Effect of Extremely Low Frequency Electromagnetic Fields (EMF) on Phospholipase Activity in the Cultured Cells

Abstract

This study was conducted to investigate the effects of extremely low frequency electromagnetic fields (EMF) on signal pathway in plasma membrane of cultured cells (RAW 264.7 cells and RBL 2H3 cells), by measuring the activity of phospholipase A(2) (PLA(2)), phospholipase C (PLC) and phospholipase D (PLD). The cells were exposed to the EMF (60 Hz, 0.1 or 1 mT) for 4 or 16 h. The basal and 0.5 µM melittin-induced arachidonic acid release was not affected by EMF in both cells. In cell-free PLA(2) assay, we failed to observe the change of cPLA(2) and sPLA(2) activity. Also both PLC and PLD activities did not show any change in the two cell lines exposed to EMF. This study suggests that the exposure condition of EMF (60 Hz, 0.1 or 1 mT) which is 2.4 fold higher than the limit of occupational exposure does not induce phospholipases-associated signal pathway in RAW 264.7 cells and RBL 2H3 cells.

Keywords: Arachidonic acid; EMF; Phospholipase A2; Phospholipase C; Phospholipase D.

Fig. 1

The changes of basal (A) and 0.5 µM melittin-induced [³H]AA release (B) in RAW 264.7 cells. The cells were exposed to EMF (60 Hz, 0.1 or 1 mT) for 4 or 16 h and were labeled with [³H]AA for 2 h. The radioactivity of released [³H]AA was measured in the presence or absence of 0.5 µM melittin. Results are indicated in mean±S.D. from four separate experiments.

Fig. 2

The changes of basal (A) and 0.5 µM melittin-induced [³H]AA release (B) in RBL 2H3 cells. The cells were exposed to EMF (60 Hz, 0.1 or 1 mT) for 4 or 16 h and were labeled with [³H]AA for 2 h. The radioactivity of released [³H]AA was measured in the presence or absence of 0.5 µM melittin. Results indicate mean±S.D. from four separate experiments.

Fig. 3

Cell-derived PLA₂ activity in the presence of 5 mM CaCl₂. PLA₂ (25 µg protein) derived from RAW 264.7 cells and RBL 2H3 cells was incubated with 1-palmitoyl-2-[¹⁴C]arachidonyl phosphatidylcholine in the presence of 10 µM AACOCF₃ (cPLA₂ inhibitor) or 1 mM DTT (sPLA₂ inhibitor) and in the absence of CaCl₂. Results indicate mean±S.D. from four separate experiments. ^*Significantly different from Control (p<0.05).

Fig. 4

Effect of EMF on cell-derived cPLA₂ activity. Cell-derived cPLA₂ was obtained from the RAW 264.7 cells (A) and RBL 2H3 cells (B) exposed to EMF (60 Hz, 0.1 or 1 mT) for 4 or 16 h. cPLA₂ (25 µg protein) was incubated with 1-palmitoyl-2-[¹⁴C]arachidonyl phosphatidylcholine in the presence of 5 mM CaCl₂ and 1 mM DTT. Results indicate mean±S.D. from four separate experiments.

Fig. 5

Effect of EMF on cell-derived sPLA₂ activity. Cell-derived sPLA₂ was obtained from the RAW 264.7 cells (A) and RBL 2H3 cells (B) exposed to EMF (60 Hz, 0.1 or 1 mT) for 4 or 16 h. sPLA₂ (100 µg protein) was incubated with 10-pyren phosphatidylcholine. Results indicate mean±S.D. from four separate experiments.

Fig. 6

Effect of EMF on cell-derived PLC activity. Cell-derived PLC was obtained from the RAW 264.7 cells (A) and RBL 2H3 cells (B) exposed to EMF (60 Hz, 0.1 or 1 mT) for 4 or 16 h. PLC (25 µg protein) was incubated with [³H]phosphatidylinositol. Results indicate mean±S.D. from four separate experiments.

Fig. 7

Effect of EMF on PLD activity. RAW 264.7 cells (A) and RBL 2H3 cells (B) were exposed to EMF (60 Hz, 1 mT) for 4 or 16 hand were labeled with [³H]oleic acid, for 3 h. The radioactivity of [³H]phosphatidylethanol produced by PLD was measured in the presence or absence of 1 µM PMA. Results indicate mean±S.D. from four separate experiment.