Extremely Low-Frequency Electromagnetic Fields Affect the miRNA-Mediated Regulation of Signaling Pathways in the GC-2 Cell Line

Extremely low-frequency electromagnetic fields (ELF-EMFs) can affect male reproductive function, but the underlying mechanism of this effect remains unknown. miRNA-mediated regulation has been implicated as an important epigenetic mechanism for regulatory pathways. Herein, we profiled miRNA expression in response to ELF-EMFs in vitro. Mouse spermatocyte-derived GC–2 cells were intermittently exposed to a 50 Hz ELF-EMF for 72 h (5 min on/10 min off) at magnetic field intensities of 1 mT, 2 mT and 3 mT. Cell viability was assessed using the CCK–8 assay. Apoptosis and the cell cycle were analyzed with flow cytometry. miRNA expression was profiled using Affymetrix Mouse Genechip miRNA 3.0 arrays. Our data showed that the growth, apoptosis or cell cycle arrest of GC–2 cells exposed to the 50 Hz ELF-EMF did not significantly change. However, we identified a total of 55 miRNAs whose expression significantly changed compared with the sham group, including 19 differentially expressed miRNAs (7 miRNAs were upregulated, and 12 were downregulated) in the 1 mT exposure group and 36 (9 miRNAs were upregulated, and 27 were downregulated) in the 3 mT exposure group. The changes in the expression of 15 selected miRNAs measured by real-time PCR were consistent with the microarray results. A network analysis was used to predict core miRNAs and target genes, including miR-30e-5p, miR-210-5p, miR-196b-5p, miR-504-3p, miR-669c-5p and miR-455-3p. We found that these miRNAs were differentially expressed in response to different magnetic field intensities of ELF-EMFs. GO term and KEGG pathway annotation based on the miRNA expression profiling results showed that miRNAs may regulate circadian rhythms, cytokine-cytokine receptor interactions and the p53 signaling pathway. These results suggested that miRNAs could serve as potential biomarkers, and the miRNA-mediated regulation of signaling pathways might play significant roles in the biological effects of ELF-EMFs.