Radiofrequency radiation-induced changes in Leydig cell function.

Leydig cells are the primary producers of testosterone and play a crucial role in male fertility. Damage to these cells can compromise spermatogenesis and lead to testicular dysfunction. While a growing body of research suggests that radiofrequency electromagnetic fields (RF-EMF) can induce oxidative stress, mitochondrial dysfunction, and sperm abnormalities, our understanding of their direct influence on cell cycle regulation and Leydig cell proliferation remains limited. This study addresses this knowledge gap by examining frequency- and time-dependent effects.

The authors exposed the murine TM-3 Leydig cell line to three different RF sources under non-thermal conditions: a commercial 4G mobile phone (Xiaomi Note 7, 2318 MHz, 9.25 V/m), an 1800 MHz unmodulated field from a signal generator (continuous wave, 14.69 V/m), and a 2450 MHz unmodulated field from a signal generator (continuous wave, 13.44 V/m). The specific absorption rate (SAR) value for the mobile phone was calculated to be 0.5 W/kg. Cells exposed to a sham field were used as controls. Exposure durations were 15, 30, 45, 60, 90, and 120 minutes. The temperature difference between the exposed cells and the controls was less than 0.1°C. The electric field was monitored using a Narda 520 system. The scientists investigated morphological changes using microscopy and examined cell proliferation and DNA synthesis using an enzyme-linked immunosorbent assay (ELISA) with bromodeoxyuridine (BrdU). They also analyzed the cell cycle using flow cytometry. All experiments were independently repeated three times (n = 3). The statistical analysis included one- and two-way ANOVA with a Dunnett’s multiple comparisons test.

All RF sources resulted in frequency- and time-dependent morphological signs of cellular stress. These included cell rounding, loss of adherence, and reduced cell density, which is a sign of cell death. The strongest and earliest effects were observed at 2450 MHz, followed by the 4G mobile phone and 1800 MHz sources. Consistent with these findings, the BrdU assay revealed a progressive reduction in DNA synthesis, indicating decreased cell division. This reduction was statistically significant with mobile phone radiation exposure after 45 minutes, with 1800 MHz exposure after 60 minutes, and with 2450 MHz exposure after 90 minutes. However, the greatest reduction in relative cell division was observed with 2450 MHz exposure, dropping below 50% after 90 and 120 minutes. Cell cycle analysis revealed an accumulation of the cell population in the G1 phase, accompanied by a decrease in the S phase. The G2/M phase percentage remained unchanged across all conditions. These results suggest cell cycle arrest in the G1 phase.

The results of this study demonstrate that non-thermal RF exposure, including that from a commercial mobile phone, can impact the morphology, division, and cycle of Leydig cells in vitro. This influence was frequency- and dose-dependent. Concurrent findings from the BrdU assay and cell cycle analysis suggest sequential damage. DNA replication is functionally inhibited, which subsequently manifests as cell cycle arrest in the G1 phase. The authors discuss oxidative stress as the mechanistic basis because they had previously observed reduced cell proliferation and testosterone production, as well as increased levels of reactive oxygen species, following RF exposure.

Editor’s note:

The strengths of the present study lie in its controlled, non-thermal experimental design. This design included independently verified exposure parameters and statistically robust replication of the experiment. Notably, the DNA synthesis and cell cycle analyses are systematically coherent and support the hypothesis of progressive replication stress. Another recent study by this research group investigated the effect of various RF sources on the redox system of Leydig cells that contributes to the elucidation of the mechanistic background of the present study (see "Disruption of cellular redox balance and apoptosis: Different effects of RF frequencies on Leydig cells" [1], which is also discussed in this issue, EREP 02/26). Interestingly, despite its lower field strength, the commercial mobile phone has almost always produced more harmful effects than the 1800 MHz signal generator. Regarding cell proliferation, the mobile phone produced statistically significant effects even with the shortest exposure duration. This suggests that real-world modulated mobile phone signals have greater biological potency than continuous waves. However, the study has limitations, including the use of an in vitro cell line, which restricts its applicability to primary cells or in vivo models. The study confirms the results of earlier findings (reviewed in ElektrosmogReport) that describe the harmful effects of RF radiation on testosterone production [2–4]. (RH)

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2.   Miao X, Lin Y, Guo J, Lin J, Gao P, Zhang W et al. (2025). Differential metabolic responses of mouse Leydig and spermatogonia cells to radiofrequency electromagnetic field exposure. Frontiers in Public Health, 13, 1623701. https://doi.org/10.3389/fpubh.2025.1623701

3.   Kaur P, Rai U, Singh R (2023). Genotoxic risks to male reproductive health from radiofrequency radiation. Cells, 12(4), 1–20. https://doi.org/10.3390/cells12040594

4.   Maluin SM, Jaffar FHF, Osman K, Zulkefli AF, Mat Ros MF, Ibrahim SF (2024). Exploring edible bird nest’s potential in mitigating Wi-Fi’s impact on male reproductive health. Reproductive Medicine and Biology, 23(1), 1–13. https://doi.org/10.1002/rmb2.12606