The effects of exposure of neuroblastoma cells to a 50 Hz magnetic field on the cell cycle and cell proliferation and the underlying signal pathways should be investigated.
Previous studies by the authors (Trillo et al. 2012, Cid et al. 2012, Martinez et al. 2012, Trillo et al. 2013) have found increased cell proliferation and MAPK-ERK-1/2 signal pathway activation in human cancer cell lines after exposure to 50 Hz magnetic fields. The present study should investigate if changes in cell cycle regulation and other signaling pathways relevant to cell proliferation might be involved in the increased cell proliferation.
Cells were treated with the p38 MAPK inhibitor SB203580 (10 µM) or the JNK inhibitor SP600125 (20 µM) to investigate the involvement of these signaling pathways. Cells were also treated with N-acetyl-L-cysteine (1 mM and 10 mM) to inhibit reactive oxygen species.
The different exposure conditions and durations for individual methods were based on results from former studies.
| Exposure | Parameters |
|---|---|
|
Exposure 1:
50 Hz
Exposure duration:
continuous for 15, 30, 60 or 120 minutes
|
|
|
Exposure 2:
50 Hz
Exposure duration:
intermittent (3 h on/3 h off) for 24, 42 or 63 hours
|
|
| Frequency | 50 Hz |
|---|---|
| Type | |
| Waveform | |
| Exposure duration | continuous for 15, 30, 60 or 120 minutes |
| Additional info | vertically polarized |
| Exposure source | |
|---|---|
| Chamber | one pair of Helmholtz coils was placed inside each of two chambers magnetically shielded chambers located within two CO2 incubators |
| Setup | in each experimental run, petri dishes (five per experimental group) were stacked in the central region of the Helmholtz coil gap, which ensured uniformity of exposure; no increase of temperature at the sample location was observed during exposure |
| Sham exposure | A sham exposure was conducted. |
| Additional info | the setup was composed of two identical exposure systems; following a random sequence, both coil sets and incubators were alternatively used for exposure or sham exposure |
| Measurand | Value | Type | Method | Mass | Remarks |
|---|---|---|---|---|---|
| magnetic flux density | 100 µT | effective value | measured | - | - |
| Frequency | 50 Hz |
|---|---|
| Type | |
| Waveform | |
| Exposure duration | intermittent (3 h on/3 h off) for 24, 42 or 63 hours |
| Additional info | vertically polarized |
| Exposure source |
|
|---|---|
| Sham exposure | A sham exposure was conducted. |
| Measurand | Value | Type | Method | Mass | Remarks |
|---|---|---|---|---|---|
| magnetic flux density | 100 µT | effective value | measured | - | - |
The cell cycle distribution showed significant changes after 24 h (increased proportion of cells in S phase) and 42 h (decreased proportion of cells in G0 phase/G1 phase) of exposure compared to the control group. Exposure to the magnetic field for 120 minutes significantly increased the protein expression of cyclin D1 and decreased that of p27Kip1.
As expected, cells exposed to the magnetic field for 63 hours showed a significantly increased cell proliferation. The proliferative effect of the magnetic field was blocked by the p38 MAPK inhibitor, but not by the JNK inhibitor. Moreover, the proliferative effect of the MF was blocked by 1 mM of N-acetyl-L-cysteine while a concentration of 10 mM significantly reduced cell proliferation compared to the control group with and without co-exposure to the magnetic field.
After 15 minutes and 30 minutes of exposure, the protein expression of phosphorylated p38 MAPK was significantly increased compared to the control group and this increase could be blocked by treatment with the p38 MAPK inhibitor or N-acetyl-L-cysteine. The protein expression of phosphorylated JNK was significantly increased after 15 min and significantly decreased after 60 min of exposure compared to the control group. The phosphorylated ERK-1/2 protein expression was significantly increased after 30 minutes of exposure and was not influenced by N-acetyl-L-cysteine treatment.
The authors conclude that exposure of neuroblastoma cells to a 50 Hz magnetic field might induce changes in cell cycle and increase cell proliferation. The effects are probably mediated by a radical-dependent activation of the p38 MAPK and a radical-independent activation of the ERK-1/2 signal pathways.
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