchers' attention. Histone H2AX is phosphorylated (denoted asγH2AX) in early response to DSBs induced by many stimuli.γH2AX recruit many DNA repair proteins, such as BRCA1, 53BP1, and formγH2AX foci. It has demonstrated that the number ofγH2AX foci is quantitatively the same as DSBs induced by various stimuli. Thus,γH2AX foci formation is proposed to be a sensitive biomarker for DSBs, though it has also been reported to form in response to single-stranded DNA resulting from incomplete nucleotide excision repair. It could be a sensitive method to detect DSBs induced by ELF EMF, which a weak stimulus in the environment. Litovitz et al. proposed that living cells were affected only by EMF which were temporally and spatially coherent, and they suggested that the bioeffects induced by EMF would be interfered when superposed with a spatially coherent but temporally incoherent "noise" magnetic field.In this study, we investigated the effects of 0.4 mT 50 Hz ELF EMF on DNA damage and repair in human lens epithelia cells (hLECs), as well as the interfering effects of noise magnetic fields on DNA damage induced by ELF EMF using comet assay andγH2AX inmmunofluorescence measurement.1. Effects of 0.4 mT 50 Hz ELF EMF on DNA damage in hLECs.Cells were divided into seven groups: sham exposure group, ELF EMF exposure for 2 h group, ELF EMF exposure for 6 h group, ELF EMF exposure for 12 h group, ELF EMF exposure for 24 h group, ELF EMF exposure for 48 h group, and positive control group (0.1 umol/L 4-nitroquinoline-1-oxide (4NQO) exposuse for 1 h). Cells were sham-exposed or continuously exposed to 50 Hz ELF EMF for 2, 6, 12, 24 or 48 h at the intensity of 0.4 mT. After exposure, cells were collected at the same time.and processed for alkaline comet assay andγH2AX inmmunofluorescence measurement, respectively. In alkaline comet assay, cells embedded in agarose were lysed, helix-untied, electrophoresed, stained with ethidium bromide (EB), and observed by fluorescence microscopy. At least 50 cells randomly selected in each sample to evaluate DNA damage, and the tail length and tail moment were adopted as the indexes for evaluating DNA damage. The results showed that tail length and tail moment were significantly increased in positive control group compared with the sham exposure group (P<0.05). With the exposure duration of ELF EMF increased, the tail length and tail moment were increasing, but no significant difference compared with the sham exposure group (P>0.05). InγH2AX inmmunofluorescence measurement, the primary antibody was mouse monoclonal antibody againstγH2AX and the secondary antibody was fluorescein isothiocyanate (FITC)-conjugated goat anti-mouse IgG. Nuclei were counterstained with 4,6-diamidino-2-phenylindole (DAPI). The samples were examined with an Olympus AX70 fluorescent microscope. For each sample, at least 200 cells were randomly selected to detectγH2AX foci. Cells were classified as positive when more than 3γH2AX foci were detected. The percentage ofγH2AX foci positive cells and the mean number of foci per cell were adopted as the indexes for evaluating DSBs. There were significant differences in the percentage ofγH2AX foci positive cells and the mean number of foci per cell in 24 h or 48 h exposure group or positive control group compared with the sham exposure group (P<0.05). However, there was no significant difference in 2, 6 or 12 h exposure group compared with the sham exposure group (P>0.05).2. Repair of DNA damage induced by 0.4 mT 50 Hz ELF EMF in hLECs.Cells were continued to incubate for 0, 1, 2 or 4 h after continuous exposure to 50 Hz ELF EMF for 48 h at the intensity of 0.4 mT. The parallel sham exposure for each exposure group were set. Then cells were fixed using 4% paraformaldehyde and processed forγH2AX immunofluorescence measurement. The results showed that no significant difference was found in the percentage ofγH2AX foci positive cells and the mean number of foci per cell among each sham exposure group, and also no significant difference in 1 h or 2 h incubation after exposure group compared with 48 h exposure group. However, the two indexes were significantly decreased in 4 h incubation after ELF EMF exposure(P<0.05).3. Interfering effect of noise magnetic fields on DNA damage induced by 0.4 mT50 Hz ELF EMF in hLECs.Cells were divided into five groups: sham exposure group, ELF EMF exposure for 24 h group, ELF EMF exposure for 48 h group, ELF EMF superposed noise MF exposue for 24 h group, and ELF EMF superposed noise MF exposure for 48 h group. Cells were sham-exposed or continuously exposed to 50 Hz ELF EMF for 24 h or 48 h at the intensity of 0.4 mT, superposed or not superposed of noise MF. After exposure, cells were fixed using 4% paraformaldehyde and processed forγH2AX immunofluorescence measurement. The results showed that the percentage ofγH2AX foci positive cells and the mean number of foci per cell in ELF EMF 24 h or 48 h exposure group were significantly higher than the sham exposure group (P<0.05). However, the two indexes were significantly decreased in ELF EMF superposed noise MF exposure for 24 h or 48 h group compared with ELF EMF 24 h or 48 h exposure group (P<0.05).Our preliminary finding showed that 0.4 mT 50 Hz ELF EMF exposed for 24 h or 48 h could induce DNA damage in hLECs, and these damages would be partly repaired after exposure. Noise MF could partly block the effect of 50 Hz ELF EMF on DNA damage in hLECs.
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