The effect of electromagnetic field exposure on the formation of DNA lesions

Abstract

In an attempt to determine whether electromagnetic field (EMF) exposure might lead to DNA damage, we exposed SnCl₂-treated pBR322 plasmids to EMF and analysed the resulting conformational changes using agarose gel electrophoresis. An EMF-dependent potentiation of DNA scission (i.e. the appearance of relaxed plasmids) was observed. In confirmation of this, plasmids pre-exposed to EMF also were less capable of transforming Escherichia coli. The results indicate that EMF, in the presence of a transition metal, is capable of causing DNA damage. These observations support the idea that EMF, probably through secondary generation of reactive oxygen species, can be clastogenic and provide a possible explanation for the observed correlation between EMF exposure and the frequency of certain types of cancers in humans.     

Short‐term exposure to 50 Hz ELF‐EMF alters the cisplatin‐induced oxidative response in AT478 murine squamous cell carcinoma cells

The aim of this study was to assess the influence of cisplatin and an extremely low frequency electromagnetic field (ELF‐EMF) on antioxidant enzyme activity and the lipid peroxidation ratio, as well as the level of DNA damage and reactive oxygen species (ROS) production in AT478 carcinoma cells. Cells were cultured for 24 and 72 h in culture medium with cisplatin. Additionally, the cells were irradiated with 50 Hz/1 mT ELF‐EMF for 16 min using a solenoid as a source of the ELF‐EMF. The amount of ROS, superoxide dismutase (SOD) isoenzyme activity, glutathione peroxidase (GSH‐Px) activity, DNA damage, and malondialdehyde (MDA) levels were assessed. Cells that were exposed to cisplatin exhibited a significant increase in ROS and antioxidant enzyme activity. The addition of ELF‐EMF exposure to cisplatin treatment resulted in decreased ROS levels and antioxidant enzyme activity. A significant reduction in MDA concentrations was observed in all of the study groups, with the greatest decrease associated with treatment by both cisplatin and ELF‐EMF. Cisplatin induced the most severe DNA damage; however, when cells were also irradiated with ELF‐EMF, less DNA damage occurred. Exposure to ELF‐EMF alone resulted in an increase in DNA damage compared to control cells. ELF‐EMF lessened the effects of oxidative stress and DNA damage that were induced by cisplatin; however, ELF‐EMF alone was a mild oxidative stressor and DNA damage inducer. We speculate that ELF‐EMF exerts differential effects depending on the exogenous conditions. This information may be of value for appraising the pathophysiologic consequences of exposure to ELF‐EMF. Bioelectromagnetics 33:641–651, 2012. © 2012 Wiley Periodicals, Inc.     

Extremely low frequency electromagnetic fields as effectors of cellular responses in vitro: possible immune cell activation

There is presently an intense discussion if electromagnetic field (EMF) exposure has consequences for human health. This include exposure to structures and appliances that emit in the extremely low frequency (ELF) range of the electromagnetic spectrum, as well as emission coming from communication devices using the radiofrequency part of the spectrum. Biological effects of such exposures have been noted frequently, although the implication for specific health effects is not that clear. The basic interaction mechanism(s) between such fields and living matter is unknown. Numerous hypotheses have been suggested, although none is convincingly supported by experimental data. Various cellular components, processes, and systems can be affected by EMF exposure. Since it is unlikely that EMF can induce DNA damage directly, most studies have examined EMF effects on the cell membrane level, general and specific gene expression, and signal transduction pathways. In addition, a large number of studies have been performed regarding cell proliferation, cell cycle regulation, cell differentiation, metabolism, and various physiological characteristics of cells. Although 50/60 Hz EMF do not directly lead to genotoxic effects, it is possible that certain cellular processes altered by exposure to EMF indirectly affect the structure of DNA causing strand breaks and other chromosomal aberrations. The aim of this article is to present a hypothesis of a possible initial cellular event affected by exposure to ELF EMF, an event which is compatible with the multitude of effects observed after exposure. Based on an extensive literature review, we suggest that ELF EMF exposure is able to perform such activation by means of increasing levels of free radicals. Such a general activation is compatible with the diverse nature of observed effects. Free radicals are intermediates in natural processes like mitochondrial metabolism and are also a key feature of phagocytosis. Free radical release is inducible by ionizing radiation or phorbol ester treatment, both leading to genomic instability. EMF might be a stimulus to induce an "activated state" of the cell such as phagocytosis, which then enhances the release of free radicals, in turn leading to genotoxic events. We envisage that EMF exposure can cause both acute and chronic effects that are mediated by increased free radical levels: (1) Direct activation of, for example macrophages (or other cells) by short-term exposure to EMF leads to phagocytosis (or other cell specific responses) and consequently, free radical production. This pathway may be utilized to positively influence certain aspects of the immune response, and could be useful for specific therapeutic applications. (2) EMF-induced macrophage (cell) activation includes direct stimulation of free radical production. (3) An increase in the lifetime of free radicals by EMF leads to persistently elevated free radical concentrations. In general, reactions in which radicals are involved become more frequent, increasing the possibility of DNA damage. (4) Long-term EMF exposure leads to a chronically increased level of free radicals, subsequently causing an inhibition of the effects of the pineal gland hormone melatonin. Taken together, these EMF induced reactions could lead to a higher incidence of DNA damage and therefore, to an increased risk of tumour development. While the effects on melatonin and the extension of the lifetime of radicals can explain the link between EMF exposure and the incidence of for example leukaemia, the two additional mechanisms described here specifically for mouse macrophages, can explain the possible correlation between immune cell system stimulation and EMF exposure.     

The effect of a low-frequency electromagnetic field on DNA molecules in aqueous solutions

A chemiluminescence study showed that hepatitis B virus (HBV) and hepatitis C virus (HCV) DNA amplicons are capable of induced radiation when exposed to electromagnetic fields (EMFs) that range from 7.5 to 30 Hz in frequency and from 24 to 40 A/m in field strength. An EMF with a frequency of 9 Hz was shown to exert the greatest effect on aqueous solutions of the hepatitis virus DNA amplicons. The hydration shell of the DNA amplicons was observed to change. The change in the DNA hydration shell on exposure to a low-frequency EMF was presumed to restore hydrogen bonds, to induce crosslinks, and to facilitate DNA repair.     

Assessing of plasma protein denaturation induced by exposure to cadmium,electromagnetic fields and their combined actions on rat

In our environment, we have numerous chances to be exposed to not only electromagnetic fields (EMFs) but also many chemicals containing mutagens. Therefore, the aim of this study was to estimate whether rat’s exposure to cadmium and/or EMFs could cause oxidative damage to molecular structure of proteins and whether and to what extent the effects of co-exposure differ from those observed under the treatment with each exposure alone. Thirty-two rats were divided into four groups. Group 1 was termed as control, group 2 was treated with cadmium (3.0?mg/Kg), group 3 was exposed to EMF (10?mT/h/day) and group 4 was treated with cadmium and exposed to EMF. Protein carbonyls (PCO) in the plasma as a marker of oxidative protein damage and total oxidant status (TOS), as well as electrical conductivity and SDS electrophoresis to estimate changes in molecular structure of protein, were determined. The exposure to Cd and/or EMF led to oxidative protein damage (increased PCO and TOS) accomplished by increased stress of electrical charges on the surface of the protein molecule (increased electrical conductivity) and changes in the molecular structure of protein. The effects were more pronounced after treatment with both Cd and EMF than at the treatment with each exposure alone. The serious damage to proteins at the co-exposure to Cd and EMF seems to be due to the interference of the EMF with the toxic activity of cadmium. This work concluded that combined exposure to Cd and EMFs might increase the risk of plasma damage via enhancing free radical generation and protein oxidation.     

Effects of prenatal exposure to a 900 MHz electromagnetic field on 60-day-old rat testis and epididymal sperm quality

We investigated the effects of exposure in utero to a 900 megahertz (MHz) electromagnetic field (EMF) on 60-day-old rat testis and epididymis. Pregnant rats were divided into control (CG; no treatment) and EMF (EMFG) groups. The EMFG was exposed to 900 MHz EMF for 1 h each day during days 13 ? 21 of pregnancy. Newborn rats were either newborn CG (NCG) or newborn EMF groups (NEMFG). On postnatal day 60, a testis and epididymis were removed from each animal. Epididymal semen quality, and lipid and DNA oxidation levels, apoptotic index and histopathological damage to the testis were compared. We found a higher apoptotic index, greater DNA oxidation levels and lower sperm motility and vitality in the NEMFG compared to controls. Immature germ cells in the seminiferous tubule lumen, and altered seminiferous tubule epithelium and seminiferous tubule structure also were observed in hematoxylin and eosin stained sections of NEMFG testis. Nuclear changes that indicated apoptosis were identified in TUNEL stained sections and large numbers of apoptotic cells were observed in most of the seminiferous tubule epithelium in the NEMFG. Sixty-day-old rat testes exposed to 900 MHz EMF exhibited altered sperm quality and biochemical characteristics.     

Decreased DNA repair rates and protection from heat induced apoptosis mediated by electromagnetic field exposure

In this study, we demonstrate that electromagnetic field (EMF) exposure results in protection from heat induced apoptosis in human cancer cell lines in a time dependent manner. Apoptosis protection was determined by growing HL-60, HL-60R, and Raji cell lines in a 0.15 mT 60 Hz sinusoidal EMF for time periods between 4 and 24 h. After induction of apoptosis, cells were analyzed by the neutral comet assay to determine the percentage of apoptotic cells. To discover the duration of this protection, cells were grown in the EMF for 24 h and then removed for 24 to 48 h before heat shock and neutral comet assays were performed. Our results demonstrate that EMF exposure offers significant protection from apoptosis (P<.0001 for HL-60 and HL-60R, P<.005 for Raji) after 12 h of exposure and that protection can last up to 48 h after removal from the EMF. In this study we further demonstrate the effect of the EMF on DNA repair rates. DNA repair data were gathered by exposing the same cell lines to the EMF for 24 h before damaging the exposed cells and non-exposed cells with H2O2. Cells were allowed to repair for time periods between 0 and 15 min before analysis using the alkaline comet assay. Results showed that EMF exposure significantly decreased DNA repair rates in HL-60 and HL-60R cell lines (P<.001 and P<.01 respectively), but not in the Raji cell line. Importantly, our apoptosis results show that a minimal time exposure to an EMF is needed before observed effects. This may explain previous studies showing no change in apoptosis susceptibility and repair rates when treatments and EMF exposure were administered concurrently. More research is necessary, however, before data from this in vitro study can be applied to in vivo systems.     

Exposure to 900 MHz electromagnetic field induces an unbalance between pro-apoptotic and pro-survival signals in T-lymphoblastoid leukemia CCRF-CEM cells

It has been recently established that low-frequency electromagnetic field (EMFs) exposure induces biological changes and could be associated with increased incidence of cancer, while the issue remains unresolved as to whether high-frequency EMFs can have hazardous effect on health. Epidemiological studies on association between childhood cancers, particularly leukemia and brain cancer, and exposure to low- and high-frequency EMF suggested an etiological role of EMFs in inducing adverse health effects. To investigate whether exposure to high-frequency EMFs could affect in vitro cell survival, we cultured acute T-lymphoblastoid leukemia cells (CCRF-CEM) in the presence of unmodulated 900 MHz EMF, generated by a transverse electromagnetic (TEM) cell, at various exposure times. We evaluated the effects of high-frequency EMF on cell growth rate and apoptosis induction, by cell viability (MTT) test, FACS analysis and DNA ladder, and we investigated pro-apoptotic and pro-survival signaling pathways possibly involved as a function of exposure time by Western blot analysis. At short exposure times (2-12 h), unmodulated 900 MHz EMF induced DNA breaks and early activation of both p53-dependent and -independent apoptotic pathways while longer continuous exposure (24-48 h) determined silencing of pro-apoptotic signals and activation of genes involved in both intracellular (Bcl-2) and extracellular (Ras and Akt1) pro-survival signaling. Overall our results indicate that exposure to 900 MHz continuous wave, after inducing an early self-defense response triggered by DNA damage, could confer to the survivor CCRF-CEM cells a further advantage to survive and proliferate.     

Induction of cell activation processes by low frequency electromagnetic fields

Electromagnetic fields (EMF) such as those from electric power transmission and distribution lines (50/60 Hz) have been associated with increased risk of childhood leukemia, cancer of the nervous system, and lymphomas. Several in vitro studies on EMF effects were performed to clarify the existing controversies, define the risks, and determine the possible mechanisms of adverse effects. In some of these reports, the effects were related to other mechanisms of carcinogenesis. Modification in cell proliferation was observed after EMF exposure and a few reports on cytotoxic effects have also been published. This limited review gives an overview of the current results of scientific research regarding in vitro studies on the effects of power line frequency EMF, but also cell biological mechanisms and their potential involvement in genotoxicity and cytotoxicity are discussed. Cell cycle control and signal transduction processes are included to elucidate the biochemical background of possible interactions. Exposure to EMF has been also linked to the incidence of leukemia and other tumors in some epidemiological studies and is considered as "possibly carcinogenic to humans", but there is no well-established biological mechanism that explains such a relation. Furthermore, EMF is also shown as a stimulus for immune relevant cells (e.g., macrophages) to release free radicals. It is known that chronic activation of macrophages is associated with the onset of phagocytosis and leads to increased formation of reactive oxygen species, which themselves may cause DNA damage and are suggested to lead to carcinogenesis. To demonstrate a possible interaction between EMF and cellular systems, we present a mechanistic model describing cell activation as a major importance for cellular response.     

Exposure to 900 MHz electromagnetic field induces an unbalance between pro‐apoptotic and pro‐survival signals in T‐lymphoblastoid leukemia CCRF‐CEM cells

The original article to which this Erratum was published in J. Cell. Physiol. 198:324–332, 2004 It has been recently established that low‐frequency electromagnetic field (EMFs) exposure induces biological changes and could be associated with increased incidence of cancer, while the issue remains unresolved as to whether high‐frequency EMFs can have hazardous effect on health. Epidemiological studies on association between childhood cancers, particularly leukemia and brain cancer, and exposure to low‐ and high‐frequency EMF suggested an etiological role of EMFs in inducing adverse health effects. To investigate whether exposure to high‐frequency EMFs could affect in vitro cell survival, we cultured acute T‐lymphoblastoid leukemia cells (CCRF‐CEM) in the presence of unmodulated 900 MHz EMF, generated by a transverse electromagnetic (TEM) cell, at various exposure times. We evaluated the effects of high‐frequency EMF on cell growth rate and apoptosis induction, by cell viability (MTT) test, FACS analysis and DNA ladder, and we investigated pro‐apoptotic and pro‐survival signaling pathways possibly involved as a function of exposure time by Western blot analysis. At short exposure times (2–12 h), unmodulated 900 MHz EMF induced DNA breaks and early activation of both p53‐dependent and ‐independent apoptotic pathways while longer continuous exposure (24–48 h) determined silencing of pro‐apoptotic signals and activation of genes involved in both intracellular (Bcl‐2) and extracellular (Ras and Akt1) pro‐survival signaling. Overall our results indicate that exposure to 900 MHz continuous wave, after inducing an early self‐defense response triggered by DNA damage, could confer to the survivor CCRF‐CEM cells a further advantage to survive and proliferate. J. Cell. Physiol. 198: 324–332, 2004. © 2003 Wiley‐Liss, Inc.     

Extremely low-frequency electromagnetic fields do not affect DNA damage and gene expression profiles of yeast and human lymphocytes

We studied the effects of extremely low-frequency (50 Hz) electromagnetic fields (EMFs) on peripheral human blood lymphocytes and DBY747 Saccharomyces cerevisiae. Graded exposure to 50 Hz magnetic flux density was obtained with a Helmholtz coil system set at 1, 10 or 100 microT for 18 h. The effects of EMFs on DNA damage were studied with the single-cell gel electrophoresis assay (comet assay) in lymphocytes. Gene expression profiles of EMF-exposed human and yeast cells were evaluated with DNA microarrays containing 13,971 and 6,212 oligonucleotides, respectively. After exposure to the EMF, we did not observe an increase in the amount of strand breaks or oxidated DNA bases relative to controls or a variation in gene expression profiles. The results suggest that extremely low-frequency EMFs do not induce DNA damage or affect gene expression in these two different eukaryotic cell systems.     

Effect of 910-MHz electromagnetic field on rat bone marrow

Aiming to investigate the possibility of electromagnetic fields (EMF) developed by nonionizing radiation to be a noxious agent capable of inducing genotoxicity to humans, in the current study we have investigated the effect of 910-MHz EMF in rat bone marrow. Rats were exposed daily for 2 h over a period of 30 consecutive days. Studying bone marrow smears from EMF-exposed and sham-exposed animals, we observed an almost threefold increase of micronuclei (MN) in polychromatic erythrocytes (PCEs) after EMF exposure. An induction of MN was also observed in polymorphonuclear cells. The induction of MN in female rats was less than that in male rats. The results indicate that 910-MHz EMF could be considered as a noxious agent capable of producing genotoxic effects.     

Effect of DNA damage on stable transformation of mammalian cells with integrative and episomal plasmids

The efficiency of stable transformation of human cells by integrative (non-replicating) plasmids carrying a selectable gene has been shown to be markedly enhanced by the introduction into the plasmid DNA of bulky damage, such as cyclobutane pyrimidine dimers or psoralen photoadducts. Enhanced transformation (ET) occurs in all human cells tested, including DNA repair-deficient cells from the hereditary syndrome xeroderma pigmentosum, but significantly less, if at all, in rodent cells. ET has been observed with a variety of integrative plasmid constructs, suggesting the generality of the phenomenon; as expected, ET is due to an increase in the number of cells carrying integrated plasmid sequences. In contrast to integrative plasmids, stable transformation by episomal (autonomously replicating) plasmids derived from the Epstein-Barr virus is only depressed by the introduction of photoproducts; furthermore, pronounced inactivation of transformation mediated by episomal plasmids becomes apparent in xeroderma pigmentosum cells. Altogether, these results suggest that DNA damage increases the probability of stable insertion of heterologous non-replicating DNA into human chromosomes. Moreover, the differential sensitivity to DNA damage of human cell transformation mediated by integrative versus episomal plasmids suggests caution in using such assay to measure host cell reactivation capacity; processing of DNA damage in mammalian cells might differ significantly between intra- versus extra-chromosomal DNA. Since ET may be induced by damage outside the selectable gene carried on integrative plasmids, we propose a model that involves local disruption of chromatin structure by helix-distorting DNA lesions flanking actively transcribed sequences; alternatively, reorganization of such altered DNA structure might be favored by the presence of topoisomerase-like activities in the proximity of active genes. Because ET can also be induced by DNA damage to the recipient cells, it is speculated that similar mechanism(s) might be involved in the generation of other types of non-homologous DNA recombination in damaged human chromosomes, including oncogenic cell transformation mediated by integrative DNA viruses.     

Regulation of c-fos is affected by electromagnetic fields

The goal of the present study was to determine if regulatory regions of the c-fos gene were responsive to electromagnetic field exposure. The research design used transfected cells to increase the sensitivity of assays designed to identify changes following exposure. HeLa cells were transiently transfected with plasmids containing upstream regulating regions of c-fos up to -700 base pairs, coupled with the prokaryotic reporter gene CAT. Cells were exposed to an environmentally relevant EMF of 60 Hz at 60 mG_rms. CAT expression above control levels in transfected cells (region +42 to -700 bp) was observed following 5 min exposure to the electromagnetic field, with a peak at 20 min. The expression was at basal levels following 40 min exposure. Deletion analysis of upstream DNA narrowed the responsive region to 138 base pairs from -363 to -225, which contains the SRE/AP-1 sites. © 1996 Wiley-Liss, Inc.     

Oxidative DNA damage in rats exposed to extremely low frequency electro magnetic fields

Extremely low frequency (ELF) electromagnetic field (EMF) is thought to prolong the life of free radicals and can act as a promoter or co-promoter of cancer. 8-hydroxy-2'-deoxyguanosine (8OHdG) is one of the predominant forms of radical-induced lesions to DNA and is a potential tool to asses the cancer risk. We examined the effects of extremely low frequency electro magnetic field (ELF-EMF) (50 Hz, 0.97 mT) on 8OHdG levels in DNA and thiobarbituric acid reactive substances (TBARS) in plasma. To examine the possible time-dependent changes resulting from magnetic field, 8OHdG and TBARS were quantitated at 50 and 100 days. Our results showed that the exposure to ELF-EMF induced oxidative DNA damage and lipid peroxidation (LPO). The 8OHdG levels of exposed group (4.39+/-0.88 and 5.29+/-1.16 8OHdG/dG.10(5), respectively) were significantly higher than sham group at 50 and 100 days (3.02+/-0.63 and 3.46+/-0.38 8OHdG/dG.10(5)) (p<0.001, p<0.001). The higher TBARS levels were also detected in the exposure group both on 50 and 100 days (p<0.001, p<0.001). In addition, the extent of DNA damage and LPO would depend on the exposure time (p<0.05 and p<0.05). Our data may have important implications for the long-term exposure to ELF-EMF which may cause oxidative DNA damage.     

DNA damage in frog erythrocytes after in vitro exposure to a high peak-power pulsed electromagnetic field

Till the present time, the genotoxic effects of high peak-power pulsed electromagnetic fields (HPPP EMF) on cultured cells have not been studied. We investigated possible genotoxic effects of HPPP EMF (8.8 GHz, 180 ns pulse width, peak power 65 kW, repetition rate 50 Hz) on erythrocytes of the frog Xenopus laevis. We used the alkaline comet assay, which is a highly sensitive method to assess DNA single-strand breaks and alkali-labile lesions. Blood samples were exposed to HPPP EMF for 40 min in rectangular wave guide. The specific absorption rate (SAR) calculated from temperature kinetics was about 1.6 kW/kg (peak SAR was about 300 MW/kg). The temperature rise in the blood samples at steady state was 3.5 +/- 0.1 degrees C. The data show that the increase in DNA damage after exposure of erythrocytes to HPPP EMF was induced by the rise in temperature in the exposed cell suspension. This was confirmed in experiments in which cells were incubated for 40 min under the corresponding temperature conditions. The results allow us to conclude that HPPP EMF-exposure at the given modality did not cause any a-thermal genotoxic effect on frog erythrocytes in vitro.     

Controversial cytogenetic observations in mammalian somatic cells exposed to extremely low frequency electromagnetic radiation: a review and future research recommendations

During the years 1990-2003, a large number of investigations were conducted using animals, cultured rodent and human cells as well as freshly collected human blood lymphocytes to determine the genotoxic potential of exposure to nonionizing radiation emitted from extremely low frequency electromagnetic fields (EMF). Among the 63 peer reviewed scientific reports, the conclusions from 29 studies (46%) did not indicate increased damage to the genetic material, as assessed from DNA strand breaks, incidence of chromosomal aberrations (CA), micronuclei (MN), and sister chromatid exchanges (SCE), in EMF exposed cells as compared with sham exposed and/or unexposed cells, while those from 14 investigations (22%) have suggested an increase in such damage in EMF exposed cells. The observations from 20 other studies (32%) were inconclusive. This study reviews the investigations published in peer reviewed scientific journals during 1990-2003 and attempts to identify probable reason(s) for the conflicting results. Recommendations are made for future research to address some of the controversial observations.     

Long-term effects of 60-Hz electric vs. magnetic fields on IL-1 and IL-2 activity in sheep

This study was designed to assess the effect of exposure to long-term extremely low-frequency electric and magnetic fields (ELF-EMF) from a 500 kV transmission line on IL-1 and IL-2 activity in sheep. The primary hypothesis was that the reduction in IL-1 activity observed in our two previous short-term studies (10 months) was due to EMF exposure from this transmission line. To repeat and expand these studies and to characterize the components of EMF responsible for the previously observed reduction in IL-1 activity, the current experiment examined not only the effect of exposure to electric and magnetic fields, but also the magnetic field component alone. In the current study, IL-2 was examined to characterize the effects of EMF exposure on an indicator of T cell responses. 45 Suffolk ewe lambs were randomized into three groups of 15 animals each. One group of animals was placed in the EMF pen, located directly beneath the transmission line. A second group was placed in the shielded MF (magnetic field only) pen, also directly beneath the transmission line. The third group of animals was placed in the control pen located several hundred meters away from the transmission line. During the 27 month exposure period, blood samples were taken from all animals monthly. When the data were analyzed collectively over time, no significant differences between the groups were found for IL-1 or IL-2 activity. In previous studies ewe lambs of 8-10 weeks of age were used as the study animals and significant differences in IL-1 activity were observed after exposure of these animals to EMF at mean magnetic fields of 3.5-3.8 microT (35-38 mG) and mean electric fields of 5.2-5.8 kV/m. At the start of the current study EMF levels were reduced as compared to previous studies. One interpretation of the current data is that magnetic field strength and age of the animals may be important variables in determining whether EMF exposure will affect IL-1 activity.