Enhanced NOR-1 gene expression by exposure of Chinese hamster cells to high-density 50 Hz magnetic fields

Enhanced expression of neuron derived orphan receptor (NOR-1) gene was observed by exposure of Chinese hamster ovary K1 (CHO-K1) cells to an extremely low frequency magnetic field (ELFMF) of 50 Hz at 400 mT, but not at 5 mT. The enhanced expression, reaching the maximum at 6 h, was transient and reduced to the control level after exposure to 400 mT ELFMF for 24 h. The NOR-1 expression induced by treatment with forskolin and TPA was further enhanced by the simultaneous treatment with 400 mT ELFMF, in which the maximum response was at 3 h. The NOR-1 expression by these treatments was induced more earlier than that by 400 mT ELFMF alone. When cells were treated with an inhibitor of the protein kinase C (calphostin C or crocetin) and Ca²⁺ entry blockers (nifedipin and dantrolen) during the 400 mT ELFMF exposure, the enhanced NOR-1 expression was not observed. Exposure of CHO-K1 cells to the high-density 400 mT ELFMF may affect the signal transduction in the cells, resulting in the enhanced NOR-1 gene expression.     

Effect of high-density extremely low frequency magnetic field on sister chromatid exchanges in mouse m5S cells

The induction of sister chromatid exchanges (SCEs) was evaluated in the cultured mouse m5S cells after exposure to extremely low frequency magnetic field (ELFMF; 5, 50 and 400 mT). Exposure to 5 mT and 50 mT ELFMF led to a very small increase in the frequency of SCEs, but no significant difference was observed between exposed and unexposed control cells. The cells exposed to 400 mT ELFMF exhibited a significant elevation of the SCE frequencies. There was no significant difference between data from treatments with mitomycin-C (MMC) alone and from combined treatments of MMC plus ELFMF (400 mT) at any MMC concentrations from 4 to 40 nM. These results suggest that exposure to highest-density ELFMF of 400 mT may induce DNA damage, resulting in an elevation of the SCE frequencies. We suppose that there may be a threshold for the elevation of the SCE frequencies, that is at least over the magnetic density of 50 mT.     

Exposure to extremely low frequency magnetic fields suppresses x-ray-induced transformation in mouse C3H10T1/2 cells

We designed and manufactured equipment for exposure of cultured cells to extremely low frequency magnetic fields (ELFMF) at 5, 50, and 400 mT and examined the effect of ELFMF on cellular transformation in mouse C3H10T1/2 cells (clone 8). Transformed foci, Type II and Type III, were independently counted as transformants. The cells were exposed to ELFMF alone at 5, 50, and 400 mT for 24 h or X-irradiated with 3 Gy followed by the ELFMF exposure. No significant difference in the transformation was observed between sham-exposed control and the ELFMF exposure from 5 to 400 mT. The transformation frequency for X-rays plus ELFMF was decreasing compared with X-rays alone. When 12-O-tetra-decanoylphorbol-13-acetate (TPA) was contained in the medium throughout the experiment, the transformation frequency by X-rays alone was elevated more. In the combined treatment with X-rays followed by ELFMF, the transformation frequency was slightly decreased at 50 and 400 mT even in the medium containing TPA. The long-term exposure at 5 mT suppressed both spontaneous and X-ray-induced transformations significantly. It is well known that overexpressing protein kinase C (PKC) failed to yield identifiable transformation of foci induced by ionizing radiation. We demonstrated previously that exposure to high-density ELFMF induced expression of several genes through an increase in PKC activity. From these results, it is suggested that ELFMF might suppress X-ray-induced transformation through activation of PKC by ELFMF.     

Effect of high-density extremely low frequency magnetic field on sister chromatid exchanges in mouse m5S cells

The induction of sister chromatid exchanges (SCEs) was evaluated in the cultured mouse m5S cells after exposure to extremely low frequency magnetic field (ELFMF; 5, 50 and 400 mT). Exposure to 5 mT and 50 mT ELFMF led to a very small increase in the frequency of SCEs, but no significant difference was observed between exposed and unexposed control cells. The cells exposed to 400 mT ELFMF exhibited a significant elevation of the SCE frequencies. There was no significant difference between data from treatments with mitomycin-C (MMC) alone and from combined treatments of MMC plus ELFMF (400 mT) at any MMC concentrations from 4 to 40 nM. These results suggest that exposure to highest-density ELFMF of 400 mT may induce DNA damage, resulting in an elevation of the SCE frequencies. We suppose that there may be a threshold for the elevation of the SCE frequencies, that is at least over the magnetic density of 50 mT.     

Increase in X-ray-induced mutations by exposure to magnetic field (60 Hz, 5 mT) in NF-kappaB-inhibited cells

It is established that extremely low frequency magnetic fields (ELFMF) at the flux densities, i.e., 5 mT and less, are not mutagenic. However, exposure to ELFMF enhances mutations induced by X-rays. In this study, we examined the effects of long-term exposure to 5 mT ELFMF on mutation induction and X-ray-induced mutations in human malignant glioma cells (MO54) with different mutant IkappaB-alpha (a critical inhibitor of NF-kappaB) genes. Cells were exposed or sham-exposed to 5 mT ELFMF for up to 8 days with or without initial X-rays (4 Gy), and the mutant frequency of hypoxanthine-guanine phosphoribosyl transferase (HPRT) gene was analyzed. An obvious increase in X-ray-induced mutations was observed after treatment with ELFMF in combination with X-irradiation in MO54 cells with tyrosine mutant IkappaB-alpha gene other than with serine mutant IkappaB-alpha gene or vector alone. Exposure to ELFMF alone increased mutations significantly in MO54 cells with tyrosine mutant IkappaB-alpha gene. In addition, X-ray-induced apoptoic cells were increased in MO54-V cells after exposure to ELFMF, while an anti-apoptotic effect of magnetic field was found in MO54-SY4 cells. Our data suggest that exposure to 5 mT ELFMF may induce mutations and enhance X-ray-induced mutations, resulting from the inactivation of NF-kappaB through the inhibition of tyrosine phosphorylation.     

An extremely low frequency magnetic field attenuates insulin secretion from the insulinoma cell line, RIN-m

In this study, we investigated the effects of exposure to an extremely low frequency magnetic field (ELFMF) on hormone secretion from an islet derived insulinoma cell line, RIN-m. We stimulated RIN-m cells to secrete insulin under exposure to an ELFMF, using our established system for the exposure of cultured cells to an ELFMF at 5 mT and 60 Hz, or under sham exposure conditions for 1 h and observed the effects. In the presence of a depolarizing concentration of potassium (45 mM KCl), exposure to ELFMF significantly attenuated insulin release from RIN-m cells, compared to sham exposed cells. Treatment with nifedipine reduced the difference in insulin secretion between cells exposed to an ELFMF and sham exposed cells. The expression of mRNA encoding synaptosomal associated protein of 25 kDa (SNAP-25) and synaptotagmin 1, which play a role in exocytosis in hormone secretion and influx of calcium ions, decreased with exposure to an ELFMF in the presence of 45 mM KCl. These results suggest that exposure to ELFMF attenuates insulin secretion from RIN-m cells by affecting calcium influx through calcium channels.     

Decrease in glucose-stimulated insulin secretion following exposure to magnetic fields

We evaluated the effects of extremely low frequency magnetic field (ELFMF) on glucose-stimulated insulin secretion from HIT-T15 cells and investigated the mechanisms of these effects. We demonstrated that exposure to ELFMF at 5mT decreased glucose-stimulated insulin secretion by preventing the increases in cellular adenosine 5'-triphosphate/adenosine 5'-diphosphate, membrane depolarization, and cytosolic free calcium ion concentration. The glucose-induced upregulation of insulin mRNA expression was also attenuated by exposure to ELFMF, although cell viability was not affected. These findings demonstrate the potential of exposure to ELFMF for clinical use as a novel inhibitory method of insulin secretion.     

Exposure to extremely low frequency magnetic fields affects insulin-secreting cells

To evaluate the effects of extremely low frequency magnetic field (ELFMF) on beta-cell survival and function, we cultured a hamster-derived insulin-secreting cell line (HIT-T15), which exhibits responsiveness to glucose in a semi-physiological range, under exposure to sham and ELFMF conditions, and assessed cell survival and function. We used our previously developed ELFMF exposure unit (a sinusoidal magnetic field at a frequency of 60 Hz, 5 mT) to culture cells under exposure to ELFMF conditions. We found that exposure to ELFMF for 5 days in the absence of glucose increased cell number, exposure for 2 days in the absence of glucose and for 5 days with 100 mg/dl glucose increased the insulin secretion to the culture medium, and exposure for 2 and 5 days with 40 and 100 mg/dl glucose increased intracellular insulin concentration in HIT-T15 cells. The increase in cell number under apoptotic culture conditions by exposure to ELFMF could lead to new therapeutic concepts in the treatment of diabetes. The ELFMF-induced increase in intracellular insulin concentration could be utilized to develop culture conditions to enhance intracellular insulin concentration in insulin-secreting cells that would be useful for cell transplantation to cure diabetes mellitus.     

Induction of stress proteins by electromagnetic fields in cultured HL-60 cells.

HL-60 cells in culture were exposed for 2 h to a sinusoidal 0.1 or 1 mT (1 or 10 Gauss) magnetic field at 60 Hz and pulse labeled after exposure with radioactive isotopes by incubation by using either [(35)S]methionine, [(3)H]leucine, or [(33)P]phosphate. The radioactive labels were incorporated into cellular proteins through synthesis or phosphorylation. Proteins were extracted from electrostatically sorted nuclei, and the heat shock/stress proteins (sp) were analyzed for synthesis and phosphorylation by two-dimensional polyacrylamide gel electrophoresis. In the control cultures (no exposure to the magnetic field), sp 72c (cognate form) was faintly observed. A 0.1 mT exposure did not show sp metabolism to be different from that of the controls; however, after a 1 mT exposure of the HL-60 cells, sp 70i (inducible form) was synthesized ([(35)S]methionine incorporation). Sp 90 was not synthesized at either field level, but was phosphorylated ([(33)P]phosphate incorporation) in the 1 mT exposure. Sp 27 (isoforms a and b) was induced after a 1 mT exposure as reflected by labeling with [(3)H]leucine. These sps were not detected after a 0.1 mT exposure. After a 1 mT exposure and labeling with [(33)P], sp 27 isoforms b and c were phosphorylated whereas isoform 'a' was not observed. Sps 70i, 72c, and 90 were identified by commercial sp antibodies. Likewise, polypeptides a, b, and c were verified as sp 27 isoforms by Western blotting. Statistical evaluation of sp areas and densities, determined from fluorographs by Western-blot analysis, revealed a significant increase in sps 90 and 27a after a 1 mT magnetic field exposure. The 1 mT magnetic field interacts at the cellular level to induce a variety of sp species. Bioelectromagnetics 20:347-357, 1999. Published 1999 Wiley-Liss, Inc.     

Extremely low frequency magnetic fields suppress the reduction of germination rate of Arabidopsis thaliana seeds kept in saturated humidity.

The effects of an extremely low frequency magnetic field (ELFMF) on the germination of plant seeds were examined. The decrease in the germination activity of the seeds of Arabidopsis thaliana WS kept in saturated humidity and high temperature (37 degrees C) was suppressed by the exposure to a 400 mT ELFMF.     

Extremely Low Frequency Magnetic Fields Suppress the Reduction of Germination Rate of Arabidopsis thaliana Seeds Kept in Saturated Humidity

The effects of an extremely low frequency magnetic field (ELFMF) on the germination of plant seeds were examined. The decrease in the germination activity of the seeds of Arabidopsis thaliana WS kept in saturated humidity and high temperature (37°C) was suppressed by the exposure to a 400 mT ELFMF.     

The effect of 100 mT SMF on activation of the hsp70 promoter in a heat shock/luciferase reporter system

Human exposure to magnetic fields, increased through use of new technologies like magnetic resonance imaging (MRI), has prompted investigations into possible effects of static magnetic fields (SMFs) on cellular processes. However, controversy still remains between many studies, which likely results from a lack of uniformity across experimental parameters, including the length of magnetic field exposure, the strength of the magnetic field, and the cell type or organism under investigation. The purpose of this research was to monitor effects of SMF exposure using real‐time luminescence photometry. The study investigated the potential interaction of a 100 mT SMF on a heat shock protein (hsp70)/luciferase reporter construct in stably transfected NIH3T3 cells. Changes in heat shock promoter activation following 100 mT SMF exposure were analyzed and detected as bioluminescence in real‐time. Two heat parameters were considered in combination with sham‐ and 100 mT‐exposed experiments: no heat or 1,800 s heat. As expected, there was a significant increase in bioluminescence in response to 1,800 s of heat alone. However, no significant difference in average hsp70 promoter activation between sham and 100 mT experiments was observed for no heat or 1,800 s heat experiments. Therefore, a 100 mT SMF was shown to have no effect on the activation of the heat shock protein promoter during SMF exposure or when SMF exposure was combined with a heat insult. J. Cell. Biochem. 108: 956–962, 2009. © 2009 Wiley‐Liss, Inc.     

Salicylic acid-mediated potentiation of Hsp70 induction correlates with reduced apoptosis in tobacco protoplasts.

BACKGROUND: Elevated temperatures jeopardize plant disease resistance, as mediated by salicylic acid (SA). SA potentiates heat-induced expression of the 70-kDa heat shock protein (Hsp70) in tomato cells. In mammalian cells, Hsp70 suppresses apoptosis. We hypothesized that potentiation of heat-induced Hsp70 by SA contributes to a reduction in apoptosis in tobacco protoplasts. METHODS: Tobacco protoplasts (Nicotiana tabacum) were exposed to SA (70 microM) at normal temperatures or in combination with heat shock. Hsp70/Hsc70 accumulation and phosphatidylserine (PS) exposure, DNA fragmentation, as well as loss of mitochondrial membrane potential were quantified by flow cytometry. RESULTS AND CONCLUSIONS: SA at normal temperatures did not influence Hsp70/Hsc70 accumulation, but were found to induce apoptosis. In contrast, SA in combination with HS potentiated heat-induced Hsp70/Hsc70 accumulation in tobacco protoplasts that correlated negatively with apoptosis, illustrated by decreased PS exposure and DNA fragmentation and enhanced mitochondrial membrane potential. We propose that this correlation supports a possible role for apoptosis suppression by Hsp70 under elevated temperatures during pathogen infection.     

ELF electromagnetic fields increase hydrogen peroxide (H2O2)-induced mutations in pTN89 plasmids

We have examined the mutational effects of hydrogen peroxide (H(2)O(2)) in the presence and absence of an extremely low-frequency magnetic field (ELFMF), using pTN89 plasmids. Mutations were detected in the supF gene carried by these plasmids in Escherichia coli. The plasmids were either treated with H(2)O(2) (1microM) alone at 37 degrees C for 4h, or were exposed to an ELFMF (60Hz, 5millitesla (mT)) simultaneously with H(2)O(2) treatment. The mutation frequency was 2.28 x 10(-4) for H(2)O(2) treatment alone, and 5.81 x 10(-4) for ELFMF exposure with H(2)O(2) treatment. We did not observe any mutations using treatment with ELFMF exposure alone. This indicates that the ELFMF may potentiate H(2)O(2)-induced mutation. Sequence analysis of the supF mutant plasmids revealed that base substitutions, G: C-->A :T transitions and G:C-->T:A transversions were dominant in both treatment groups, and there was no difference in the mutation spectrum or the hotspots between the groups. Therefore, ELFMFs may interact and potentiate the damage induced by H(2)O(2), resulting in an increase in the number of mutations.