Wednesday, April 15, 2015

Low-Frequency Electromagnetic Field Exposure Enhances Extracellular Trap Formation by Human Neutrophils through the NADPH Pathway

Low-Frequency Electromagnetic Field Exposure Enhances Extracellular Trap Formation by Human Neutrophils through the NADPH Pathway

Golbach LA, Scheer MH, Cuppen JJ, Savelkoul H, Verburg-van Kemenade BM. Low-Frequency Electromagnetic Field Exposure Enhances Extracellular Trap Formation by Human Neutrophils through the NADPH Pathway. J Innate Immun. 2015 Apr 10. [Epub ahead of print]


Low-frequency (LF) electromagnetic fields (EMFs) are abundantly present in modern society, and the potential biological consequences of exposure to these fields are under intense debate. Immune cells are suggested as possible target cells, though a clear mechanism is lacking.

Considering their crucial role in innate immune activation, we selected an ex vivo exposure set-up with human neutrophils to investigate a possible correlation between neutrophil extracellular trap (NET) formation and LF EMF exposure.

Our study shows that formation of NETs is enhanced by LF EMF exposure. Enhanced NET formation leads to increased antimicrobial properties as well as damage to surrounding cells. We found that LF-EMF-induced NET formation is dependent on the NADPH oxidase pathway and production of reactive oxygen species. Additionally, LF EMF exposure does not influence autophagy and PAD4 activity.

Our study provides a mechanism by which exposure to LF EMFs could influence the innate immune system.
Open Access Paper:


Environmental exposure to low-frequency (LF) electromagnetic fields (EMFs) produced by household appliances, communication equipment and power lines is increasing. Consequently, societal concern regarding potential adverse health effects of chronic exposure to LF EMFs is rising [1]. Reported effects of LF EMFs on biological systems range from harmful interactions [2] to beneficial effects [3]. No precise mechanism of action of LF EMFs has yet been elucidated, although immune cell activation through receptor interactions, calcium mobilisation and free radical production has been suggested [4]. Mechanistic studies to elucidate the biological response to LF EMF exposure so far mainly focused on macrophages and monocytes [3,4,5]. Neutrophils, on the other hand, were not studied extensively in this context. Neutrophils, however, fulfil a key role in the innate immune system by rapid eradication of invading pathogens via several strategies. They quickly migrate towards an infection and dominate the infected site already after a couple of minutes [6]. They are highly reactive, mobile and sensitive cells, which makes them putative targets to investigate possible cell modulation by LF EMF exposure [4].

Initially, phagocytosis of pathogens and degranulation of lytic enzymes were considered to be the only two antimicrobial strategies neutrophils used, but in 2004 a third special defence mechanism was described: neutrophil extracellular trap (NET) formation [7]. During NET formation, nuclear DNA is mixed with antimicrobial proteins to release an antimicrobial extracellular trap that captures and destroys extracellular microbes. Recent studies showed the importance of NADPH oxidase activity and consequential reactive oxygen species (ROS) formation that triggers dissociation of the peptide complexes, which contain the antimicrobial proteins [8,9]. Subsequently, peptidylarginine deiminase 4 (PAD4) [10] decondensates the chromatin, followed by disintegration of the nuclear envelop through autophagy to facilitate intermingling of these antimicrobial peptides with chromosomal DNA [11]. These findings underscore that NET formation involves specific signal transduction pathways and is tightly regulated to ensure a well-balanced innate immune response. NET formation requires an active NADPH oxidase complex to generate large amounts of superoxide anion radicals (O2-) that are rapidly dismutated to hydrogen peroxide (H2O2). Patients with chronic granulomatous disease highlight the importance of functional NADPH oxidase activity, since these patients fail to maintain phagocytosis or generate NETs [12]. Even though NET formation involves activation of intracellular signalling pathways that have been linked to the effects of LF EMFs, such as ROS production and NADPH oxidase activation [13,14], a correlation between NET formation by neutrophils and LF EMF exposure has not been studied yet. In an ex vivo experimental set-up we show that LF EMF exposure is able to enhance NET formation in neutrophils for which the NADPH pathway and subsequent ROS production are essential. These NETs are able to capture and kill bacteria, as well as damage epithelial cells, thereby indicating that NET formation by neutrophils is a vital but also delicately regulated process, which may be modulated by LF EMF exposure. Our study provides a reliable mechanism by which exposure to LF EMFs could influence the innate immune system.

Cell exposure was performed with a custom-made exposure system (Immunent BV, Veldhoven, The Netherlands), which fits inside a standard cell culture incubator to ensure optimal cell culture conditions with 37 ± 0.2°C and 5% CO2. An irregular combination of four block waves, described previously [16], with frequencies of 320, 730, 880 and 2,600 Hz was applied at a magnetic field intensity of 300 µT. Sham treatment consisted of identical culture conditions with a unenergised coil in a second identical incubator.

This is the first study to show that LF EMF exposure is able to significantly enhance NET formation ex vivo. Cellular activation by PMA was needed, since LF EMF exposure alone was not able to promote NET formation. The pathways involved in PMA-stimulated NET formation are partly known [32]; with the use of selective pharmacological inhibitors, we demonstrated that the NADPH pathway is crucial for LF-EMF-enhanced NET formation, possibly by upregulated ROS production. These data provide a basic mechanism of action on immune cells to explain potential health effects of LF EMFs generated by household appliances and power lines.


Joel M. Moskowitz, Ph.D., Director
Center for Family and Community Health
School of Public Health
University of California, Berkeley

Electromagnetic Radiation Safety

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