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In the last years, radiofrequency (RF) has demonstrated that it can reduce DNA damage induced by a subsequent treatment with chemical or physical agents in different cell types, resembling the adaptive response, a phenomenon well documented in radiobiology. Such an effect has also been reported by other authors both in vitro and in vivo, and plausible hypotheses have been formulated, spanning from the perturbation of the cell redox status, to DNA repair mechanisms, and stress response machinery, as possible cellular mechanisms activated by RF pre-exposure. These mechanisms may underpin the observed phenomenon, and require deeper investigations. The present study aimed to determine whether autophagy contributes to RF-induced adaptive response. To this purpose, SH-SY5Y human neuroblastoma cells were exposed for 20 h to 1950 MHz, UMTS signal, and then treated with menadione. The results obtained indicated a reduction in menadione-induced DNA damage, assessed by applying the comet assay. Such a reduction was negated when autophagy was inhibited by bafilomycin A1 and E64d. Moreover, CRISPR SH-SY5Y cell lines defective for ATG7 or ATG5 genes did not show an adaptive response. These findings suggest the involvement of autophagy in the RF-induced adaptive response in human neuroblastoma cells; although, further investigation is required to extend such observation at the molecular level.

In previous studies we demonstrated that radiofrequency (RF) electromagnetic fields (EMF) is able to reduce DNA damage induced by a subsequent treatment with genotoxic agents, resembling the adaptive response, a phenomenon well known in radiobiology. In this study we report on the capability of the culture medium from SH-SY5Y neuroblastoma cells exposed to 1950 MHz to elicit, in recipient non-exposed cells, a reduction of menadione-induced DNA damage (P < 0.05; comet assay), indicating the capability of non-ionizing radiation to elicit a bystander effect. A comparable reduction was also detected in cultures directly exposed to the same EMF conditions (P < 0.05), confirming the adaptive response. In the same exposure conditions, we also evidenced an increase of heat shock protein 70 (hsp70) in culture medium of cells exposed to RF with respect to sham exposed ones (P < 0.05; western blot analysis), while no differences were detected in the intracellular content of hsp70. On the whole, our results evidence a protective effect of RF against menadione-induced DNA damage in directly and non-directly exposed cells, and suggest hsp70 pathway to be investigated as one of the potential candidate underpinning the interaction between RF exposure and biological systems.

Exposure to radiofrequency (RF) electromagnetic fields (EMF) (frequencies of 100 kHz to 300 GHz) has been steadily increasing. In addition to heat-related effects of RF EMF, other yet-unspecified biological effects, might exist which could possibly lead to health effects. Given the large number of health endpoints that have been studied, we wanted to prioritize those that would merit systematic reviews. We developed a survey listing of all health endpoints reported in the literature and we asked 300 RF EMF experts and researchers to prioritize these health effects for systematic review as critical, important or unimportant. We also asked the experts to provide the rationale for their prioritization. Of the 300 RF EMF experts queried, 164 (54%) responded. They rated cancer, heat-related effects, adverse birth outcomes, electromagnetic hypersensitivity, cognitive impairment, adverse pregnancy outcomes and oxidative stress as outcomes most critical regarding RF EMF exposure. For these outcomes, systematic reviews are needed. For heat-related outcomes, the experts based their ranking of the critical outcomes on what is known from human or animal studies, and for cancer and other outcomes, they based their rating also on public concern. To assess health risks of an exposure in a robust manner, it is important to prioritize the health outcomes that should be systematically reviewed. Here we have shown that it feasible to do so in an inclusive and transparent way.

Background Exposure to radiofrequency electromagnetic fields (RF-EMF, 100 kHz – 300 GHz) emitted by wireless communication technologies is pervasive and ubiquitous. Concern has been raised about possible adverse effects to human health. In 2011 the International Agency for Research on Cancer has classified RF-EMF as possibly carcinogenic to humans, highlighting that the evidence is weak and far from conclusive. Updated systematic reviews of the scientific literature on this topic are lacking, especially for mechanistic studies. Objectives To develop a protocol for a systematic review of experimental studies investigating genotoxic effects induced by RF-EMF in in vitro cellular models. Genotoxicity is one of the key-biological indicators of carcinogenicity, and the most common characteristics of established carcinogens. The predefined procedures for conducting the systematic review are outlined below. Methods We will follow the guidelines developed by the National Toxicology Program-Office of Health Assessment and Translation (NTP-OHAT), adapted to the evaluation of in vitro studies. Eligibility criteria We will include experimental in vitro studies addressing the relationship between controlled exposures to RF-EMF and genotoxicity in mammalian cells only. Eligibility for inclusion will be further restricted to peer reviewed articles reporting findings from primary studies. Information sources We will search the scientific literature databases NCBI PubMed, Web of Science, and EMF-Portal. No filter on publication date will be applied. Only studies published in English will be considered. The reference lists of the included papers and available reviews will be screened for unidentified relevant papers. References will be managed through Endnote X9 software. Data extraction and synthesis of results Data from included papers will be extracted according to predefined forms. Heterogeneity within the available evidence will determine the type of evidence synthesis that is appropriate. Findings will be summarized in tables, graphical displays and in a narrative synthesis of the available evidences. A meta-analysis will be carried out if subgroups of studies homogeneous in terms of exposure characteristics, endpoint, and cell types will be identified. Risk of bias The internal validity of included studies will be assessed using the NTP-OHAT Risk of Bias Rating Tool for animal studies, adapted to in vitro studies. This stage of the process will be managed through the Health Assessment Workspace Collaborative (HAWC). Evidence appraisal To rate confidence in the body of evidence, we will use the OHAT GRADE-based approach for animal studies. Framework and funding This protocol concerns one of the evidence streams considered in a larger systematic review of the scientific literature on the potential carcinogenicity of RF-EMF, performed by scientists from several Italian public research agencies. The project is supported by the Italian Workers’ Compensation Authority (INAIL) in the framework of the CRA with the Istituto Superiore di Sanita “BRiC 2018/06 – Scientific evidence on the carcinogenicity of radiofrequency electromagnetic fields”.

Magnetic resonance imaging (MRI) is one of the most-used diagnostic imaging methods worldwide. There are ~50,000 MRI scanners worldwide each of which involves a minimum of five workers from different disciplines who spend their working days around MRI scanners. This review analyzes the state of the art of literature about the several aspects of the occupational exposure to electromagnetic fields (EMF) in MRI: regulations, literature studies on biological effects, and health surveillance are addressed here in detail, along with a summary of the main approaches for exposure assessment. The original research papers published from 2013 to 2021 in international peer-reviewed journals, in the English language, are analyzed, together with documents published by legislative bodies. The key points for each topic are identified and described together with useful tips for precise safeguarding of MRI operators, in terms of exposure assessment, studies on biological effects, and health surveillance. Graphical abstract: [Figure not available: see fulltext.]

The last decades have seen a huge increase in applications and devices using and emitting non-ionizing radiation, otherwise referred to as “electromagnetic fields” (EMF) [...]

In previous investigations, we demonstrated that pre-exposure of different cell cultures to radiofrequency fields can reduce the damage induced by genotoxic agents, an effect resembling the so-called adaptive response. In this study, we pre-exposed human peripheral blood lymphocytes and Chinese hamster lung fibroblast cell line to 1950 MHz, UMTS (Universal Mobile Telecommunication System) signal, for 20 h, and then treated cultures with Mitomycin-C. After confirming the induction of an adaptive response in terms of the reduction of micronuclei formation, we observed that such a response was negated by treatments with 3-aminobenzamide. Since 3-aminobenzamide is an inhibitor of poly (ADP-ribose) polymerase enzyme, which is involved in DNA repair, these results support the possible involvement of DNA repair mechanisms in radiofrequency-induced adaptive response.

In questo intervento verrà presentata una analisi degli studi sperimentali condotti in vitro e in vivo volti alla valutazione degli effetti di esposizioni a campi elettromagnetici nel range delle onde millimetriche, incluse le frequenze che saranno impiegate per la tecnologia 5 G. Verranno descritti i risultati disponibili in letteratura con l'obiettivo di valutare se esistono evidenze di una risposta, robusta e riproducibile, nei sistemi biologici in condizioni di regime non termico e che siano in grado di fornire indicazioni sul possibile rischio per la salute umana. Inoltre, verranno anche considerate le implicazioni di tali effetti per le applicazioni diagnostiche e terapeutiche. Infine sarà discussa la qualità della ricerca in bioelettromagnetismo, che rappresenta un punto critico nella determinazione del peso dell'evidenza per la valutazione del rischio associato all'esposizione.

The occurrence of modulation-specific effects after co-exposures to radio-frequency (RF) and other agents has been discussed in the literature. In this paper, the influence of modulation and bandwidth in eliciting the DNA damage of RF alone and in combination with mitomycin-C (MMC) is analyzed in human lymphocytes. Blood cultures from healthy donors were exposed to 1950 MHz, and continuous wave (CW), wideband direct-sequence code division multiple access (WCDMA, 4.5 MHz bandwidth), and additive white Gaussian noise (AWGN, 9 MHz bandwidth) signals were considered. For each signal, specific absorption rate (SAR) values of 0.15, 0.3, 0.6, 1.25 W/kg were tested. RF exposure alone never induced DNA damage in the micronucleus assay. When RF exposure was followed by MMC treatment, the effect depended on modulation and bandwidth. CW exposure never altered the MMC-induced DNA damage, while such damage was reduced when either signals WCDMA at 0.3 W/kg SAR or AWGN at 0.15 and 0.3 W/kg were applied. These results indicate the influence of modulation for the occurrence of the protective effect, with a relation between the bandwidth and the power absorbed by samples. If confirmed in vivo , clinical applications using modulated RF signals could be devised to protect cells from side effects of therapeutic treatments.

La diffusione di tecnologie basate sull'utilizzo di campi elettromagnetici (CEM) negli ambienti di lavoro, ha determinato un crescente interesse verso lo sviluppo di strumenti e metodi per la valutazione del rischio per i lavoratori sottoposti a tale tipologia di esposizione. La direttiva definisce un nuovo sistema di limiti e valori di azione, e prescrive l'obbligo per i datori di lavoro di effettuare la valutazione del rischio, e di informare e formare i lavoratori rispetto all'esito di tale valutazione. I sistemi per Imaging a Risonanza Magnetica (MRI) sono sorgenti di esposizioni multiple a CEM: campi di induzione magnetica statici (fino ad alcuni T), CM di bassa frequenza (campi di gradiente) e CEM impulsivi a radiofrequenza (RF). Gli scenari di esposizione dipendono dall'attività svolta (tecnici, infermieri, medici, addetti alla pulizia o alla manutenzione) e dal layout dell'impianto MRI. Dal 2012 al 2017 l'INAIL-CONTARP Regione Campania e il CNR-IREA hanno svolto un'intensa attività di valutazione del rischio a CM, bio-monitoraggio, formazione e informazione dei lavoratori in ambienti MRI, mettendo a punto strumenti e procedure che possono essere utilizzati dal datore di lavoro e dai responsabili per la sicurezza.

The acquired resistance by cells and organisms pre-exposed to low doses of ionizing radiation or selected genotoxic chemicals to the damaging effect of higher doses of the same or other agents, is commonly referred to as adaptive response (AR). It has been demonstrated in several cell types as well as in in vivo experiments, by measuring chromosomal and DNA damage, apoptosis, oxidative stress and survival. Despite a large amount of experimental results, several hypothesis have been proposed to explain this phenomenon but at the moment there is not yet an action mechanism universally accepted. Recently, our research group evidenced the capability of radiofrequency (RF) to offer protection against the effects of chemical and physical agents in different cell types, resembling the phenomenon of AR. Other research groups confirmed this observation also in in vivo studies. In this presentation, the results of RF-induced AR, collected so far, and the ongoing investigation aimed to characterize the phenomenon will be presented. Moreover, the potential action mechanisms involved in the RF-induced AR will be discussed.

This study aims to assess whether a 1950 MHz radiofrequency (RF) electromagnetic feld could protect human neuroblastoma SH-SY5Y cells against a subsequent treatment with menadione, a chemical agent inducing DNA damage via reactive oxygen species formation. Cells were pre-exposed for 20h to specifc absorption rate of either 0.3 or 1.25W/kg, and 3h after the end of the exposure, they were treated with 10µM menadione (MD) for 1h. No diferences were observed between sham- and RFexposed samples. A statistically signifcant reduction in menadione-induced DNA damage was detected in cells pre-exposed to either 0.3 or 1.25W/kg (P

Reversible electroporation is used to increase the uptake of chemotherapeutic drugs in local tumor treatment (electrochemotherapy) by applying the pulsing protocol (8 rectangular pulses, 1000 V/cm, 100 µs) standardized in the framework of the European Standard Operating Procedure on Electrochemotherapy multicenter trial. Currently, new electrochemotherapy strategies are under development to extend its applicability to tumors with different histology. Electrical parameters and drug type are critical factors. A possible approach is to test pulse parameters different from European Standard Operating Procedure on Electrochemotherapy but with comparable electroporation yield (European Standard Operating Procedure on Electrochemotherapy-equivalent protocols). Moreover, the use of non-toxic drugs combined with electroporation represents the new frontier for electrochemotherapy applications; calcium electroporation has been recently proposed as a simple tool for anticancer therapy. In vitro investigations facilitate the optimization of electrical parameters and drugs for in vivo and clinical testing. In this optimization study, new pulsing protocols have been tested by increasing the pulse number and reducing the electric field with respect to the standard. European Standard Operating Procedure on Electrochemotherapy-equivalent protocols have been identified in HL-60 and A431 cancer cell models, and a higher sensitivity in terms of electroporation yield has been recorded in HL-60 cells. Moreover, cell killing efficacy of European Standard Operating Procedure on Electrochemotherapy-equivalent protocols has been demonstrated in the presence of increasing calcium concentrations on both cell lines. Equivalent European Standard Operating Procedure on Electrochemotherapy protocols can be used to optimize the therapeutic effects in the clinic, where different regions of the same cancer tissue, with different electrical properties, might result in a differential electroporation yield of the standard protocol over the same tissue, or, eventually, in an override of the operational limits of the instrument. Moreover, using calcium can help overcome the drawbacks of standard drugs (side effects, high costs, difficult handling, preparation, and storage procedures). These results support the possibility of new treatment options in both standard electrochemotherapy and calcium electroporation, with clear advantages in the clinic.

In this paper, we study the influence of radiofrequency signal modulation and bandwidth, in eliciting the genotoxic effect of radiofrequency per se and in combination with Mitomycin-C, a chemical mutagen, in a combined exposure protocol on cultured human lymphocytes.

ELF-MF exposures have been described to modulate oxidative responses. A recent review indicated changes in oxidative status as an early response to ELF-MF, although the interaction mechanisms are still lacking. Here we aim to address Reactive Oxygen Species formation in immune relevant cells during ELF-MF exposure, and to investigate if intracellular localized temperature increase (hot spots) is involved.

Pulsed electric fields (PEFs) for permeabilization of biological membranes have gained increasing consensus in the last decades for medical and industrial applications. Since ten years, our research groups have been studying the basic mechanisms of cellular responses to microsecond and nanosecond PEFs, by developing innovative experimental and modeling approaches

An ongoing research project aimed to investigate if autophagy is involved in the radiofrequency-induced adaptive response will be presented. It is funded by the ANSES and the activities, carried out in cooperation between Italian and France laboratories, will be described

I sistemi per Imaging a Risonanza Magnetica (MRI) costituiscono una sorgente di esposizioni multiple a campi elettromagnetici (CEM) per gli operatori del settore. La valutazione del rischio da esposizione ai CEM è particolarmente complessa considerando anche i diversi scenari di esposizione che dipendono dall'attività specifica svolta (tecnici, infermieri, medici, addetti alla pulizia o alla manutenzione), oltre che dal layout dell'impianto MRI. In questo lavoro verranno presentati i risultati di uno studio pilota condotto sui lavoratori MRI dell'Istituto Nazionale Tumori, Fondazione G. Pascale di Napoli, con il duplice obiettivo di valutare i livelli di esposizione e di identificare potenziali biomarcatori per la valutazione del rischio.

This study aims to assess whether a 1950 MHz radiofrequency (RF) electromagnetic field could protect human neuroblastoma SH-SY5Y cells against a subsequent treatment with menadione, a chemical agent inducing DNA damage via reactive oxygen species formation. Cells were pre-exposed for 20 h to specific absorption rate of either 0.3 or 1.25 W/kg, and 3 h after the end of the exposure, they were treated with 10 µM menadione (MD) for 1 h. No differences were observed between sham- and RF-exposed samples. A statistically significant reduction in menadione-induced DNA damage was detected in cells pre-exposed to either 0.3 or 1.25 W/kg (P

Misure di campi magnetici di gradiente con metodo del picco ponderato presso impianti MRI di ospedali Campani. Bio-monitoraggio dei lavoratori esposti e di soggetti di controllo mediante analisi del danno cromosomico in campioni di mucosa buccale. Diffusione e disseminazione dei risultati in strutture pubbliche e private della Regione Campania

In questa comunicazione saranno presentate e discusse le applicazioni biomedicali dei campi elettromagnetici non ionizzanti, dalle basse frequenze alle radiofrequenze. Dopo una introduzione di carattere generale, sarà presentata una panoramica della letteratura sull'argomento, inclusi i più recenti risultati. Infine, verranno illustrati i risultati sperimentali della recente letteratura scientifica riguardanti nuove potenziali applicazioni non ancora impiegate nella pratica clinica.

In this study, the effect of radiofrequency (RF) exposure to 1950 MHz, Universal Mobile Telecommunication System signal, was investigated in Chinese hamster lung fibroblast cell line (V79). Genotoxic and cytotoxic effects of 20-h exposure at specific absorption rate (SAR) values from 0.15 W/kg to 1.25 W/kg were measured by means of cytokinesis-block micronucleus (MN) assay. Exposure was carried out blinded under strictly controlled conditions of dosimetry and temperature. The effect of RF exposure alone at four SAR values was tested, that is, 0.15, 0.3, 0.6, and 1.25 W/kg. A statistically significant increase in MN frequency was found in cultures exposed to 0.15 and 0.3 W/kg (P < 0.05) compared to sham-exposed ones, in the absence of cytotoxicity. SAR values of 0.6 and 1.25 W/kg did not exert any effect. Moreover, to evaluate the ability of RF to exert protective effects with respect to a chemical mutagen, cell cultures were also pre-exposed for 20 h at 0.3 or 1.25 W/kg, and then treated with 500 ng/ml of mitomycin-C (MMC). A significant reduction in the frequency of MN was detected in cultures pre-exposed to 1.25 W/kg compared to cultures treated with MMC alone (P < 0.05), indicating induction of adaptive response. Such a decrease was not induced by pre-exposure at 0.3 W/kg SAR. Taken together, our results indicated that V79 is a sensitive cell model to evidence either adverse or beneficial effects of RF exposure, depending on experimental conditions applied.

L'analisi dell'incidenza di micronuclei nelle cellule di mucosa buccale viene impiegata in studi di epidemiologia molecolare volti a valutare l'impatto dei fattori legati sia allo stile di vita sia all'esposizione occupazionale ad agenti chimici e fisici a potenziale effetto genotossico. Infatti, l'aumento dell'incidenza di micronuclei è stato associato al rischio di invecchiamento precoce, cancro e malattie neurodegenerative. La finalità di questo lavoro è verificare se la valutazione dell'incidenza di micronuclei in cellule di mucosa buccale può essere impiegata come bio-marcatore di rischio per i lavoratori in ambiente MRI. A questo scopo, mediante la compilazione di un questionario ad-hoc, sono stati reclutati 12 lavoratori in ambiente MRI, selezionati sulla base dell'età, sesso, stato di salute, stile di vita (fumo, consumo di alcool) ed esposizioni professionali a cancerogeni chimici o fisici. Inoltre, è stato selezionato un gruppo di 12 lavoratori non esposti (controllo) da confrontare agli esposti. Sebbene sia stata riscontrata una alta variabilità tra i lavoratori esposti, i risultati ottenuti indicano un lieve aumento di micronuclei che però, confrontato con il gruppo di controllo, non era statisticamente significativo. Al fine di validare l'impiego di questo test come marcatore di rischio per i lavoratori in ambiente MRI, sarebbe necessario aumentare il numero di donatori da coinvolgere nello studio.

Lo staff di Risonanza Magnetica è continuamente esposto al campo magnetico statico e spazialmente eterogeneo presente in ogni momento nella sala RM. E' importante che i lavoratori siano costantemente informati sui rischi associati all'esposizione ai Campi Elettromagnetici. A questo scopo è stato creato un portale web (www.mriworkers.eu ), nato da un'idea degli istituti IFC ed IREA del CNR, che si configura come uno strumento interattivo attraverso il quale ogni operatore di Risonanza Magnetica può avere a disposizione informazioni tecniche, normative vigenti, elenco di "buone norme" e di comportamenti da evitare.

The acquired resistance by cells and organisms pre-exposed to low doses of ionizing radiation or selected genotoxic chemicals to the damaging effect of higher doses of the same or other agents, is commonly referred to as adaptive response (AR) It has been demonstrated in several cell types as well as in vivo experiments, by measuring chromosomal and DNA damage, apoptosis, oxidative stress and survival. Although AR has been repeatedly demonstrated to be induced by different genotoxicants, under variable time schedule and biomarkers and in different cell types, the basic mechanisms still remain to be elucidated. Recently, our research group evidenced the capability of radiofrequency (RF) to induce AR by offering protection against the effects of chemical and physical agents in different cell types. Other research groups confirmed this observation also in in vivo studies. In this presentation, the results of RF-induced AR, collected so far, and the ongoing investigation aimed to characterize the phenomenon and elucidate the interaction mechanisms will be presented. Moreover, the interesting perspectives of such phenomenon in the field of radio-protection and biomedical applications will be discussed.

It has been shown that magnetic fields in the extremely low frequency range (ELF-MF) can act as a stressor in various in vivo or in vitro systems, at flux density levels below those inducing excitation of nerve and muscle cells, which are setting the limits used by most generally accepted exposure guidelines, such as the ones published by the International Commission on Non-Ionizing Radiation Protection. In response to a variety of physiological and environmental factors, including heat, cells activate an ancient signaling pathway leading to the transient expression of heat shock proteins (HSPs), which exhibit sophisticated protection mechanisms. A number of studies suggest that also ELF-MF exposure can activate the cellular stress response and cause increased HSPs expression, both on the mRNA and the protein levels. In this review, we provide some of the presently available data on cellular responses, especially regarding HSP expression, due to single and combined exposure to ELF-MF and heat, with the aim to compare the induced effects and to detect possible common modes of action. Some evidence suggests that MF and heat can act as costressors inducing a kind of thermotolerance in cell cultures and in organisms. The MF exposure might produce a potentiated or synergistic biological response such as an increase in HSPs expression, in combination with a well-defined stress, and in turn exert beneficial effects during certain circumstances.

Electrochemotherapy (ECT) is a clinical procedure for the local treatment of solid tumors, which combines the application of pulsed electric fields (PEFs) and chemotherapeutic drugs, by exploiting the PEF-induced membrane permeabilization (electroporation, EP). The ESOPE pulsing protocol (8 pulses with 100 μs duration and 1 kV/cm electric field strength, given at 1 Hz or 5 kHz as repetition rate) is the only protocol approved for application on human patients and is currently used in the clinical practice. Moving from an experimental study comparing the efficacy of ESOPE to that of modified PEF protocol with lower electric field intensity and higher pulse number, in this paper an electrophysiological model is used to provide theoretical support to the experimental results. The pore density and the transmembrane voltage time behavior at the upper pole of a single spherical cell are considered, in order to assess the EP effect of the applied pulse train, considered as boundary condition in the field based solution of the problem. The simulation results highlight that, 40 pulses 750 V/cm is a good candidate as ESOPE equivalent protocol.

La risposta adattativa è un fenomeno ben documentato in radiobiologia: pre-trattamenti con dosi molto basse di agenti mutageni chimici o fisici inducono protezione da un trattamento successivo con una alta dose dello stesso agente o di altri agenti. Tale fenomeno è stato evidenziato sia in colture cellulari sia nell'uomo e in particolare nei lavoratori. La maggior parte degli studi ha impiegato dosi molto basse e dosi elevate di radiazioni ionizzanti, ma è stato anche dimostrato che agenti chimici o combinazioni di agenti fisici e chimici sono altrettanto efficaci nell'indurre adattamento. Sebbene sia disponibile una discreta quantità di risultati pubblicati e siano state formulate diverse ipotesi, ad oggi non esiste ancora un consolidato meccanismo di azione in grado di spiegare l'insorgenza del fenomeno. Nel 2009 il nostro gruppo di ricerca ha evidenziato per la prima volta che pre-esposizioni di colture di linfociti umani alle frequenze in uso per la telefonia cellulare sono in grado di offrire protezione dal danno indotto al DNA da mutageni chimici. Successivamente, lo studio è stato esteso ed i risultati ottenuti ad oggi hanno mostrato che in diversi tipi cellulari, diverse frequenze e modulazioni offrono protezione nei confronti del danno indotto da svariati mutageni chimici e da radiazioni ionizzanti. Tali risultati sono stati anche confermati da altri gruppi di ricerca internazionali sia in vitro sia in vivo. In questa relazione verranno illustrati i principali risultati ottenuti dal nostro gruppo di ricerca. Verranno inoltre evidenziate le conoscenze da acquisire nonché le implicazioni di tale fenomeno per la popolazione in generale e i lavoratori in particolare.

A numerical model, usually employed to analyze the electroporation phenomenon in eukaryotic cells exposed to high voltage electric pulses with duration in the ms to ?s time scale, is extended to investigate the electroporation onset and dynamics induced by nanosecond pulsed electric fields (nsPEFs). The model allows to track the primary events and dynamics of the electroporation process on a time scale where available experimental methods do not provide reliable information. In particular the model, solved in the time domain, provides the spatial and temporal distribution of the transmembrane voltage and pore density of fixed radii around the plasma membrane. Moreover, it allows to optimize the experimental design of biological investigations in the framework of the mechanistic understanding of nsPEF. The obtained simulation results are correlated to biological observations of plasma membrane permeabilization in human lymphoblastoid Jurkat T-cells exposed to nsPEFs with variable pulse amplitude, as assessed by YO-PRO1 dye uptake.

The aim of this preliminary investigation was to assess whether human peripheral blood lymphocytes which have been pre-exposed to non-ionizing radiofrequency fields exhibit an adaptive response (AR) by resisting the induction of genetic damage from subsequent exposure to ionizing radiation. Peripheral blood lymphocytes from four healthy donors were stimulated with phytohemagglutinin for 24 h and then exposed for 20 h to 1950 MHz radiofrequency fields (RF, adaptive dose, AD) at an average specific absorption rate of 0.3 W/kg. At 48 h, the cells were subjected to a challenge dose (CD) of 1.0 or 1.5 Gy X-irradiation (XR, challenge dose, CD). After a 72 h total culture period, cells were collected to examine the incidence of micronuclei (MN). There was a significant decrease in the number of MN in lymphocytes exposed to RF + XR (AD + CD) as compared with those subjected to XR alone (CD). These observations thus suggested a RF-induced AR and induction of resistance to subsequent damage from XR. There was variability between the donors in RF-induced AR. The data reported in our earlier investigations also indicated a similar induction of AR in human blood lymphocytes that had been pre-exposed to RF (AD) and subsequently treated with a chemical mutagen, mitomycin C (CD). Since XR and mitomycin-C induce different kinds of lesions in cellular DNA, further studies are required to understand the mechanism(s) involved in the RF-induced adaptive response.

International Expert Group Evaluations on Biological/Health Effects of Electromagnetic Fields

Alzheimer's disease involves the formation of plaques of amyloid beta (A?), that induce cellular damage, including oxidative DNA damage. This study aims to investigate the capability of RF exposure to reduce the A?-induced DNA damage in SH-SY5Y cells. To this purpose a dose-response curve of A? has been set up in terms of DNA migration, and preliminary results on the effect of RF are reported.

Pulsed power is applied to gases and liquids to produce plasmas; these may interact with living organisms. Plasma, known as the fourth state of matter, consists of charged and neutral particles and is characterized by a quasi-neutral state. Two types of plasma exist, nonthermal and thermal equilibrium. Nonthermal plasma, the type most frequently employed in the bioelectric field, has a gas temperature much lower than its electron temperature. The biocidal agents which may be produced by nonthermal plasmas are heat, electric fields, ultraviolet radiation, shock waves, and reactive chemical species. Additionally, secondary biological effects can be induced through post-discharge reactions of chemical species produced by plasma in gases and liquid.

To evaluate possible health risk related to occupational exposure in MRI, the micronucleus frequency in buccal mucosa cells was measured in workers and in controls. Pilot results indicate no differences between the two groups, but a larger number of donors is required. A procedure for the evaluation of occupational exposure to electromagnetic fields is being disseminated for workers formation.

The induction of an adaptive response (AR) was examined in human peripheral blood lymphocytes exposed to non-ionizing radiofrequency fields (RF). Cells from nine healthy human volunteers were stimulated for 24 h with phytohaemagglutinin and then exposed for 20 h to an adaptive dose (AD) of a 1950 MHz RF UMTS (universal mobile telecommunication system) signal used for mobile communications, at different specific absorption rates (SAR) of 1.25, 0.6, 0.3, and 0.15 W/kg. This was followed by treatment of the cells at 48 h with a challenge dose (CD) of 100 ng/ml mitomycin C (MMC). Lymphocytes were collected at the end of the 72 h total culture period. The cytokinesis-block method was used to record the frequency of micronuclei (MN) as genotoxicity end-point. When lymphocytes from six donors were pre-exposed to RF at 0.3 W/kg SAR and then treated with MMC, these cells showed a significant reduction in the frequency of MN, compared with the cells treated with MMC alone: this result is indicative of induction of AR. The results from our earlier study indicated that lymphocytes that were stimulated for 24 h, exposed for 20 h to a 900 MHz RF GSM (global system for mobile communication) signal at 1.25 W/kg SAR and then treated with 100 ng/ml MMC, also exhibited AR. These overall data suggest that the induction of AR depends on RF frequency, type of the signal and SAR. Further characterization of RF-induced AR is in progress.

International audience; It has sometimes been assumed that children are more sensitive than adults to the effects of radiofrequency (RF) fields associated with cellular wireless telephones. However, relatively few in vitro or animal models have examined this possibility. In vitro studies have used several cell types, from both humans and rodents, including primary cells, embryonic cell lines, undifferentiated cancer cell lines, and stem cells. Overall, the balance of evidence does not suggest that field-related effects occur in any cell type: gene and protein expression were not significantly changed by exposure in nine out of 15 studies; genotoxicity was evaluated in 13 papers and in most, of these studies, no damage to DNA was detected; eight studies failed to demonstrate induction of apoptosis; and three studies reported lack of oxidative stress induction by RF-exposures. Five of eight studies investigating the effects of combined exposures to RF fields and chemical or physical agents reported a lack of field-related effects. In addition, few papers have been published on the effects of low level exposure of immature animals. The available results are very limited, both in terms of signals used and biological endpoints investigated, but the evidence does not indicate that prenatal or early postnatal exposures are associated with acute adverse responses or the development of detrimental changes in the long-term. Overall, this suggests that young animals may not be significantly more sensitive than adults, but there is clearly a need for further studies to be carried out.

Intense nanosecond pulsed electric fields (nsPEFs) have been shown to induce, on intracellular structures, interesting effects dependent on electrical exposure conditions (pulse length and amplitude, repetition frequency and number of pulses), which are known in the literature as "bioelectrical effects" (Schoenbach et al., IEEE Trans Plasma Sci 30:293-300, 2002). In particular, pulses with a shorter width than the plasma membrane charging time constant (about 100 ns for mammalian cells) can penetrate the cell and trigger effects such as permeabilization of intracellular membranes, release of Ca(2+) and apoptosis induction. Moreover, the observed effects have led to exploration of medical applications, like the treatment of melanoma tumors (Nuccitelli et al., Biochem Biophys Res Commun 343:351-360, 2006). Pulsed electric fields allowing such effects usually range from several tens to a few hundred nanoseconds in duration and from a few to several tens of megavolts per meter in amplitude (Schoenbach et al., IEEE Trans Diel Elec Insul 14:1088-1109, 2007); however, the biological effects of subnanosecond pulses have been also investigated (Schoenbach et al., IEEE Trans Plasma Sci 36:414-422, 2008). The use of such a large variety of pulse parameters suggests that highly flexible pulse-generating systems, able to deliver wide ranges of pulse durations and amplitudes, are strongly required in order to explore effects and applications related to different exposure conditions. The Blumlein pulse-forming network is an often-employed circuit topology for the generation of high-voltage electric pulses with fixed pulse duration. An innovative modification to the Blumlein circuit has been recently devised which allows generation of pulses with variable amplitude, duration and polarity. Two different modified Blumlein pulse-generating systems are presented in this article, the first based on a coaxial cable configuration, matching microscopic slides as a pulse-delivery system, and the other based on microstrip transmission lines and designed to match cuvettes for the exposure of cell suspensions.

This study was designed to assess if radiofrequency (RF) radiation induces oxidative stress in cultured mammalian cells when given alone or in combination with ferrous ions (FeSO4). For this purpose the production of reactive oxygen species (ROS) was measured by flow cytometry in human lymphoblastoid cells exposed to 1950 MHz signal used by the third generation wireless technology of the Universal Mobile Telecommunication System (UMTS) at Specific Absorption Rate of 0.5 and 2.0 W/kg. Short (5–60 min) or long (24 h) duration exposures were carried out in a waveguide system under strictly controlled conditions of both dosimetry and environment. Cell viability was also measured after 24 h RF exposure using the Resazurin and Neutral Red assays. Several co-exposure protocols were applied to test if RF radiation is able to alter ROS formation induced by FeSO4 (RF given before or concurrently to FeSO4). The results obtained indicate that non-thermal RF exposures do not increase spontaneous ROS formation in any of the experimental conditions investigated. Consistent with the lack of ROS production, no change in cell viability was observed in Jurkat cells exposed to RF radiation for 24 h. Similar results were obtained when co-exposures were considered: combined exposures to RF radiation and FeSO4 did not increase ROS formation induced by the chemical treatment alone. In contrast, in cultures treated with FeSO4 as positive control, a dose-dependent increase in ROS formation was recorded, validating the sensitivity of the method employed. Bioelectromagnetics 30:525–535, 2009. © 2009 Wiley-Liss, Inc.

The incidence of micronuclei was evaluated to assess the induction of an adaptive response to non-ionizing radiofrequency (RF) radiation in peripheral blood lymphocytes collected from five different human volunteers. After stimulation with phytohemagglutinin for 24 h, the cells were exposed to an adaptive dose of 900 MHz RF radiation used for mobile communications (at a peak specific absorption rate of 10 W/kg) for 20 h and then challenged with a single genotoxic dose of mitomycin C (100 ng/ml) at 48 h. Lymphocytes were collected at 72 h to examine the frequency of micronuclei in cytokinesis-blocked binucleated cells. Cells collected from four donors exhibited the induction of adaptive response (i.e., responders). Lymphocytes that were pre-exposed to 900 MHz RF radiation had a significantly decreased incidence of micronuclei induced by the challenge dose of mitomycin C compared to those that were not pre-exposed to 900 MHz RF radiation. These preliminary results suggested that the adaptive response can be induced in cells exposed to non-ionizing radiation. A similar phenomenon has been reported in cells as well as in animals exposed to ionizing radiation in several earlier studies. However, induction of adaptive response was not observed in the remaining donor (i.e., non-responder). The incidence of micronuclei induced by the challenge dose of mitomycin C was not significantly different between the cells that were pre-exposed and unexposed to 900 MHz RF radiation. Thus the overall data indicated the existence of heterogeneity in the induction of an adaptive response between individuals exposed to RF radiation and showed that the less time-consuming micronucleus assay can be used to determine whether an individual is a responder or non-responder.

The aim of this study was to investigate DNA damage in human dermal fibroblasts from a healthy subject and from a subject affected by Turner's syndrome that were exposed for 24 h to radiofrequency (RF) radiation at 900 MHz. The RF-radiation exposure was carried out alone or in combination with 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX), a well-known environmental mutagen and carcinogen produced during the chlorination of drinking water. Turner's syndrome fibroblasts were also exposed for a shorter time (1 h). A signal similar to that emitted by Global System for Mobile Communications (GSM) mobile phones was used at a specific absorption rate of 1 W/kg under strictly controlled conditions of temperature and dosimetry. To evaluate DNA damage after RF-radiation exposure alone, the alkaline comet assay and the cytokinesis-block micronucleus assay were used. In the combined-exposure experiments, MX was given at a concentration of 25 microM for 1 h immediately after the RF-radiation exposure, and the effects were evaluated by the alkaline comet assay. The results revealed no genotoxic and cytotoxic effects from RF radiation alone in either cell line. As expected, MX treatment induced an increase in DNA migration in the comet assay, but no enhancement of the MX-induced DNA damage was observed in the cells exposed to RF radiation.

In this study, the induction of apoptosis after exposure to 900 MHz radiofrequency radiation (GSM signal) was investigated by assessing caspase 3 activation in exponentially growing Jurkat cells and in quiescent and proliferating human peripheral blood lymphocytes (PBLs). The exposure was carried out at an average specific absorption rate of 1.35 W/kg in a dual wire patch cell exposure system where the temperature of cell cultures was accurately controlled. After 1 h exposure to the radiofrequency field, a slight but statistically significant increase in caspase 3 activity, measured 6 h after exposure, was observed in Jurkat cells (32.4%) and in proliferating human PBLs (22%). In contrast, no effect was detected in quiescent human PBLs. In the same experimental conditions, apoptosis was also evaluated in Jurkat cells by Western blot analysis and in both cell types by flow cytometry. To evaluate late effects due to caspase 3 activity, flow cytometry was also employed to assess apoptosis and viability 24 h after radiofrequency-radiation exposure in both cell types. Neither the former nor the latter was affected. Since in recent years it has been reported that caspases are also involved in processes other than apoptosis, additional cell cycle studies were carried out on proliferating T cells exposed to radiofrequency radiation; however, we found no differences between sham-exposed and exposed cultures. Further studies are warranted to investigate the biological significance of our findings of a dose-response increase in caspase 3 activity after exposure to radiofrequency radiation.

The last decades have seen increased interest toward possible adverse effects arising from exposure to intense static magnetic fields. This concern is mainly due to the wider and wider applications of such fields in industry and clinical practice; among them, Magnetic Resonance Imaging (MRI) facilities are the main sources of exposure to static magnetic fields for both general public (patients) and workers. In recent investigations, exposures to static magnetic fields have been demonstrated to elicit, in different cell models, both permanent and transient modifications in cellular endpoints critical for the carcinogenesis process. The World Health Organization has therefore recommended in vitro investigations as important research need, to be carried out under strictly controlled exposure conditions. Here we report on the absence of effects on cell viability, reactive oxygen species levels and DNA integrity in MRC-5 human foetal lung fibroblasts exposed to 370 mT magnetic induction level, under different exposure regimens. Exposures have been performed by using an experimental apparatus designed and realized for operating with the static magnetic field generated by permanent magnets and confined in a magnetic circuit, to allow cell cultures exposure in absence of confounding factors like heating or electric field components.