Medical/biological study (experimental study)

A 60 Hz uniform electromagnetic field promotes human cell proliferation by decreasing intracellular reactive oxygen species levels med./bio.

By: Song K, Im SH, Yoon YJ, Kim HM, Lee HJ, Park GS

Published in: PLoS One 2018; 13 (7): e0199753

Aim of study (acc. to author)

To investigate the effects of a uniform 60 Hz magnetic field on cell proliferation and its underlying mechanism of action in human lung fibroblasts and HeLa cells.

Background/further details

In previous studies, the authors showed that an exposure to a gradient magnetic field with 60 Hz (6 or 7 mT) induced DNA damage and apoptosis (Kim et al., 2012; Kim et al., 2010). However, other studies showed contradictory results (e.g., Jin et al., 2014; Hong et al., 2012). Because of the inconsistency, the authors hypothesize that there is an overlooked parameter which plays a key role and paid attention to the uniformity of the magnetic field in this study.
Cell viability and reactive oxygen species level were also tested in the presence of glucose oxidase which produces low amounts of intracellular hydrogen peroxide and reduces cell viability. If the exposure to the 60 Hz-magnetic field decreased the intracellular reactive oxygen species level, the anti-proliferative effect of the glucose oxidase would be mitigated.
A positive control was used to evaluate DNA damage.

Endpoint

cell viability/cell division/proliferation: cell viability and underlying mechanism of action

Exposure

- 50/60 Hz
- magnetic field

Exposure	Parameters
Exposure 1: 60 Hz Exposure duration: 10, 30, or 60 minutes single exposure	magnetic flux density: 6 mT
Exposure 2: 60 Hz Exposure duration: 30 minutes daily for 3 days repetitive exposure	magnetic flux density: 3 mT magnetic flux density: 6 mT
Exposure 3: 60 Hz Exposure duration: 24, 48, 72, or 168 hours continuous exposure	magnetic flux density: 1 mT magnetic flux density: 6 mT magnetic flux density: 10 mT

General information

Two exposure devices were tested: an open-type Helmholtz coil device and a closed-type Helmholtz coil device. The uniformity is better in the closed-type device, therefore it was used for all experiments and is only described here.

Exposure 1

Main characteristics

Frequency	60 Hz
Type	magnetic field
Exposure duration	10, 30, or 60 minutes
Additional info	single exposure

Exposure setup

Exposure source	coil(s)
Chamber	cells were exposed in a petri dish (diameter 35 mm)
Setup	exposure system consisted of 28 ferrite cores (85 mm x 105 mm) and had a total length of 280 mm; Helmholtz-like coils were used to generate the magnetic field; each coil was 0.4 mm in diameter and had 1,000 turns; 6 dishes were exposed simultaneously; exposure system was put into a CO₂ incubator where temperature was monitored and maintained at 37°C ± 0.5°C

Parameters

Measurand	Value	Type	Method	Mass	Remarks
magnetic flux density	6 mT	-	measured	-	-

Exposure 2

Main characteristics

Frequency	60 Hz
Type	magnetic field
Exposure duration	30 minutes daily for 3 days
Additional info	repetitive exposure

Exposure setup

Exposure source	same setup as exposure 1

Parameters

Measurand	Value	Type	Method	Mass	Remarks
magnetic flux density	3 mT	-	measured	-	-
magnetic flux density	6 mT	-	measured	-	-

Exposure 3

Main characteristics

Frequency	60 Hz
Type	magnetic field
Exposure duration	24, 48, 72, or 168 hours
Additional info	continuous exposure

Exposure setup

Exposure source	same setup as exposure 1

Parameters

Measurand	Value	Type	Method	Mass	Remarks
magnetic flux density	1 mT	-	measured	-	-
magnetic flux density	6 mT	-	measured	-	-
magnetic flux density	10 mT	-	measured	-	-

Exposed system:

intact cell/cell culture
HeLa cells (human cervical carcinoma cell line); IMR-90 cells (human lung fibroblasts)

Methods Endpoint/measurement parameters/methodology

cell viability/cell division/proliferation: cell viabilty with and without glucose oxidase (MTT assay and haemocytometer); cell cycle distribution (propidium iodide staining, flow cytometry)
level of reactive oxygen species (ROS) with and without glucose oxidase (dichlorofluorescein, fluorescence spectrometry and flow cytometry)
molecular biosynthesis: protein expression of proteins involved in cell cycle, reactive oxygen species (ROS) generation and DNA damage: cyclin-dependent kinase 4 (CDK4) as a marker for G1 progression, proliferation cell nuclear antigen (PCNA) as a marker for DNA synthesis in S-phase, phosphorylated histone H3 (p-H3) as a marker for G2 phase/M phase; phosphorylated ERK 1/2, ERK 1/2, phosphorylated proteine kinase B (p-Akt) and protein kinase B (Akt) as evidence for generation of reactive oxygen species (ROS); protein expression of gamma-H2AX as a marker of DNA damage (Western blot)

Investigated system:

isolated bio./chem. substance
intact cell/cell culture

Main outcome of study (acc. to author)

The following results are always valid for both cell lines:
Neither a single exposure nor a repetitive exposure influenced cell viability or induced DNA damage. However, continuously exposed cell cultures showed a significantly increased cell viability compared to control cells dependent on the magnetic flux density: The higher the magnetic field, the stronger the increase in cell viability. This effect was reversible. When the cells were exposed for 72 hours and further incubated for 96 hours without exposure, the increase in cell viability was no longer present.
All markers indicating different cell cycle phases were increased in exposed cells compared to control cells. However, no significant changes in the distribution were observed.
The level of ROS was significantly decreased after continuous exposure to the magnetic field when compared with control cells. This finding was also supported by an increased phosphorylation of ERK 1/2 and Akt (but mainly not significant) in exposed cells compared to control cells. Additionally, the experiments with glucose oxidase indicated the involvement of ROS.
The authors suggest that continuous exposure to a 60 Hz magnetic field increased the cell viability in human lung fibroblasts and HeLa cells by reducing intracellular ROS levels.

Study character:

medical/biological study
experimental study
full/main study

Study funded by

National Research Foundation (NRF) of Korea

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Kim J et al. (2012): Time-varying magnetic fields of 60 Hz at 7 mT induce DNA double-strand breaks and activate DNA damage checkpoints without apoptosis
Kim J et al. (2010): Repetitive exposure to a 60-Hz time-varying magnetic field induces DNA double-strand breaks and apoptosis in human cells
Koh EK et al. (2008): A 60-Hz sinusoidal magnetic field induces apoptosis of prostate cancer cells through reactive oxygen species
Palumbo R et al. (2006): Effects on apoptosis and reactive oxygen species formation by Jurkat cells exposed to 50 Hz electromagnetic fields

A 60 Hz uniform electromagnetic field promotes human cell proliferation by decreasing intracellular reactive oxygen species levels med./bio.

Aim of study (acc. to author)

Background/further details

Endpoint

Exposure

General information

Exposure 1

Main characteristics

Exposure setup

Parameters

Exposure 2

Main characteristics

Exposure setup

Parameters

Exposure 3

Main characteristics

Exposure setup

Parameters

Methods Endpoint/measurement parameters/methodology

Main outcome of study (acc. to author)

Study funded by

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