Involvement of midkine expression in the inhibitory effects of low-frequency magnetic fields on cancer cells

Effects of magnetic fields (MFs) on cancer cells may depend on cell type and exposure conditions. Gene expression levels are different among cancer cells. However, the effect of MFs on cancer cells with different gene expressions is still unclear. In this study, the cancer cell lines BGC-823, MKN-45, MKN-28, A549, SPC-A1, and LOVO were exposed to a low-frequency MF. Specific parameters of MFs were determined. Furthermore, the potential of the MF to influence cancer cell growth with midkine (MK) expression was evaluated. Cell proliferation and cell cycle were detected using the CCK-8 assay and flow cytometry. Cell ultrastructure was observed by transmission electron microscopy. BGC-823 cells with over-expression of MK (BGC-MK cells) and stanniocalcin-1 were generated by plasmid construction and transfection. Results showed that exposure to a 0.4-T, 7.5 Hz MF inhibited the proliferation of BGC-823, MKN-28, A549, and LOVO cells, but not MKN-45 and SPC-A1 cells. Moreover, the inhibitory effect of the MF on BGC-MK cells was lower (12.3%) than that of BGC-823 cells (20.3%). Analysis of the cell cycle showed that exposure to the MF led to a significant increase in the S phase in BGC-823 cells, but not in BGC-MK cells. In addition, organelle morphology was modified in BGC-823 cells exposed to the MF. These results suggest that exposure to a 0.4-T, 7.5 Hz MF could inhibit tumor cell proliferation and disturb the cell cycle. The alteration of MK expression in cancer cells may be related to the inhibitory effect of the MF on these cells.     

The effect of oscillating low intensity magnetic field on the Na+, K+, Ca++, and Mg++ concentrations in the maternal and fetal circulation of the dually perfused human placental cotyledon

Dual-sided perfusions of the human placental cotyledon in vitro were used to study effects of low intensity magnetic fields (MFs) of 2 mT, 50 Hz (E1, 10 perfusions) and 5 mT, 50 Hz (E2, 10 perfusions). In the control group C (10 experiments) no field was used. Perfusions lasted 180 min each. Increased release of calcium ions from the placental cotyledon was found in the fetal circulation during perfusion when the 2 mT, 50 Hz MF was used. No changes in the release of sodium and magnesium ions were observed compared to the control group. The 5 mT, 50 Hz oscillating MF intensified the release of sodium ions from the perfused cotyledon both to the fetal and maternal circulation up to the 150th min of the experiment. Increased release of magnesium ions was observed only to the fetal circulation between 120 and 180 min and of calcium ions to the fetal circulation between 60 and 180 min. No significant differences in K concentrations were found between the control and MF exposed cotyledons under conditions of these experiments.     

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.     

Effect of ELF magnetic fields on protein synthesis in Escherichia coli K12

Escherichia coli K12 was used as a model system to determine whether ELF magnetic fields (MFs) are a general stress factor. The cells were exposed to ELF MFs (5-100 Hz) at a maximum intensity of 14 mT r. m.s. for circularly polarized MFs and 10 mT r.m.s. for vertically polarized MFs. The response of the cells to the MFs was estimated from the change in protein synthesis by using 2D PAGE. Approximately 1,000 proteins were separated on the 2D gels. The stress-responsive proteins such as CH10, DNAK, CH60, RECA, USPA, K6P1 and SODM were identified from the SWISS-2DPAGE database on the 2D gels. These proteins respond to most stress factors, including temperature change, chemical compounds, heavy metals, and nutrients. When the bacterial cells were exposed to each MF at 5-100 Hz under aerobic conditions (6.5 h) or at 50 Hz under anaerobic conditions (16 h) at the maximum intensity (7.8 to 14 mT r.m.s.), no reproducible changes were observed in the 2D gels. Changes in protein synthesis were detected by 2D PAGE with exposure to heat shock (50 degrees C for 30 min) or under anaerobic conditions (no bubbling for 16 h). Increases in the levels of synthesis of the stress proteins were observed in heat-shocked cells (CH60, CH10, HTPG, DNAK, HSLV, IBPA and some unidentified proteins) and in cells grown under anaerobic conditions (DNAK, PFLB, RECA, USPA and many unidentified proteins). These results suggest that 2D PAGE is sufficient to detect cell responses to environmental stress. The high-intensity ELF MFs (14 mT at power frequency) did not act as a general stress factor.     

Evaluation of the Effects of Power-Frequency Magnetic Field Exposure on B-Cell Differentiation From Human Hematopoietic Stem/Progenitor Cells

The causal relationship between exposure to power-frequency magnetic fields (MFs) and childhood leukemia has long been controversial. The most common type of childhood leukemia is acute B-lymphoblastic leukemia caused by abnormal proliferation of B cells in the early differentiation process. Here, we focused on B-cell early differentiation and aimed to evaluate the effects of exposing cells to power-frequency MF. First, we optimized an in vitro differentiation protocol of human hematopoietic stem/progenitor cells (HSPCs) to B-cell lineages. Following validation of the responsiveness of the protocol to additional stimulations and the uniformity of the experimental conditions, human HSPCs were continuously exposed to 300 mT of 50 Hz MF for 35 days of the differentiation process. These experiments were performed in a blinded manner. The percentages of myeloid or lymphoid cells and their degree of differentiation from pro-B to immature-B cells in the MF-exposed group showed no significant changes compared with those in the control group. Furthermore, the expression levels of recombination-activating gene (RAG)1 and RAG2 in the B cells were also similar to those in the control group. These results indicate that exposure to 50 Hz MF at 300 mT does not affect the human B-cell early differentiation from HSPCs. © 2023 The Authors. Bioelectromagnetics published by Wiley Periodicals LLC on behalf of Bioelectromagnetics Society.     

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.     

Effects of exposure to a 50 Hz sinusoidal magnetic field during the early adolescent period on spatial memory in mice

Adolescence is a critical developmental stage during which substantial remodeling occurs in brain areas involved in emotional and learning processes. Although a robust literature on the biological effects of extremely low frequency magnetic fields (ELF‐MFs) has been documented, data on the effects of ELF‐MF exposure during this period on cognitive functions remain scarce. In this study, early adolescent male mice were exposed from postnatal day (P) 23–35 to a 50 Hz MF at 2 mT for 60 min/day. On P36–45, the potential effects of the MF exposure on spatial memory performance were examined using the Y‐maze and Morris water maze tasks. The results showed that the MF exposure did not affect Y‐maze performance but improved spatial learning acquisition and memory retention in the water maze task under the present experimental conditions. Bioelectromagnetics 34:275–284, 2013. © 2012 Wiley Periodicals, Inc.     

EXPOSURE TO 50 Hz MAGNETIC FIELDS DOES NOT INDUCE THE PHOSPHORYLATION OF SEK1/MKK4 IN CULTURED CELLS

In our previous studies, we found that 50 Hz magnetic fields (MFs) could induce the phosphorylation of stress-activated protein kinase (SAPK) and enhance its enzymatic activity. In order to clarify the relationship between MF exposure and the SAPK pathway clearly, we studied the effects of 50 Hz MF exposure on phosphorylation (activation) of SEK1/MKK4 (the upstream kinase of SAPK). A Chinese hamster lung (CHL) cell line was exposed to 50 Hz MFs at two intensities (0.4 and 0.8 mT) for different durations, and Western blot analysis was used to measure the degree of phosphorylation (activation), and nonphosphorylation (non-activation) of SEK1/MKK4 with corresponding antibodies. The results showed that the exposures at both intensities could not induce the phosphorylation of SEK1/MKK4. However, treatment with high osmotic pressure NaCl could induce the phosphorylation of SEK1/MKK4 in cultured cells. It is suggested that 50 Hz MFs may activate the SAPK through a kinase other than SEK1/MKK4.     

ELF magnetic fields induce internalization of gap junction protein connexin 43 in Chinese hamster lung cells

We have previously demonstrated that exposure of Chinese hamster lung (CHL) cells to 50 Hz magnetic fields (MFs) and/or 12-O-tetradecanoylphorbol-3-acetate (TPA)-inhibited gap junctional intercellular communication (GJIC). To explore and compare the mechanisms of GJIC inhibition induced by extremely low frequency (ELF) MF and TPA, the number and localization of connexin 43 (C x 43) were studied. The localization of C x 43 was determined with indirect immunofluorescence histochemical analysis and detected by confocal microscopy after exposing CHL cells to 50 Hz sinusoidal magnetic field at 0.8 mT for 24 h without or with TPA (5 ng/ml) for the last 1 h. The C x 43 levels in nuclei and in cytoplasm were examined by Western blotting analysis. The results showed that the cells exposed to MF and/or TPA displayed individual plaques at regions of intercellular contact, which were fewer than the normal cells in number, while the number of C x 43 in cytoplasm increased and congregated near the nuclei. Western blot analysis further demonstrated the quantity of changes in location of Cx43. These results suggest that reduction of C x 43 at regions of intercellular contact may be one of the mechanisms of GJIC inhibition induced by ELF MF.     

Intracellular calcium activity in isolated bovine adrenal chromaffin cells in the presence and absence of 60 Hz magnetic fields

This study examined whether 60 Hz magnetic field (MF) exposure alters intracellular calcium levels ([Ca(2+)](i)) in isolated bovine adrenal chromaffin cells, a classic model of neural responses. [Ca(2+)](i) was monitored by fluorescence video imaging of cells loaded with the calcium indicator fluo-4 during exposures to magnetic flux densities of 0.01, 0.1, 1.0, 1.4, or 2.0 mT. MFs generated by Helmholtz coils constructed from bifilar wire allowed both 60 Hz field and sham exposures. Following a 5 min monitoring period to establish baseline patterns, cells were subjected for 10 min to a 60 Hz MF, sham field or no field. Reference calcium responses and assessment of cell excitability were obtained by the sequential addition of the nicotinic cholinergic receptor agonist dimethylphenylpiperazinium (DMPP) and a depolarizing concentration of KCl. Throughout an 8 day culture period, cells exhibited spontaneous fluctuations in [Ca(2+)](i). Comparisons of the number of cells exhibiting transients, the number and types of calcium transients, as well as the time during monitoring when transients occurred showed no significant differences between MF exposed cells and either sham exposed or nonexposed cells. With respect to the percentage of cells responding to DMPP, differences between 1 and 2 mT exposed cells and both nonexposed and sham exposed cells reached statistical significance during the first day in culture. No statistically significant differences were observed for responses to KCl. In summary, our data indicate that [Ca(2+)](i) in chromaffin cells is unaffected by the specific 60 Hz MF intensities used in this study. On the other hand, plasma membrane nicotinic receptors may be affected in a manner that is important for ligand-receptor interactions.     

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.     

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.     

Effects of magnetic field on the antioxidant enzyme activities of suspension-cultured tobacco cells

Effects of magnetic fields (MFs) on the activities of antioxidant enzymes of suspension-cultured tobacco cells were investigated. Compared with the control cells, exposure of the cells to static MF with the magnitudes of 10 and 30 mT for 5 days, 5 h each day, increased the activity of superoxide dismutase (SOD). In contrast, the activity of the catalase (CAT) and ascorbate peroxidase (APX) was decreased by MF, compared with those of the control cells. Level of lipid peroxidation was also increased by MF. It suggests that MF could deteriorate antioxidant defense system of plant cells.     

The effect of 100 Hz magnetic field combined with X-ray on hepatoma-implanted mice

Our previous cellular experiments demonstrated that 100 Hz magnetic field (MF) was effective at enhancing apoptosis of liver cancer cells BEL‐7402 induced by X‐ray irradiation. This study was performed to further explore the possible synergism between 100 Hz MF and X‐ray in treatment of hepatoma‐implanted Balb/c mice. 100 Hz MF exposure with a mean flux density of 0.7 mT was performed inside an energized solenoid coil. Six MV X‐ray irradiation was generated using a linear accelerator. Tumor growth and survival of mice implanted with H22 cells were evaluated by measuring the tumor diameters and overall days of survival. Six groups treated with 100 Hz MF or X‐ray alone or a combination of MF and X‐ray were examined. Furthermore, the effects of different numbers of MF exposure periods on tumor growth and mice survival were examined when combined with 4 Gy X‐ray. Data referring to overall survival days and tumor diameters of the above groups were compared using log‐rank test and Student's t‐test. Our results showed that five periods of combined 100 Hz MFs and 4 Gy X‐ray could significantly extend the overall days of survival and reduce the tumor size compared to MF or X‐ray alone. Also, a greater number of 100 Hz MF exposure periods could further improve the survival and inhibit tumor growth in hepatoma‐implanted mice when combined with 4 Gy X‐ray. In conclusion, these findings suggested that 100 Hz MF could possibly synergize with 4 Gy X‐ray in terms of survival improvement and tumor inhibition in hepatoma‐implanted mice. Bioelectromagnetics 32:322–324, 2011. © 2011 Wiley‐Liss, Inc.     

Magnetic field exposure enhances mRNA expression of σ32 in E. coli

The mechanism of interaction between weak electromagnetic fields and cells is not understood. As a result, the health effect(s) induced by exposure to these fields remains unclear. In addition to questions relating to the site of initial magnetic field (MF) interactions, the nature of the cell's response to these perturbations is also unclear. We examined the hypothesis that the cells respond to MFs in a manner similar to other environmental stressors such as heat. Using the bacterium Escherichia coli, we examined the mRNA levels of σ³², a protein that interacts with RNA polymerase to help it recognize a variety of stress promoters in the cell. Our data show that the intracellular level of σ³² mRNA is enhanced following a 15-min exposure to a 60 Hz, 1.1 mT magnetic field. J. Cell. Biochem. 68:1–7, 1998. © 1998 Wiley-Liss, Inc.     

Effects of magnetic fields on mammary tumor development induced by 7, 12-dimethylbenz(a)anthracene in rats

A series of epidemiological studies have indicated associations between exposure to magnetic fields (MFs) and a variety of cancers, including breast cancer. In order to test the possibility that MF acts as a cancer promoter or copromoter, four separate experiments have been conducted in rats in which the effects of chronic exposure to MFs on the development of mammary tumors induced by 7,12-dimethylbenz(a)anthracene (DMBA) were determined. Female rats were exposed in magnetic coils for 91 days (24 h/day) to either alternating current (AC; 50 Hz)-MF or direct current (DC)-MF. Magnetic flux density of the DC-MF was 15 mT. Two AC-MF exposures used a homogeneous field with a flux density of 30 mT (rms); one used a gradient field with flux density ranging from 0.3–1 μT. DMBA (5 mg) was administered orally at the onset of MF exposure and was repeated thrice at intervals of 1 week. In each experiment, 18–36 animals were exposed in 6 magnetic coils. The same number of rats were used as sham-exposed control. These control animals were treated with DMBA and were placed in dummy coils in the same room as the MF-exposed rats. Furthermore, groups of age-matched rats (reference controls) were treated with DMBA but housed in another room to exclude any MF exposure due to the magnetic stray field from the MF produced by coils. At the end of the exposure or sham-exposure period, tumor number and weight or size of tumors were determined at necropsy. Results were as follows: In sham-exposed animals or reference controls, the tumor incidence varied between 50 and 78% in the 4 experiments. The average number of mammary tumors per tumor-bearing animal varied between 1.6 and 2.9. In none of the experiments did MFs significantly alter tumor incidence, but in one of the experiments with AC-MF exposure at 30 mT, the number of tumors per tumor-bearing animal was significantly increased. Furthermore, exposure to a DC-MF at 15 mT significantly enhanced the tumor weight. Exposure to a gradient AC-MF at 0.3–1 μT exerted no significant effects. These experiments seem to indicate that MFs at high flux densities may act as a promoter or copromoter of breast cancer. However, this interpretation must be considered only a tentative conclusion because of the limitations of this study, particularly the small sample size used for MF exposure and the lack of repetition of data. © 1993 Wiley-Liss. Inc.     

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.     

Influence of pulsed electromagnetic and pulsed vector magnetic potential field on the growth of tumor cells

Aims and Background: Tumor diseases cause 20% of deaths in Europe and they are the second most common cause of death and morbidity after cardiovascular diseases. Thus, tumor cells are target of many therapeutic strategies and tumor research is focused on searching more efficient and specific drugs as well as new therapeutic approaches. One of the areas of tumor research is an issue of external fields. In our work, we tested influence of a pulsed electromagnetic field (PEMF) and a hypothetic field of the pulsed vector magnetic potential (PVMP) on the growth of tumor cells; and further the possible growth inhibition effect of the PVMP. Methods: Both unipolar and bipolar PEMF fields of 5?mT and PVMP fields of 0?mT at frequencies of 15?Hz, 125?Hz and 625?Hz were tested on cancer cell lines derived from various types of tumors: CEM/C2 (acute lymphoblastic leukemia), SU-DHL-4 (B-cell lymphoma), COLO-320DM (colorectal adenocarcinoma), MDA-BM-468 (breast adenocarcinoma), and ZR-75-1 (ductal carcinoma). Cell morphology was observed, proliferation activity using WST assay was measured and simultaneous proportion of live, early apoptotic and dead cells was detected using flow cytometry. Results: A PEMF of 125?Hz and 625?Hz for 24?h–48?h increased proliferation activity in the 2 types of cancer cell lines used, i.e. COLO-320DM and ZR-75-1. In contrast, any of employed methods did not confirm a significant inhibitory effect of hypothetic PVMP field on tumor cells.