No effects of extremely low frequency magnetic fields found on cytotoxic activities and cytokine production of human peripheral blood mononuclear cells in vitro

Several epidemiologic studies have suggested an association between exposure to extremely low frequency (ELF) magnetic fields (MFs) and cancer in adults and children. A possible target of MFs is the immune system. The effects of the exposure to ELF MFs on the immunological functions of human peripheral blood mononuclear cells (PBMCs) obtained from healthy male volunteers were assessed by measuring the natural killer (NK) and lymphokine activated killer (LAK) activities and the production of interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), interleukin-2 (IL-2), and interleukin-10 (IL-10). The PBMCs were exposed to three different MF: linearly polarized (vertical), circularly polarized, and elliptically polarized, at 50 and 60 Hz. Magnetic flux densities were set at 500, 100, 20, and 2 microT (rms) for vertical field and at 500 microT (rms) for the rotating fields. Using cytotoxicity assay and enzyme-linked immunosorbent assay (ELISA) for cytokine production, we could not find any effects of ELF MFs on the cytotoxic activities and the cytokines production of human PBMCs.     

Development and evaluation of intermediate frequency magnetic field exposure system for studies of in vitro biological effects

We have developed an intermediate frequency (IF) magnetic field exposure system for in vitro studies. Since there are no previous studies on exposure to heating-frequency magnetic fields generated from an induction heating (IH) cook top, there is a strong need for such an exposure system and for biological studies of IF magnetic fields. This system mainly consists of a magnetic-field-generating coil housed inside an incubator, inside which cultured cells can be exposed to magnetic field. Two systems were prepared to allow the experiment to be conducted in a double-blind manner. The level of the generated magnetic field was set to 532 microT rms in the exposure space, 23 kHz, 80 times the value in the International Commission on Non-ionizing Radiation Protection (ICNIRP) guidelines, with a spatial field uniformity better than 3.8%. The waveforms were nearly sinusoidal. It was also confirmed that the parasitic electric field was 157 V/m rms and the induced electric field was 1.9 V/m rms. The temperature was maintained at 36.5 +/- 0.5 degrees C for 2 h. Furthermore, leaked magnetic flux density was 0.7 microT rms or lower at extremely low frequency (ELF) and IF in the stopped system when the other system was being operated, and the environmental magnetic flux density was 0.1 microT rms or lower at the center of the coils. As a result, it was confirmed that this system could be successfully used to evaluate the biological effects of exposure to IF magnetic fields.     

The influence of 1.2 microT, 60 Hz magnetic fields on melatonin- and tamoxifen-induced inhibition of MCF-7 cell growth

We independently examined the findings of Harland and Liburdy, who reported that 1.2 microT(rms), 60 Hz magnetic fields could significantly reduce the inhibitory action of physiological levels of melatonin (10(-9) M) and of pharmacological levels of tamoxifen (10(-7) M) on the growth of MCF-7 human breast cancer cells in vitro. We used two testing protocols. In the melatonin study, the cell numbers per dish on day 7 of treatment were determined using a hemocytometer assay. In the tamoxifen study we used an expanded protocol, employing an alternative cell counting assay to characterize the cell numbers per dish on days 4, 5, 6, and 7. In both the melatonin and tamoxifen studies, cells were plated on 35 mm dishes and placed in each of two exposure chambers inside 5% CO(2) incubators. One exposure chamber was energized to produce 1.2 microT(rms), 60 Hz magnetic fields and the other chamber was not energized. Treatment was continuous until assays were performed. Cells were harvested at selected times, and enumerated without knowledge of treatment. In the melatonin study, the experiment was repeated three times, whereas in the tamoxifen study, each experiment was repeated nine times. In the melatonin study, cell numbers per dish were significantly reduced (by 16.7%) in the melatonin treated cultures after 7 days of incubation compared to control cultures, whereas in the presence of 1.2 microT(rms), 60 Hz magnetic fields, the melatonin treated cultures had the same cell populations as the control cultures. In the tamoxifen study, tamoxifen reduced the cell growth by 18.6 and 25% on days 6 and 7, respectively, in the chamber not energized, while in 1.2 microT(rms), 60 Hz fields, tamoxifen reduced the cell growth only by 8.7 and 13.1%, respectively. These results are consistent with those reported by Harland and Liburdy. A critical element of this successful replication effort was the constructive communication established and maintained with the original investigators. Bioelectromagnetics 22:122-128, 2001. Published 2001 Wiley-Liss, Inc.     

Sinusoidal 50 Hz magnetic fields do not affect structural morphology and proliferation of human cells in vitro

The effects of electromagnetic fields on several processes related to cell physiology and proliferation are currently being investigated. Although the results are still not conclusive and even conflicting, there seems to be a fairly good agreement on the early effects of electromagnetic fields on the generation of free radicals and on Ca++-intracellular concentration and transport. To evaluate the long-lasting consequences of these precocious events, we examined the effects of short- and long-term magnetic field exposure on structural organization (cytokeratin or actin detection), proliferation (bromodeoxyuridine incorporation and propidium iodide staining), colony forming ability and viability (trypan blue exclusion test) of highly proliferating MCF-7 cells (from human breast carcinoma) and on slowly proliferating normal human fibroblasts (from healthy donors). Cells were exposed to either 20 or 500 microT sinusoidally oscillating (50Hz) magnetic fields for different lengths of time (1 to 4 days). Short (1 day)- and long (4 days)-time exposure to the two intensities did not affect MCF-7 growth and viability, colony number and size, or cellular distribution along the cell cycle; neither were the cell morphology and the intracellular distribution and amount of cytokeratin modified. Similarly, no modifications in the actin distribution and proliferative potential were observed in normal human fibroblasts. These findings suggest that under our experimental conditions, continuous exposure to magnetic fields does not result in any appreciable effect in both normal and tumor cells in vitro.     

Exposure to ELF magnetic field tuned to Zn inhibits growth of cancer cells

The effects of ELF alternating magnetic fields tuned to Zn(2+) on the growth of cancer cells with different status of p53 were investigated using a cell proliferation assay. Human cancer cells HeLa (cervix cancer, p53(+/+)), Saos-2 and Saos-2-His-273 (osteosarcoma, p53(-/-) and p53 His-273 mutant, respectively), H1299tTA and H1299tTA-His175 (lung carcinoma, p53(-/-) and p53 His-175 mutant), and normal human fibroblasts VH-10 (p53(+/+)) were used. Exposure parameters were calculated for the first harmonic of Zn(2+) based either on the magnetic parametric resonance (MPR) model of Lednev or the ion parametric resonance (IPR) model of Blanchard and Blackman. ELF exposure was for 72 and 96 h. The vertical alternating field was 20 Hz at amplitudes of either 38.7 or 77.4 microT (peaks, IPR or MPR, respectively). The vertical static magnetic field was 43 microT, and the horizontal static magnetic field was zeroed. Treatments of cells with PRIMA-1 and gamma-rays were used as positive controls. Growth inhibition was observed in cells after exposure to ELF at 38.7 microT. Inhibition of HeLa, VH-10, and Saos-2-His-273 cells was statistically significant, P=0.0003, 0.02, and 0.006, respectively. No consistent ELF effects following exposure 77.4 microT were seen. PRIMA-1 inhibited the growth of all cell lines with the strongest effect in mutant p53-carrying cell line H1299tTA-His175. The effects of gamma-rays were relatively weak, suggesting that the cell proliferation assay under conditions employed in this study is not very sensitive to apoptosis. In conclusion, ELF under conditions of exposure tuned to Zn(2+) according to the IPR model inhibited the growth of cancer and normal cells. No clear relationship of the observed growth inhibition to p53 status was found. Further experiments, using complementary techniques, are required to test whether p53 reactivation by ELF is feasible.     

Effects of combined DC and AC magnetic fields on germination of hornwort seeds

Seeds of hornwort (Cryptotaenia japonica Hassk) were exposed to sinusoidally time-varying extremely low frequency (ELF) magnetic fields (AC fields) in combination with the local geomagnetic field (DC field). Exposure lasted 24 h/day for 16 days. Three directions of the AC magnetic fields were considered; the vertical (magnetic flux density B ACV, the directions parallel B ACparallel), and perpendicular B ACperpendicular to the direction of total geomagnetic field (magnetic flux density BG) in the geomagnetic plane (GP). Controls consisted of seeds exposed to zero AC magnetic fields in combination with the DC magnetic field. The B ACV in combination with BG effectively promoted the germination of hornwort seeds when applied at 750 microT (RMS) at 7 Hz or 500 microT (RMS) at 14 Hz from among the cases of individual frequencies f = 3.5, 7.0, 10.5, 14.0 Hz at 500 and 750 microT. The B ACparallel promoted the germination of hornwort seeds more effectively than the B ACperpendicular in combination with BG when 500 and 750 microT at 7 Hz were applied.     

Developmental toxicity evaluation of ELF magnetic fields in Sprague-Dawley rats

To identify possible effects of horizontally polarized magnetic field (MF) exposure on maintenance of pregnancy and embryo-fetal development, an MF exposure system was designed and constructed and 96 time-mated female Sprague-Dawley (SD) rats (24/group) received continuous exposure to 60 Hz MF at field strengths of 0 (sham control) and 5, 83.3, or 500 microT (50, 833, or 5000 mG). Dams received MF or sham exposures for 22 h/day on gestational day 6-20. MF was monitored continuously throughout the study. There were no evidences of maternal toxicity or developmental toxicity in any MF exposed groups. Mean maternal body weight, organ weights, and hematological and serum biochemical parameters in groups exposed to MF did not differ from those in sham control. No exposure related differences in fetal deaths, fetal body weight, and placental weight were observed between MF exposed groups and sham control. External, visceral, and skeletal examination of fetuses demonstrated no significant differences in the incidence of fetal malformations between MF exposed and sham control groups. In conclusion, exposure of pregnant rats to 60 Hz at MF strengths up to 500 microT during gestation day 6-20 did not produce any biologically significant effect in either dams or fetuses.     

Investigation of protein expression in magnetic field-treated human glioma cells

We previously reported phenotypic changes in human breast cancer cells following low-level magnetic field (MF) exposure. Here proteomic methods were used to investigate the biochemical effect of MF exposure in SF767 human glioma cells. Protein alterations were studied after exposure to 1.2 microTesla (microT) MF [12 milliGauss (mG), 60 Hertz (Hz)] +/- epidermal growth factor (EGF). SF767 cells were exposed for 3 h to sham conditions (<0.2 microT ambient field strength) or 1.2 microT MF (+/-EGF; 10 ng/ml). Solubilized protein fractions (sham; 1.2 microT; sham + EGF; 1.2 microT + EGF) were loaded for electrophoresis by 2D-PAGE and stained using a colloidal Coomassie blue technique to resolve and characterize the proteins. Protein patterns were compared across groups via Student's t-test using PDQUEST software. Cell profiles revealed significant alterations in the spot density of a subset of treated cells. Automated spot excision and processing was performed prior to peptide mass fingerprinting proteins of interest. Fifty-seven proteins from the detectable pool were identified and/or found to differ significantly across treatment groups. The mean abundance of 10 identified proteins was altered following 1.2 microT exposure. In the presence of EGF six proteins were altered after low magnetic field treatment by increasing (4) or decreasing (2) in abundance. The results suggest that the analysis of differentially expressed proteins in SF767 cells may be useful as biomarkers for biological changes caused by exposure to magnetic fields.     

Effects of 50 Hz magnetic field on cell cycle kinetics and the colony forming ability of budding yeast exposed to ultraviolet radiation.

To investigate the effects of extremely low frequency magnetic fields on ultraviolet radiation (UV) exposed budding yeast, haploid yeast (Saccharomyces cerevisiae) cells of the strain SEy2101a were exposed to 50 Hz sine wave magnetic field (MF) of 120 microT with simultaneous exposure to UV radiation. Most of the UV energy was in the UVB range (280-320 nm). The biologically weighted (CIE action spectrum) dose level for the UV radiation was 175 J/m2. We examined whether 50 Hz MF affected the ability of UV irradiated yeast cells to form colonies (Colony Forming Units, CFUs). In addition, the effect of coexposure on cell cycle kinetics was investigated. Although the significant effect of MF on the cell cycle phases of UV exposed yeast cells was seen only at one time point, the overall results showed that MF exposure may influence the cell cycle kinetics at the first cycle after UV irradiation. The effect of our particular MF exposure on the colony forming ability of the UV irradiated yeast cells was statistically significant 420 min after UV irradiation. Moreover, at 240, 360, and 420 min after UV irradiation, there were fewer CFUs in every experiment in (UV+MF) exposed populations than in only UV exposed yeast populations. These results could indicate that MF exposure in conjunction with UV may have some effects on yeast cell survival or growth.     

Effects of a 60 Hz magnetic field on photosynthetic CO2 uptake and early growth of radish seedlings

Photosynthetic CO2 uptake rate and early growth parameters of radish Raphanus sativus L. seedlings exposed to an extremely low frequency magnetic field (ELF MF) were investigated. Radish seedlings were exposed to a 60 Hz, 50 microT(rms) (root mean square) sinusoidal magnetic field (MF) and a parallel 48 microT static MF for 6 or 15 d immediately after germination. Control seedlings were exposed to the ambient MF but not the ELF MF. The CO2 uptake rate of ELF MF exposed seedlings on day 5 and later was lower than that of the control seedlings. The dry weight and the cotyledon area of ELF MF exposed seedlings on day 6 and the fresh weight, the dry weight and the leaf area of ELF MF exposed seedlings on day 15 were significantly lower than those of the control seedlings, respectively. In another experiment, radish seedlings were grown without ELF MF exposure for 14 d immediately after germination, and then exposed to the ELF MF for about 2 h, and the photosynthetic CO2 uptake rate was measured during the short-term ELF MF exposure. The CO2 uptake rate of the same seedlings was subsequently measured in the ambient MF (control) without the ELF MF. There was no difference in the CO2 uptake rate of seedlings exposed to the ELF MF or the ambient MF. These results indicate that continuous exposure to 60 Hz, 50 microT(rms) sinusoidal MF with a parallel 48 microT static MF affects the early growth of radish seedlings, but the effect is not so severe that modification of photosynthetic CO2 uptake can observed during short-term MF exposure.     

Effects of information and 50 Hz magnetic fields on cognitive performance and reported symptoms

The aim of this study was to explore the role of expectancies and beliefs about the potential effects of electromagnetic fields (EMFs) (what the subject thought the effect was going to be) and the effects of 50 Hz magnetic fields (400 microT(rms)) acute exposure on cognitive performance, the reporting of physical symptoms and some psychological and physiological parameters. Seventy-four healthy male volunteers aged between 40 and 60 years of age were randomly assigned to one of five groups, which differed in (1) the type of information they were given concerning the expected magnetic field effect on performance in cognitive tests (positive = enhancement of the performance; negative = impairment of the performance; neutral) and (2) the type of exposure (real or sham). Three groups were sham exposed with positive (group+), negative (group-) and neutral information (group+/-); one group was really exposed with neutral information (group expo) and one group was not exposed, though they wore the helmet, and did not receive any field-related information (control group). All the volunteers, except the control group, were led to believe that they would be exposed to a magnetic field of 400 microT(rms). The experimental design respected a double blind procedure and the experimental session involved three steps (pre-testing, exposure, and post-testing). Various measurements were taken, including cognitive performance, psychological parameters such as mood, vigilance, and reporting of symptoms. Physiological parameters such as blood pressure and pulse rate were also recorded. The information given did not significantly modify beliefs. No significant difference was found among the five groups depending on the type of information and the type of exposure in cognitive performance, psychological and physiological parameters. In the context of the study, with our population, the type of information given failed to induce expected changes in parameters measured. Our results do not support the hypothesis that an acute exposure to extremely low frequency magnetic fields (50 Hz, 400 microT(rms)) affects the parameters measured.     

A magnetic field exposure facility for evaluation of animal carcinogenicity

Several animal studies have been carried out at the Institut Armand Frappier (IAF) to determine whether chronic exposure to 60 Hz linearly polarized sinusoidal magnetic fields might increase the risk of cancer development of female Fisher rats. The magnetic field exposure facility was developed to meet the requirements of the study protocol for chronic exposure of large number of animals to field intensities of sham < 0.2 microT, 2 microT, 20 microT, 200 microT, and 2000 microT. At each exposure level, including sham, the animals are distributed in a group of four exposure units. Each exposure unit contains two exposure volumes having uniform distribution of magnetic fields for the animals, while the magnetic field external to the unit falls off rapidly due to the "figure-eight" coil topography used. A program of "shake down" tests, followed by verification and calibration of the exposure facility, was carried out prior to starting the animal experiments. Continuous monitoring of the magnetic field and other environmental parameters was an important part in the overall quality assurance program adopted.     

Effect of low-intensity magnetic fields on the development of satellite muscle cells of a newborn rat in the primary culture

The influence of Earth magnetic field shielded down to 0.3 microT and static magnetic field (60-160 microT) on the proliferation and differentiation of satellite muscle cells in the primary culture has been investigated. A stimulatory effect of static magnetic fields on the rate of the formation of massive multinucleated myotubes and an increase in the intracellular calcium concentration ([Ca2+]i) have been detected for magnetic fields of the microtesla range. On the other hand, it was shown that the reduction of earth magnetic fields to 0.3 microT leads to the inhibition of proliferation and differentiation of skeletal muscle cells in the primary culture. Since the formation of contractile myotubes during in vitro experiments is similar to the regeneration of skeletal muscle fibers under muscle damage in vivo, it may be concluded that weak magnetic fields have a strong effect on intracellular processes by influencing all phases of muscle fiber formation. It is necessary to take this fact into consideration when forecasting probable complications of skeletal muscle regeneration during long-term exposure of man to low-intensity magnetic fields and also for the potential use of low static magnetic fields as a tool to recover the affected myogenesis.     

50 Hz magnetic field exposure alters onset of S-phase in normal human fibroblasts.

This study was undertaken to investigate whether power frequency magnetic fields can affect the kinetics of cell cycle progression in exposed human cells. To achieve this, cultures of normal human fibroblasts were synchronised in the G(0) phase of the cell cycle and exposed to 50 Hz magnetic fields at a range of flux densities. Progression through the cycle was monitored by examining the timing of entry into S phase, as characterised by the onset of DNA synthesis. Simultaneous positive controls were exposed to human recombinant fibroblast growth factor to demonstrate that the system was responsive to external stimuli. Exposure to magnetic fields at 20 and 200 microT induced a small but significant increase in the length of the G(1) phase of the cell cycle. However, exposure at higher flux densities of 2 and 20 mT had no significant effect. These results are discussed in relation to weak magnetic field effects on free radical concentration.     

Studies of 50 Hz circularly polarized magnetic fields of up to 350 microT on reproduction and embryo-fetal development in rats: exposure during organogenesis or during preimplantation

Groups of mated female Sprague-Dawley rats were simultaneously exposed to 0 (sham exposed), 7, 70, or 350 microT (rms) circularly polarized 50 Hz magnetic fields (MF) for 22 h/day on gestational day 8-15, the period of rat fetal organogenesis (organogenesis study) or from day 0 to day 7 of gestation, the rat preimplantation period (preimplantation study). Developmental toxicity was assessed on gestational day 20. Identical experiments were repeated to confirm reproducibility of both studies. In both studies, statistically significant differences between exposed and sham exposed animals were observed in several measured parameters; however, these differences only appeared in one, but not both replicate experiments and generally at only an isolated exposure level. Because these differences were not reproducible and did not show a dose response relationship, they were not considered related to MF exposure. In the organogenesis study, lower kidney weights of dams were seen at 70 and 350 microT in Experiment 1. Lower dam liver weights and lower mean body weights of viable female and male fetuses were seen at 70 microT in Experiment 2. Otherwise, there were no differences in these parameters or in group means for fetal loss after implantation, number of viable fetuses, fetal body weight and sex ratio, incidences of external, visceral, and skeletal abnormalities or variations, or tissue abnormalities after histopathological examination. In the preimplantation study, dam health and indices for reproduction and embryo-fetal development, including pre or postimplantation loss, number and body weight of live fetuses, and sex ratio, external, skeletal abnormalities and variations, and skeletal ossification did not differ. Dam inorganic phosphorous concentration at 350 microT was elevated in one experiment and depressed in another. In one experiment, visceral abnormalities, primarily thymic remnant in neck and accessory liver lobe, were increased in the 7 microT group. Based on these results from two studies, we conclude that circularly polarized 50 Hz MF exposure of up to 350 microT during the fetal organogenesis or during the preimplantation period does not affect reproduction and embryo-fetal development in Sprague-Dawley rats.     

Effects of continuous and intermittent exposure to RF fields with a wide range of SARs on cell growth, survival, and cell cycle distribution

To examine the biological effects of radio frequency (RF) electromagnetic fields in vitro, we have examined the fundamental cellular responses, such as cell growth, survival, and cell cycle distribution, following exposure to a wide range of specific absorption rates (SAR). Furthermore, we compared the effects of continuous and intermittent exposure at high SARs. An RF electromagnetic field exposure unit operating at a frequency of 2.45 GHz was used to expose cells to SARs from 0.05 to 1500 W/kg. When cells were exposed to a continuous RF field at SARs from 0.05 to 100 W/kg for 2 h, cellular growth rate, survival, and cell cycle distribution were not affected. At 200 W/kg, the cell growth rate was suppressed and cell survival decreased. When the cells were exposed to an intermittent RF field at 300 W/kg(pk), 900 W/kg(pk) and 1500 W/kg(pk) (100 W/kg(mean)), no significant differences were observed between these conditions and intermittent wave exposure at 100 W/kg. When cells were exposed to a SAR of 50 W/kg for 2 h, the temperature of the medium around cells rose to 39.1 degrees C, 100 W/kg exposure increased the temperature to 41.0 degrees C, and 200 W/kg exposure increased the temperature to 44.1 degrees C. Exposure to RF radiation results in heating of the medium, and the thermal effect depends on the mean SAR. Hence, these results suggest that the proliferation disorder is caused by the thermal effect.     

Effects of sinusoidal magnetic field observed on cell proliferation, ion concentration, and osmolarity in two human cancer cell lines

Low frequency magnetic fields have previously been shown to affect cell functions. In this article, the effects of 20 mT, 50 Hz sinusoidal magnetic field on cell proliferation, ion concentration, and osmolarity in two human cancer cell lines (HL-60 and SK-Hep-1) were investigated. Inhibition of cell growth was observed. On the other hand, the exposure also increased the Na+, K+ ion concentration and osmolarity in cell supernatant compared to the control group. To our knowledge, this is the first study on cancer cells where magnetic fields affect osmolarity in cell supernatant. In addition, a model of cells exposed to the oscillating magnetic field is described as well as the characteristics of ions in and out of cells. The experimental data appears to be consistent with the theoretical analysis. The results are also discussed in terms of the relationships among cell growth, ion concentration, and osmolarity. Magnetic field inhibitions of cell growth in vitro may relate to changes in cell ion concentration and osmolarity.