Effect of intermittent and continuous exposure to electromagnetic fields on cultured hippocampal cells

This study was designed to assess the effect of 50 Hz electromagnetic fields (EMFs) on hippocampal cell cultures in the presence or absence of either sodium nitroprusside (SNP, a NO donor) or Fe2+ induced oxidative stress. One week old cultured rat hippocampal cells were exposed to either intermittent EMFs (IEMFs, 50 Hz, 0-5 mT, 1 min ON/OFF cycles, repeated 10 times every 2 h, 6 times/day during 48 h) or continuous EMFs (CEMFs, 50 Hz, 0-5 mT for 48 h). In a second set of experiments, the effect on such EMFs applied in combination with oxidative stress induced by 0.5 microM Fe2+ or SNP was estimated. At the end of both sets of experiments, cell mortality was assessed by lactate dehydrogenase measurements (LDH). Neither type of exposure to EMFs was observed to modify the basal rate of cell mortality. The exposure to CEMFs in presence of either NO or Fe2+ did not induce any significant increase in cell death. However, when cells were exposed to EMFs in the presence of NO, we observed a significant increase in cell death of 11 and 23% (P<0.001) at 2.5 and 5 mT, respectively. This effect had some specificity because IEMFs did not modify the effect of Fe2+ on cell mortality. Although the effects of IEMFs reported in this study were only observed at very high intensities, our model may prove valuable in trying to identify one cellular target of EMFs.     

50Hz 1．8mT电磁场对成骨细胞增殖与分化成熟影响的波形比较研究

在50 Hz 1.8 mT的4种不同波形电磁场(electromagnetic fields,EMFs)中筛选促进体外培养大鼠成骨细胞(rat osteoblasts,ROB)增殖与分化成熟的最佳波形.体外分离培养大鼠颅骨成骨细胞,传代后随机分为5组,分别用频率50 Hz,EMFs强度为0 mT(对照组)和1.8 mT的正弦波、三角波、方波和锯齿波处理ROB,30 min/(次.天).在磁场处理后4～8天细胞呈现特征样分布.方波促进成骨细胞增殖,正弦波抑制成骨细胞增殖.三角波和正弦波增加ALP活性,其中ALP染色、茜素红钙化结节染色和胶原Ⅰ(collagen-Ⅰ)免疫组织化学检测结果与ALP活性一致.在EMFs处理后的24 h、96 h和72 h后EMFs分别提高Runx-2、Opg和Igf基因表达水平,其中尤以正弦波和三角波作用最为显著.上述结果表明:50 Hz 1.8 mT方波促进成骨细胞增殖,正弦波抑制成骨细胞增殖.50 Hz 1.8 mT EMFs能促进体外培养成骨细胞分化成熟,其中尤以正弦波和三角波促进成骨细胞分化成熟作用最为显著.     

Suppression of heat-induced HSP-70 by simultaneous exposure to 50 mT magnetic field

Effect of extremely low frequency magnetic field (ELFMF) at 50 mT and 60 Hz on heat-induced expression of heat shock protein 70 (hsp-70) was examined in HL60RG cells. No increase in hsp-70 production was observed in the cells after exposure to 50 mT ELFMF alone. Simultaneous exposure to 50 mT ELFMF in combination with mild heat at 42 and 40 degrees C suppressed heat-induced hsp-70 expression. The suppression of hsp-70 occurred when cells were simultaneously exposed to both for longer periods of more than 5 h. However, the suppression of hsp-70 was not observed at a magnetic density of 5 and 0.5 mT. This result suggests that exposure to 50 mT ELFMF may act on a protection against the concomitant mild heat stress in HL60RG cells.     

Effects of 50 Hz magnetic fields on gap junctional intercellular communication.

To explore whether the extremely low frequency (ELF) electromagnetic fields (EMFs) may act as cancer promoters or be synergistic with 12-O-tetradecanoylphorbol-13-acetate (TPA) in cancer promotion, an experiment was conducted on the effects of 50 Hz magnetic fields (MFs) on gap junctional intercellular communication (GJIC) of Chinese hamster lung (CHL) cells. Lucifer dye was loaded into CHL cells by iontophoretic injection, and the number of dye-coupled cells (DCC) 5 min after the injection was adopted as the index of GJIC. The effects of TPA at different concentrations and magnetic fields at different intensities, combined with 5 ng/ml TPA, were studied. The results showed that the suppression of TPA on GJIC was dependent on TPA concentration; the threshold concentration of TPA for CHL cells was between 1 and 5 ng/ml. After exposure to 0.8 mT magnetic field for 24 h, the number of DCC decreased to 6.08 +/- 1.59, whereas the number of DCC in the control group was 9.84 +/- 2.27 (P < .05). When the cells were exposed at 0.2, 0.4, and 0.8 mT for 24 h, combined with 5 ng/ml TPA treatment during the last 1 h, the number of DCC decreased to 5.52 +/- 1.53, 5.00 +/- 1.22, and 4.00 +/- 1.29, respectively, which were significantly lower than the values for the group treated with 5 ng/ml TPA alone (6.38 +/- 1.39). It is suggested that certain intensities of 50 Hz magnetic field might act as cancer promoters, be additive with other promoters in cancer promotion, or both.     

Large granular lymphocytic (LGL) leukemia in rats exposed to intermittent 60 Hz magnetic fields

An animal model for large granular lymphocytic (LGL) leukemia in male Fischer 344 rats was utilized to determine whether magnetic field exposure can be shown to influence the progression of leukemia. We previously reported that exposure to continuous 60 Hz, 1 mT magnetic fields did not significantly alter the clinical progression of LGL leukemia in young male rats following injection of spleen cells from donor leukemic rats. Results presented here extend those studies with the following objectives: (a) to replicate the previous study of continuous 60 Hz magnetic field exposures, but using fewer LGL cells in the inoculum, and (b) to determine if intermittent 60 Hz magnetic fields can alter the clinical progression of leukemia. Rats were randomly assigned to four treatment groups (18/group) as follows: (1) 1 mT (10 G) continuous field, (2) 1 mT intermittent field (off/on at 3 min intervals), (3) ambient controls ( < 0.1 microT), and (4) positive control (5 Gy whole body irradiation from cobalt-60 four days prior to initiation of exposure). All rats were injected intraperitoneally with 2.2 x 10(6) fresh, viable LGL leukemic spleen cells at the beginning of the study. The fields were activated for 20 h per day, 7 days per week, and all exposure conditions were superimposed over the natural ambient magnetic field. The rats were weighed and palpated for splenomegaly weekly. Splenomegaly developed 9-11 weeks after transplantation of the leukemia cells. Hematological evaluations were performed at 6, 8, 10, 12, 14, and 16 weeks of exposure. Peripheral blood hemoglobin concentration, red blood cells, and packed cell volume declined, and total white blood cells and LGL cells increased dramatically in all treatment groups after onset of leukemia. Although the positive control group showed different body weight curves and developed signs of leukemia earlier than other groups, differences were not detected between exposure groups and ambient controls. Furthermore, there were no overall effects of magnetic fields on splenomegaly or survival in exposed animals. In addition, no significant and/or consistent differences were detected in hematological parameters between the magnetic field exposed and the ambient control groups.     

No Evidence of Cellular Alterations by MilliTesla-Level Static and 50 Hz Magnetic Fields on S. cerevisiae

In an attempt to determine whether magnetic field (MF) exposures might induce cellular alterations, S. cerevisiae yeast cells were exposed to static or sinusoidal 50?Hz homogeneous MF (0.35?mT, 1.4?mT, and 2.45?mT) for 1?h and 72?h. Unsynchronized cells grown exponentially while exposed to MF, containing cells in all stages of the mitotic cell cycle. MF was generated by a pair of Helmholtz coils (40?cm in diameter, coaxial, separated by 20?cm). Survival, cell cycle distribution, colony forming ability, and mutation frequency were assayed. No differences in the above-mentioned parameters were observed in MF exposed samples in relation to unexposed controls, suggesting that homogeneous MF at these intensities do not produce appreciable cellular alterations in this organism under typical in vitro growth conditions.     

No evidence of cellular alterations by MilliTesla-level static and 50 Hz magnetic fields on S. cerevisiae

In an attempt to determine whether magnetic field (MF) exposures might induce cellular alterations, S. cerevisiae yeast cells were exposed to static or sinusoidal 50?Hz homogeneous MF (0.35?mT, 1.4?mT, and 2.45?mT) for 1?h and 72?h. Unsynchronized cells grown exponentially while exposed to MF, containing cells in all stages of the mitotic cell cycle. MF was generated by a pair of Helmholtz coils (40?cm in diameter, coaxial, separated by 20?cm). Survival, cell cycle distribution, colony forming ability, and mutation frequency were assayed. No differences in the above-mentioned parameters were observed in MF exposed samples in relation to unexposed controls, suggesting that homogeneous MF at these intensities do not produce appreciable cellular alterations in this organism under typical in vitro growth conditions.     

Exposure of mammalian cells to 60-Hz magnetic or electric fields: analysis for DNA single-strand breaks

Chinese hamster ovary (CHO) cells were exposed for 1 h to 60-Hz magnetic fields (0.1 or 2 mT), electric fields (1 or 38 V/m), or to combined magnetic and electric fields (2 mT and 38 V/m, respectively). Following exposure, the cells were lysed, and the DNA was analyzed for the presence of single-strand breaks (SSB), using the alkaline elution technique. No significant differences in numbers of DNA SSB were detected between exposed and sham-exposed cells. A positive control exposed to X-irradiation sustained SSB with a dose-related frequency. Cells exposed to nitrogen mustard (a known cross-linking agent) and X-irradiation demonstrated that the assay could detect cross-linked DNA under our conditions of electric and magnetic field exposures.     

No effect of exposure to static and sinusoidal magnetic fields on nitric oxide production by macrophages

The effects of exposure to static (1–100 mT) or sinusoidal (1 Hz, 1.6 mT) magnetic fields on the production of nitric oxide (NO) by murine BCG-activated macrophages were investigated. In these cells, the inducible isoform of NO synthase is present. No significant differences were observed in nitrite levels among exposed, sham-exposed, or control macrophages after exposure for 14 h to static fields of 1, 10, 50, and 100 mT and to sinusoidal 1.6 mT, 1 Hz magnetic fields. © 1996 Wiley-Liss, Inc.     

Overexpression of miR-26b-5p regulates the cell cycle by targeting CCND2 in GC-2 cells under exposure to extremely low frequency electromagnetic fields

The increasing prevalence of extremely low frequency electromagnetic fields (ELF-EMFs) exposure has raised considerable public concern regarding the potential hazardous effects of ELF-EMFs on male reproductive function. Increasing evidence indicates that miRNAs are necessary for spermatogenesis and male fertility. However, the regulation of miRNA expression and the roles of miRNAs in response to ELF-EMFs remain unclear. In our study, mouse spermatocyte-derived GC-2 cells were intermittently exposed to a 50 Hz ELF-EMF for 72 h (5 min on/10 min off) at magnetic field intensities of 1 mT, 2 mT and 3 mT. MiR-26b-5p was differentially expressed in response to different magnetic field intensities of ELF-EMFs. The host gene CTDSP1 showed an unmethylation status in GC-2 cells at different magnetic field intensities of ELF-EMF exposure. MiR-26b-5p had no significant, obvious influence on the cell viability, apoptosis or cell cycle of GC-2 cells. However, the overexpression of miR-26b-5p significantly decreased the percentage of G0/G1 phase cells and slightly increased the percentage of S phase cells compared to the sham group that was exposed to a 50 Hz ELF-EMF. Computational algorithms identified Cyclin D2 (CCND2) as a direct target of miR-26b-5p. MiR-26b-5p and a 50 Hz ELF-EMF altered the expression of CCND2 at both the mRNA and protein levels. Overexpressed miR-26b-5p in GC-2 cells can change the mRNA expression of CCND2 following 50 Hz ELF-EMF at 3 mT. These findings demonstrate that miR-26b-5p could serve as a potential biomarker following 50 Hz ELF-EMF exposure, and miR-26b-5p-CCND2-mediated cell cycle regulation might play a pivotal role in the biological effects of ELF-EMFs.     

Three dimensional (3D) analysis of the morphological changes induced by 50 Hz magnetic field exposure on human lymphoblastoid cells (Raji)

Human Raji B lymphoid cells after exposure for 64 h to a 1 mT (rms) 50 Hz sinusoidal magnetic field showed a reorganization of membrane and cytoskeletal components. Atomic force microscopy in air revealed several modifications in 80% of the exposed cells, such as loss of microvilli-like structures followed by progressive appearance of membrane introflections. This change in plasma membrane morphology was also accompanied by a different actin distribution, as detected by phalloidin fluorescence. These observations support our previous hypothesis that electric and magnetic fields may modify the plasma membrane structure.     

Protein kinase C activity following exposure to magnetic field and phorbol ester

We examined the separate and combined effects of 60 Hz sinusoidal magnetic fields (MFs) and a phorbol ester on protein kinase C (PKC) activity in HL60 cells. No enhancement in PKC activity was observed when a cell culture was exposed to a 1.1 mT (rms) MF alone or to a combination of MF and 2 μM phorbol 12-myristate 13-acetate (PMA) for 1 h. In a second set of experiments, cells were preexposed to a less than optimal concentration of PMA (50 nM) for 45 min, followed by a 15 min exposure to both PMA and MF. The data showed a greater decrease in cytosolic PKC activity and a larger increase in membrane activity than was induced by either 1 h PMA treatment alone or PMA and sham MF exposure. One logical conclusion from these data is that MFs may be acting in a synergistic manner on a pathway that has already been activated. Therefore, we suggest that MFs, rather than producing biological effects by a new pathway or mechanism of interaction, exert their effect(s) by interacting with already functioning reactions or pathways. If correct, the question of an MF's mechanism of interaction refocuses on how weak fields might enhance or depress a molecular reaction in progress, rather than on finding a new transduction pathway. Bioelectromagnetics 19:469–476, 1998. © 1998 Wiley-Liss, Inc.     

极低频磁场对人肝癌细胞生长、代谢及细胞周期的影响

目的研究一定参数的极低频磁场对人肝癌细胞(SK-HEP-1)在诸多方面的影响。方法在整个SK-HEP-1细胞的培养周期中用50Hz,20mT的极低频磁场对其进行作用,并检测作用后细胞的增殖活性、生长动力学、代谢以及细胞周期的变化。结果50Hz,20mT的极低频磁场对SK-HEP-1细胞的生长与代谢有抑制作用,并能阻碍其有丝分裂的进行。结论50Hz,20mT的极低频磁场为治疗人类恶性肿瘤提供了一种可能的手段。     

No influence of magnetic fields on cell cycle progression using conditions relevant for patients during MRI

The purpose of this study was to examine whether exposure to magnetic fields (MFs) relevant for magnetic resonance imaging (MRI) in clinical routine influences cell cycle progression in two tumor cell lines in vitro. HL60 and EA2 cells were exposed to four types of MFs: (i) static MF of 1.5 and 7.05 T, (ii) extremely low frequency magnetic gradient fields (ELFMGFs) with +/- 10 mT/m and 100 Hz, as well as +/- 100 mT/m and 100 Hz, (iii) pulsed high frequency MF in the radiofrequency (RF) range (63.6 MHz, 5.8 microT), and (iv) a combination of (i-iii). Exposure periods ranged from 1 to 24 h. Cell cycle distribution (G(0)/G(1), S, and G(2)/M phases) was analyzed by flow cytometry. Cell cycle analysis did not reveal differences between the exposed and the control cells. As expected, positive controls with irradiated (8 Gy) HL60 and EA2 cells showed a strong G(2)/M arrest. Using conditions that are relevant for patients during MRI, no influence of MFs on cell cycle progression was observed in these cell lines. Care was taken to control secondary parameters of influence, such as vibration by the MR scanner or temperature to avoid false positive results.     

Alteration in cellular functions in mouse macrophages after exposure to 50 Hz magnetic fields

The aim of the present study is to investigate whether extremely low frequency electromagnetic fields (ELF-EMF) affect certain cellular functions and immunologic parameters of mouse macrophages. In this study, the influence of 50 Hz magnetic fields (MF) at 1.0 mT was investigated on the phagocytic activity and on the interleukin-1beta (IL-1beta) production in differentiated macrophages. MF-exposure led to an increased phagocytic activity after 45 min, shown as a 1.6-fold increased uptake of latex beads in MF-exposed cells compared to controls. We also demonstrate an increased IL-1beta release in macrophages after 24 h exposure (1.0 mT MF). Time-dependent IL-1beta formation was significantly increased already after 4 h and reached a maximum of 12.3-fold increase after 24 h compared to controls. Another aspect of this study was to examine the genotoxic capacity of 1.0 mT MF by analyzing the micronucleus (MN) formation in long-term (12, 24, and 48 h) exposed macrophages. Our data show no significant differences in MN formation or irregular mitotic activities in exposed cells. Furthermore, the effects of different flux densities (ranging from 0.05 up to 1.0 mT for 45 min) of 50 Hz MF was tested on free radical formation as an endpoint of cell activation in mouse macrophage precursor cells. All tested flux densities significantly stimulated the formation of free radicals. Here, we demonstrate the capacity of ELF-EMF to stimulate physiological cell functions in mouse macrophages shown by the significantly elevated phagocytic activity, free radical release, and IL-1beta production suggesting the cell activation capacity of ELF-EMF in the absence of any genotoxic effects.     

Extremely low frequency electromagnetic fields (ELF-EMFs) induce in vitro angiogenesis process in human endothelial cells

Effects of extremely low frequency (ELF) electromagnetic fields (EMFs) on activation of angiogenesis were analysed using cultured umbilical human vein endothelial cells (HUVECs). The cultures were exposed to a sinusoidal EMF to intensity of 1 mT, 50 Hz for up to 12 h. EMFs increased the degree of endothelial cell proliferation and tubule formation, coupled by an acceleration in the process of wound healing. Since this process is physiologically accompanied by a large modification in the structural organization of actin and focal adhesions, we analyzed the rearrangement of some cytoskeleton elements demonstrating a major reorganization of the fibres and of the focal adhesion complexes after EMF exposure. Finally, Western blot analysis revealed a significant increase in phosphorylation as well as the overall expression of VEGF receptor 2 (KDR/Flk-1) suggesting that EMFs may modulate in vitro some endothelial functions correlated to angiogenesis through signal transduction pathways dependent on VEGF.