Effect of ELF Pulsed Electromagnetic Fields on Protein Kinase C Activation Process in HL-60 Leukemia Cells

The activation of protein kinase C in HL-60 cells has been used as a model to investigate the molecular effects of the interaction of ELF pulsed electromagnetic fields (PEMFs) with the living systems. The shape of the pulsed magnetic field used in our experiments was a positive triangle (50 Hz, 8 mT peak). Protein kinase C is activated by association with plasma membranes; the membrane-associated enzyme binds phorbol esters. In the present study the process of protin kinase C activation was studied by measuring the binding of [³H]-phorbol-12,13-dibutyrate. The extent of labelled PDBu binding to HL-60 cells was increased by exposing the cells to the ELF electromagnetic field. Scatchard analysis of PDBu binding data showed an increased number of binding sites for the PDBu in the cells exposed to the electromagnetic field for 10, 15 or 20 min. Addition of EGTA to the culture medium resulted in a smaller stimulation of protein kinase C activation in the cells exposed to PEMF.     

Changes in the chromatin structure of lymphoid cells under the influence of low-intensity extremely high-frequency electromagnetic radiation against the background of inflammatory process

Using the alkaline single cell gel electrophoresis technique (comet assay), changes in chromatin structure of peripheral blood leukocytes and peritoneal neutrophils have been studied in mice exposed to low-intensity extremely high-frequency electromagnetic radiation (42.2 GHz, 0.1 mW/cm2, 20 min at 1 h after induction of inflammation) against the background of the systemic inflammatory process. It was revealed that the exposure of mice with the developing inflammation leads to a pronounced decrease in the level of DNA damage to peripheral blood leukocytes and peritoneal neutrophils. It is supposed that the changes in the chromatin structure of lymphoid cells have a genoprotective character in the inflammatory process and can underlie the mechanisms of realization of antiinflammatory effects of the electromagnetic radiation.     

Low electromagnetic field (50 Hz) induces differentiation on primary human oral keratinocytes (HOK)

This work concerns the effect of low frequency electromagnetic fields (ELF) on biochemical properties of human oral keratinocytes (HOK). Cells exposed to a 2 mT, 50 Hz, magnetic field, showed by scanning electron microscopy (SEM) modification in shape and morphology; these modifications were also associated with different actin distribution, revealed by phalloidin fluorescence analysis. Moreover, exposed cells had a smaller clonogenic capacity, and decreased cellular growth. Indirect immunofluorescence with fluorescent antibodies against involucrin and beta-catenin, both differentiation and adhesion markers, revealed an increase in involucrin and beta-catenin expression. The advance in differentiation was confirmed by a decrease of expression of epidermal growth factor (EGF) receptor in exposed cells, supporting the idea that exposure to electromagnetic field carries keratinocytes to higher differentiation level. These observations support the hypothesis that 50 Hz electromagnetic fields may modify cell morphology and interfere in differentiation and cellular adhesion of normal keratinocytes.     

Changes in the chromatin structure of lymphoid cells under the influence of low-intensity extremely high-frequency electromagnetic radiation against the background of inflammatory process

Using the alkaline single cell gel electrophoresis technique (comet assay), changes in chromatin structure of peripheral blood leukocytes and peritoneal neutrophils have been studied in mice exposed to low-intensity extremely high-frequency electromagnetic radiation (42.2 GHz, 0.1 mW/cm², 20 min at 1 h after induction of inflammation) against the background of the systemic inflammatory process. It was revealed that the exposure of mice with the developing inflammation leads to a pronounced decrease in the level of DNA damage to peripheral blood leukocytes and peritoneal neutrophils. It is supposed that the changes in the chromatin structure of lymphoid cells have a genoprotective character in the inflammatory process and can underlie the mechanisms of realization of antiinflammatory effects of the electromagnetic radiation.     

Effect of 72 Hz pulsed magnetic field exposure on macromolecular synthesis in CCRF-CEM cells.

Cells from the T-lymphoblastoid cell line, CCRF-CEM, have been exposed in vitro to a quasirectangular, asymmetric electromagnetic field pulsed at 72 Hz at 37 degrees for periods of 30 min to 24 h. RNA synthesis, assessed by incorporation of 3H-uridine, increased (relative to control cells) 2-fold after 30 min in exposed cells and achieved its greatest increase of 3.2-fold relative to controls after 2 h exposure. Increased precursor incorporation was observed at all subsequent exposure times up to 24 h. Synthesis of mRNA was similar, but not identical to that observed with total cellular RNA. Additionally, protein synthesis, determined by incorporation of radioactive precursor into acid-precipitable material, was increased 2.8-fold, compared to controls, after 2 h exposure. Longer exposure times resulted in an exponential decrease in precursor incorporation to 1.1-times control levels after 24 h. Using a dye reduction assay, mitochondrial activity was also found to be increased over a 24 h exposure period. No effect of electromagnetic field exposure was found on cellular synthesis of DNA. These data are generally consistent with other reports documenting effects of electromagnetic field exposure on macromolecular synthesis in vitro.     

Enhanced responsiveness to parathyroid hormone and induction of functional differentiation of cultured rabbit costal chondrocytes by a pulsed electromagnetic field

Pulsed electromagnetic fields promote healing of delayed united and ununited fractures by triggering a series of events in fibrocartilage. We examined the effects of a pulsed electromagnetic field (recurrent bursts, 15.4 Hz, of shorter pulses of an average of 2 gauss) on rabbit costal chondrocytes in culture. A pulsed electromagnetic field slightly reduced the intracellular cyclic adenosine 3',5'-monophosphate (cAMP) level in the culture. However, it significantly enhanced cAMP accumulation in response to parathyroid hormone (PTH) to 140% of that induced by PTH in its absence, while it did not affect cAMP accumulation in response to prostaglandin E1 or prostaglandin I2. The effect on cAMP accumulation in response to PTH became evident after exposure of the cultures to the pulsed electromagnetic field for 48 h, and was dependent upon the field strength. cAMP accumulation in response to PTH is followed by induction of ornithine decarboxylase, a good marker of differentiated chondrocytes, after PTH treatment for 4 h. Consistent with the enhanced cAMP accumulation, ornithine decarboxylase activity induced by PTH was also increased by the pulsed electromagnetic field to 170% of that in cells not exposed to a pulsed electromagnetic field. Furthermore, stimulation of glycosaminoglycan synthesis, a differentiated phenotype, in response to PTH was significantly enhanced by a pulsed electromagnetic field. Thus, a pulsed electromagnetic field enhanced a series of events in rabbit costal chondrocytes in response to PTH. These findings show that exposure of chondrocytes to a pulsed electromagnetic field resulted in functional differentiation of the cells.     

Electromagnetic fields alter the expression of estrogen receptor cofactors in breast cancer cells

Breast cancer is the most common malignancy of women in Western societies. The increasing exposure to electromagnetic fields has been suspected to contribute to the rising incidence of breast cancer in industrialized countries. The majority of breast tumors is treated with the partial antiestrogen tamoxifen. Most tumors become resistant to tamoxifen in the course of treatment resulting in treatment failure. Electromagnetic fields reduce the efficacy of tamoxifen similar to tamoxifen resistance. In this study we investigated the mechanism by which electromagnetic fields influence the sensitivity to tamoxifen. In cells exposed to 1.2 microT of a 50 Hz electromagnetic field gene expression of cofactors of the estrogen receptors was compared to sham exposed cells. Using a gene array technology several cofactors were found to be differentially expressed. The expression of the coactivators, SRC-1 and AIB1, and of two corepressors, N-Cor and SMRT, was quantified by RT-PCR. Both coactivators were expressed more strongly in the exposed cells while the expression of two corepressors decreased. The RNA analysis was confirmed by Western blots. The contradirectional changes in gene expression of coactivators and corepressors by electromagnetic fields results in a lower sensitivity to tamoxifen. Electromagnetic fields may contribute to the induction of tamoxifen resistance in vivo.     

Endothelial cell response to pulsed electromagnetic fields: stimulation of growth rate and angiogenesis in vitro

The effects of pulsed electromagnetic fields on the repopulation rate of denuded regions of endothelial cell monolayers and on endothelial cell reorganization into complex vessellike structures was monitored in vitro by using human umbilical vein and bovine aortic endothelial cells. A small (20-40%) but statistically significant enhancement in growth rate of partially denuded endothelial cell monolayers as determined by tritiated thymidine incorporation was observed in the presence of pulsed electromagnetic fields. Morphologically, endothelial cells entering the denuded regions were observed to be elongated, often connecting end to end to form a mycelial or "sprouting" pattern when exposed to pulsed electromagnetic fields. This was in contrast to cells outside of the field which had a more cuboidal morphology. Complete disruption of the endothelial cell monolayer by passaging the cells with EDTA-trypsin resulted in reorganization of some of the cells into three-dimensional vessellike structures after as little as 5-8 hours in the presence of the pulsed electromagnetic field. This reorganization occurred in the presence of heparin, endothelial cell growth factor, and a competent fibronectin matrix. Vascularization for comparable cultures outside of the field did not occur during the time-course of the experiments. Discrete stages of neovascularization were observed in the presence of the field that were qualitatively similar to stages of angiogenesis observed in vivo.     

Effect of low-level, 60-Hz electromagnetic fields on human lymphoid cells: I. Mitotic rate and chromosome breakage in human peripheral lymphocytes

Dividing human peripheral lymphocytes from 10 normal adults (5 males and 5 females) were exposed in vitro to low level 60-Hz electromagnetic fields for 69 hours. The current density of the electrical field was 30 microA/cm2, while the magnetic field was either 1 or 2 gauss. The cytological endpoints measured were mitotic rate and chromosome breakage. No statistically significant differences, indicative of a field effect, were observed between treated and control cells whether exposed to an electric field, a magnetic field, or to various combinations of the two.     

Biological and morphological effects on the brain after exposure of rats to a 1439 MHz TDMA field

We investigated the effects of exposure to a 1439 MHz TDMA (Time Division Multiple Access) field, as used in cellular phones, on the permeability of the blood-brain barrier (BBB), on the morphological changes of the brain, and on body-mass fluctuations. Male Sprague-Dawley (SD) rats were divided into three groups of eight rats each. The rats in the EM(+) group, which had their heads arrayed in a circle near the central antenna of an exposure system, were exposed to a 1439 MHz field for one hour a day. The rats in EM(-) group were also in the exposure system, however, without high-frequency electromagnetic wave (HF-EMW) exposure. The animals in the control group were neither placed in the system nor exposed to HF-EMWs. The exposure period was two or four weeks. The energy dose rate peaked at 2 W/kg in the brain; the average over the whole body was 0.25 W/kg. The changes in the permeability of BBB were investigated by Evans blue injection method and by immunostaining of serum albumin. HF-EMWs had no effect on the permeability of BBB. The morphological changes in the cerebellum were investigated by assessing the degeneration of Purkinje cells and the cell concentration in the granular layer. No significant changes were observed in the groups of rats exposed to HF-EMWs for two or four weeks. Averaged body masses were not affected by HF-EMWs exposure. In conclusion, a 1439 MHz TDMA field did not induce observable changes in the permeability of the BBB, morphological changes in the cerebellums, or body mass changes in rats, as evaluated by the conventional methods.     

Chromosomal aberrations in human amniotic cells after intermittent exposure to fifty hertz magnetic fields

Our recent studies have shown a significant increase in the frequency of chromosomal aberrations in human amniotic cells after exposure to a sinusoidal 50 Hz, 30 μT (rms) magnetic field. To evaluate further interactions between chromosomes and electromagnetic fields, we have analyzed the effects of intermittent exposure. Amniotic cells were exposed for 72 h to a 50 Hz, 30 μT (rms) magnetic field in a 15 s on and 15 s off fashion. Eight experiments with cells from different fetuses were performed. The results show a 4% mean frequency of aberrations among exposed cells compared to 2% in sham-exposed cells. The difference is statistically significant, with P < 0.05 both excluding and including gaps. In another series of eight experiments, the cells were exposed in the same way but with the field on for 2 s and off for 20 s. Also in these experiments a similar increase in the frequency of chromosomal aberrations was seen, but only when the analysis included gaps. Continuous exposure for 72 h to 300 μT, 50 Hz, did not increase the frequency of chromosomal aberrations. The background electromagnetic fields at different locations within the two incubators used was carefully checked and was nowhere found to exceed 120 nT. Likewise, the background level of chromosomal aberrations in cells cultured at different locations in the incubators showed no significant interculture differences. © 1994 Wiley-Liss, Inc.     

Calcium homeostasis of isolated heart muscle cells exposed to pulsed high-frequency electromagnetic fields

The intracellular calcium concentration ([Ca²⁺]_i) of isolated ventricular cardiac myocytes of the guinea pig was measured during the application of pulsed high-frequency electromagnetic fields. The high-frequency fields were applied in a transverse electromagnetic cell designed to allow microscopic observation of the myocytes during the presence of the high-frequency fields. The [Ca²⁺]_i was measured as fura-2 fluorescence by means of digital image analysis. Both the carrier frequency and the square-wave pulse-modulation pattern were varied during the experiments (carrier frequencies: 900, 1,300, and 1,800 MHz pulse modulated at 217 Hz with 14% duty cycle; pulsation pattern at 900 MHz: continuous wave, 16 Hz, and 50 Hz modulation with 50% duty cycle and 30 kHz modulation with 80% duty cycle). The mean specific absorption rate (SAR) values in the solution were within one order of magnitude of 1 mW/kg. They varied depending on the applied carrier frequency and pulse pattern. The experiments were designed in three phases: 500 s of sham exposure, followed by 500 s of field exposure, then chemical stimulation without field. The chemical stimulation (K⁺-depolarization) indicated the viability of the cells. The K⁺ depolarization yielded a significant increase in [Ca²⁺]_i. Significant differences between sham exposure and high-frequency field exposure were not found except when a very small but statistically significant difference was detected in the case of 900 MHz/50 Hz. However, this small difference was not regarded as a relevant effect of the exposure. © 1996 Wiley-Liss, Inc.     

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.     

The use of nonlinear dielectric spectroscopy to monitor the bioelectromagnetic effects of a weak pulsed magnetic field in real time

Nonlinear dielectric spectroscopy (NLDS) was used to detect interaction of a pulsed magnetic field (PMF) with membrane protein dynamics in aggregating Dictyostelium discoideum amoebae. In the experiments reported here, a strong nonlinear dielectric response of Dictyostelium discoideum cells is shown, and a distinctive nonlinear dielectric response of cells previously exposed to PMF is shown. The method of NLDS is shown to be capable of monitoring and charting the dynamic frequency response of the cell to an electromagnetic field.     

A Study of the Effects of Flux Density and Frequency of Pulsed Electromagnetic Field on Neurite Outgrowth in PC12 Cells

The aim of this study was to investigate the influence of pulsed electromagnetic fields with various flux densities and frequencies on neurite outgrowth in PC12 rat pheochromocytoma cells. We have studied the percentage of neurite-bearing cells, average length of neurites and directivity of neurite outgrowth in PC12 cells cultured for 96 hours in the presence of nerve growth factor (NGF). PC12 cells were exposed to 50 Hz pulsed electromagnetic fields with a flux density of 1.37 mT, 0.19 mT and 0.016 mT respectively. The field was generated through a Helmholtz coil pair housed in one incubator and the control samples were placed in another identical incubator. It was found that exposure to both a relatively high flux density (1.37 mT) and a medium flux density (0.19 mT) inhibited the percentage of neurite-bearing cells and promoted neurite length significantly. Exposure to high flux density (1.37 mT) also resulted in nearly 20% enhancement of neurite directivity along the field direction. However, exposure to low flux density field (0.016 mT) had no detectable effect on neurite outgrowth. We also studied the effect of frequency at the constant flux density of 1.37 mT. In the range from 1 ∼ 100 Hz, only 50 and 70 Hz pulse frequencies had significant effects on neurite outgrowth. Our study has shown that neurite outgrowth in PC12 cells is sensitive to flux density and frequency of pulsed electromagnetic field.     

APOPTOSIS OF EHRLICH MOUSE ASCITES CELLS INDUCED BY LONG-TERM EXPOSURE OF WEAK ELECTROMAGNETIC FIELDS IN VIVO

To study the possibility of apoptosis of tumor cells induced by weak electromagnetic fields (EMFs) in vivo, mice were inoculated with Ehrlich ascites cells and exposed to a long-term electromagnetic field (1 mT, 700 KHz). During the treatment, growth curves of mice were measured and compared between exposed and sham-exposed mice. The results show that the growth curves of healthy controls agree well with the ideal curve of logistic growth, but the growth curves of cancer mice deviate from the ideal curve. There is no difference in growth curves between exposed, and sham-exposed healthy mice, and they both agree with the ideal curve. However, a notable difference in growth curves between exposed and sham-exposed cancer mice was obtained. Moreover, the curves of sham-exposed mice deviate even more than those of the exposed mice; in other words, the growth curves of Ehrlich ascites mice deviate from the ideal curve of healthy mice but are shifted toward it by the EMF treatments. After the treatment, apoptosis of Ehrlich ascites cells from inoculated mice was analyzed by several methods, including flow cytometry, fluorescence microscopy, and DNA gel electrophoresis. Statistical analysis from flow cytometry shows that the apoptotic ratio of cells from exposed Ehrlich ascites mice was significantly higher than that from sham-exposed treated mice. Microscopic observation of Ehrlich ascites cells stained with acridine orange (AO) and propidium iodide (PI) showed typical apoptotic changes in exposed animals whose cell nuclei were highly condensed or fragmented and uniformly stained green by the AO, whereas cell nuclei from sham-exposed mice were stained green and showed a fine reticular pattern. Agarose gel electrophoresis of DNA from exposed mice showed that the chromatin DNA exhibited ladders, a characteristic feature of internucleosomal degradation of DNA by EMF treatments. For interactions between external electromagnetic fields and DNA, the mechanism of apoptosis of tumor cells induced by weak EMFs is discussed.     

Growth inhibition of Staphylococcus aureus induced by low‐frequency electric and electromagnetic fields

Magnetic field therapy is an established technique in the treatment of pseudarthrosis. In cases of osteomylitis, palliation is also observed. This study focuses on the impact of different electric and electromagnetic fields on the growth of Staphylococcus aureus by in vitro technologies. Cultures of Staphylococcus aureus in fluid and gel‐like medium were exposed to a low‐frequency electromagnetic field, an electromagnetic field combined with an additional electric field, a sinusoidal electric field and a static electric field. In gel‐like medium no significant difference between colony‐forming units of exposed samples and non‐exposed references was detected. In contrast, Staphylococcus aureus concentrations in fluid medium could clearly be reduced under the influence of the four different applied fields within 24 h of experiment. The strongest effects were observed for the direct current electric field which could decrease CFU/ml of 37%, and the low‐frequency electromagnetic field with additional induced electric alternating field with a decrease of Staphylococci concentration by 36%. The effects of the electromagnetic treatment on Staphylococci within fluid medium are significantly higher than in gel‐like medium. The application of low‐frequency electromagnetic fields corroborates clinical situations of bone infections during magnetic field therapy. Bioelectromagnetics 30:270–279, 2009. © 2009 Wiley‐Liss, Inc.     

Nonlinear mechanism for the generation of electromagnetic fields in a magnetized plasma by the beatings of waves

The modulational instability in a plasma in a strong constant external magnetic field is considered. The plasmon condensate is modulated not by conventional low-frequency ion sound but by the beatings of two high-frequency transverse electromagnetic waves propagating along the magnetic field. The instability reduces the spatial scales of Langmuir turbulence along the external magnetic field and generates electromagnetic fields. It is shown that, for a pump wave with a sufficiently large amplitude, the effect described in the present paper can be a dominant nonlinear process.     

Calcium Ion Cyclotron Resonance (ICR), 7.0 Hz, 9.2 μT Magnetic Field Exposure Initiates Differentiation of Pituitary Corticotrope-Derived AtT20 D16V Cells

The aim of this work is the study of the effect of electromagnetic radiations (ELF-EMF) tuned to the calcium cyclotron resonance condition of 7.0?Hz, 9.2?μT on the differentiation process of pituitary corticotrope-derived AtT20 D16V cells. These cells respond to nerve growth factor by extending neurite-like processes. To establish whether exposure to the field could influence the molecular biology of the pituitary gland, a corticotrope-derived cells line (AtT20 D16V) was exposed to ELF-EMF at a frequency of 7.0?Hz, 9.2?μT electromagnetic field by a Vega Select 719 power supply. Significant evidence was obtained to conclude that as little as 36?h exposure to the Ca²⁺ ICR condition results in enhanced neurite outgrowth, with early expression and aggregation of the neuronal differentiation protein NF-200 into neurite structures.