900 MHz Radiofrequency Field Induces Mitochondrial Unfolded Protein Response in Mouse Bone Marrow Stem Cells

Abstract

Objective: To examine whether exposure of mouse bone marrow stromal cells (BMSC) to 900 MHz radiofrequency fields used in mobile communication devices can induce mitochondrial unfolded protein response (UPR^mt). Methods: BMSCs were exposed to continuous wave 900 MHz radiofrequency fields (RF) at 120 μW/cm² power intensity for 4 h/d for 5 consecutive days. Cells in sham group (SH) were cultured in RF exposure system, but without RF radiation. The positive control cells were irradiated with 6 Gy X-ray at a dose rate of 1.103 Gy/min (XR). To inhibit the upstream molecular JNK2 of UPR^mt, cells in siRNA + RF, and siRNA + XR group were also pretreated with 100 nM siRNA-JNK2 for 48 h before RF/XR exposure. Thirty minutes, 4 h, and 24 h post-RF/XR exposure, cells were collected, the level of ROS was measured with flow cytometry, the expression levels of UPR^mt-related proteins were detected using western blot analysis. Results: Compared with Sham group, the level of ROS in RF and XR group was significantly increased 30 min and 4 h post-RF/XR exposure (P < 0.05), however, the RF/XR-induced increase of ROS level reversed 24 h post-RF/XR exposure. Compared with Sham group, the expression levels of HSP10/HSP60/ClpP proteins in cells of RF and XR group increased significantly 30 min and 4 h post-RF/XR exposure (P < 0.05), however, the RF/XR-induced increase of HSP10/HSP60/ClpP protein levels reversed 24 h post-RF exposure. After interfering with siRNA-JNK2, the RF/XR exposures could not induce the increase of HSP10/HSP60/ClpP protein levels any more. Conclusions: The exposure of 900 MHz RF at 120 μW/cm² power flux density could increase ROS level and activate a transient UPR^mt in BMSC cells. Mitochondrial homeostasis in term of protein folding ability is restored 24 h post-RF exposure. Exposure to RF in our experimental condition did not cause permanent and severe mitochondrial dysfunctions. However, the detailed underlying molecular mechanism of RF-induced UPR^mt remains to be further studied.

Keywords: heat shock protein; microwave; mitochondria; radiofrequency fields; reactive oxygen species; unfolded protein response.

Figure 1

ROS levels in BMSCs at 30 min, 4 h, and 24 h post exposure. SH, Sham; RF, RF-exposed; XR, X-rays irradiated. RF and XR vs. SH: ^*P < 0.05; XR vs. RF: ^#P < 0.05.

Figure 2

The expression level of JNK2 mRNA after siRNA transfection for 48 h. siRNA vs. Sham: ^*P < 0.05.

Figure 3

Protein levels of HSP10, HSP60, ClpP in BMSCs after exposure. (A) Western blot analysis of HSP10, HSP60, ClpP in BMSCs at 30 min post exposure. (B) Relative expression on level of HSP10, HSP60, ClpP protein at 30 min post exposure. (C) Western blot analysis of HSP10, HSP60, ClpP in BMSCs at 4 h post exposure. (D) Relative expression on level of HSP10, HSP60, ClpP protein at 4 h post exposure. (E) Western blot analysis of HSP10, HSP60, ClpP in BMSCs at 24 h post exposure. (F) Relative expression on level of HSP10, HSP60, ClpP protein at 24 h post exposure. SH, Sham; RF, RF-exposed; XR, X-rays irradiated; si + SH, siRNA + Sham; si + RF, siRNA + RF-exposed; si + XR, siRNA + X-rays irradiated. RF and XR vs. SH: ^*P < 0.05; si + RF vs. RF: ^#P < 0.05.