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Updated: Dec 6, 2025
Assessing Microglial Phagocytosis of Myelin Debris in vitro Under Repeated Magnetic Stimulation
Published on: June 17, 2025
Cell physiological responses of RAW264 macrophage cells to a 50-Hz magnetic field
Chihiro Nishigaki1, Maresuke Nakayama1, Hidetake Miyata1
1Department of Physics, Tohoku University, Sendai, Japan.
Purpose:
We previously showed that a 0.5-mT, 50-Hz sinusoidal magnetic field (LFMF) enhanced DNA single-strand breaks (SSB) and necrosis in RAW264 macrophages that had been stimulated by bacterial endotoxin (lipopolysaccharide; LPS). LPS enhances production of nitric oxide (NO) and superoxide anion (O2-) that react with each other to generate peroxynitrite (ONOO-). ONOO- causes DNA strand breaks. Hence, we anticipated that 0.5-mT, 50-Hz sinusoidal magnetic field increased production of NO, thereby increasing intracellular ONOO- concentration and promoted DNA strand breaks. However, the NO production was not increased. In this study, we examined if the exposure of the cell to 0.5-mT, 50-Hz magnetic field for 24 h (1) promotes O2- production, (2) elevated the degree of apoptosis, because apoptosis is an upstream event of necrosis, (3) lowers mitochondrial membrane potential (ΔΦm), because it would also promote necrosis.
Materials And Methods:
O2-, was measured with nitroblue tetrazolium and water-soluble tetrazolium salt. Necrosis and apoptosis were quantified with propidium iodide and fluorescence labelling of caspases, respectively. The ΔΦm was measured with a fluorescent probe (JC-1) that reflects ΔΦm. Results and conclusions: In the LPS-stimulated macrophage, the LFMF did not promote O2- production. Thus, the LFMF-promoted DNA strand breaks did not result from the increase in the O2- production. The LFMF did not promote apoptosis, whereas it tended to increase the degree of necrosis, as we showed previously. The ΔΦm slightly declined in the LFMF-exposed cell without statistical significance.
Insights
Low-frequency magnetic fields (LFMF) did not increase nitric oxide or superoxide anion production in LPS-stimulated macrophages. LFMF exposure did not promote apoptosis but tended to increase necrosis, suggesting alternative mechanisms for DNA damage.
Area of Science:
- Cell Biology
- Biophysics
- Toxicology
Background:
- Bacterial endotoxin (lipopolysaccharide; LPS) stimulates macrophages, increasing nitric oxide (NO) and superoxide anion (O2-) production.
- These reactive oxygen species can form peroxynitrite (ONOO-), leading to DNA single-strand breaks (SSB).
- Previous studies indicated that a 0.5-mT, 50-Hz sinusoidal magnetic field (LFMF) enhances SSB and necrosis in LPS-stimulated macrophages.
Purpose of the Study:
- To investigate the mechanisms by which LFMF enhances DNA damage and necrosis in LPS-stimulated macrophages.
- To determine if LFMF affects superoxide anion (O2-) production, apoptosis, or mitochondrial membrane potential (ΔΦm).
Main Methods:
- RAW264 macrophages were stimulated with LPS and exposed to a 0.5-mT, 50-Hz sinusoidal magnetic field for 24 hours.
- Superoxide anion (O2-) production was measured using nitroblue tetrazolium and water-soluble tetrazolium salt assays.
- Apoptosis and necrosis were quantified using propidium iodide and caspase labeling, respectively.
- Mitochondrial membrane potential (ΔΦm) was assessed using the fluorescent probe JC-1.
Main Results:
- LFMF exposure did not increase O2- production in LPS-stimulated macrophages.
- LFMF did not promote apoptosis but showed a trend towards increased necrosis.
- A slight, statistically insignificant decline in mitochondrial membrane potential (ΔΦm) was observed in LFMF-exposed cells.
Conclusions:
- The LFMF-induced DNA strand breaks in LPS-stimulated macrophages are not mediated by increased O2- production.
- LFMF does not appear to induce apoptosis, but may contribute to necrosis through other pathways.
- Further research is needed to elucidate the precise mechanisms underlying LFMF-induced DNA damage and necrosis.
