ID Number		537
Study Type		In Vitro
Model		Microwave exposure to cell lines and yeast and analysis of growth, apoptosis, oxidative stress, genotoxicity and cancer.
Details		Human SH-SY5Y neuroblastoma and mouse L929 fibroblast cells were exposed to 872 MHz (CW or GSM) either alone or in combination with chemicals that induce reactive oxygen species (menadione) or lipid peroxidation (tert-butylhydroperoxide or t-BOOH). Preliminary experiments determined that SH-SY5Y neuroblastoma cells were much more sensitive to chemically induced oxidation than the mouse L929 fibroblast cells and thus concentrations were adjusted accordingly to achieve a similar baseline level of oxidative stress. RF exposure was performed at 5 W/kg for either 1 or 24 hours using an enclosed resonator system described in prior studies (Hoyto et al 2006, Radiat Environ. Biophys. 45:231-235). Temperature control was performed by water circulation underneath the cell cultures (convective cooling). Following RF exposure, indicators of oxidative stress (total glutathione, lipid peroxidation), proliferation, apoptosis (caspase 3 activity, DNA fragmentation), and cell viability were measured. The authors report no effect due to RF exposure alone with either CW or GSM signal modulation, and no effects due to CW RF exposure in combination with either oxidizing agent. However, GSM exposure in combination with t-BOOH increased lipid peroxidation in SH-SY5Y cells (p = 0.002), but not L929 cells. GSM in combination with menadione increased caspase 3 activity (caspase dependent apoptotic marker) in L929 cells (p = 0.008), but not SH-SY5Y cells. In the case of RF + menadione in L929 cells, CW exposure also resulted in a slight (although non-significant) increase in caspase-3 activity. No effect of RF (CW or GSM) exposure, either alone or in combination with oxidizing agents, was observed on cell proliferation or viability. In earlier studies with yeast (SEy2101a wild type, KFy437 apoptosis prone, and KFy417 control), the authors reported exposure to 900 MHz (CW or GSM) alone for 1 hour at up to 3 W/kg did not affect apoptosis, colony formation, or gene expression, but in combination with UV-B (another oxidizing agent), GSM-specific exposure (not CW) increased apoptosis in the mutant KFy437 strain and decreased colony formation in both mutant and wild type strains as compared to UV-only controls (Markkanen et al, Bioelectromagnetics (2004) 25:127-133). In an earlier study, SEy2101a (wild type), KFy437 (apoptosis prone), and KFy417 (control) yeast were exposed to 900 MHz (CW or GSM) RF at 0.5 W/kg or 872 MHz (CW or GSM) at 3.0 W/kg for 1 hour at constant temperature, either alone or in combination with UVB (250 J/m2), and analyzed for apoptosis (via annexin V-FITC flow cytometry analyzing loss of membrane asymmetry), colony forming ability, and gene expression. RF alone (CW and GSM) did not affect any markers of apoptosis in either mutant or wild type strains. GSM-specific RF exposure at both 0.5 and 3.0 W/kg + UV increased apoptosis in mutant (but not wild type) yeast and decreased colony formation in both mutant and wild type yeast compared to UV-only controls. The authors conclude the lack of effects of unmodulated RF at identical SAR levels offers a "persuasive argument" that the observed effects are "nonthermal". In a subsequent study, SH-SY5Y cells were exposed as above (872 MHz CW, GSM at 5 W/kg for 1 hr followed by treatment subsequent treatment with menadione. The authors confirm that CW 1hr exposure plus menadione increased reactive oxygen species over menadione treatment alone. GSM plus menadione also increased reactive oxygen species, but not as effectively and with a delayed onset. No genotoxic effect was observed under any of the experimental conditions. AUTHORS' ABSTRACT: Herrala et al. 2018 (IEEE #6971): PURPOSE: We examined genotoxicity, co-genotoxicity and induced genomic instability (IGI) in primary astrocytes exposed to radiofrequency (RF) radiation. MATERIALS AND METHODS: Rat primary astrocytes were exposed to 872 MHz GSM-modulated or continuous wave (CW) RF radiation at specific absorption rates of 0.6 or 6.0 W/kg for 24 h. Menadione (MQ) and methyl methanesulfonate (MMS; only in genotoxicity experiments) were used as co-exposures. Alkaline Comet assay and flow cytometric micronucleus scoring were used to detect genetic damage. RESULTS: No IGI was observed from RF radiation alone or combined treatment with MQ. RF radiation alone was not genotoxic. RF radiation combined with chemical exposure showed some statistically significant differences: increased DNA damage at 6.0 W/kg but decreased DNA damage at 0.6 W/kg in cells exposed to GSM-modulated RF radiation and MQ, and increased micronucleus frequency in cells exposed to CW RF radiation at 0.6 W/kg and MMS. CONCLUSIONS: Exposure to GSM modulated RF radiation at levels up to 6.0 W/kg did not induce or enhance genomic instability in rat primary astrocytes. Lack of genotoxicity from RF radiation alone was convincingly shown in multiple experiments. Co-genotoxicity of RF radiation and genotoxic chemicals was not consistently supported by the results. AUTHORS' ABSTRACT: Naarala et al. 2019 (IEEE #7273): This review discusses the use of systems biology in understanding the biological eects of electromagnetic fields, with particular focus on induction of genomic instability and cancer. We introduce basic concepts of the dynamical systems theory such as the state space and attractors and the use of these concepts in understanding the behavior of complex biological systems. We then discuss genomic instability in the framework of the dynamical systems theory, and describe the hypothesis that environmentally induced genomic instability corresponds to abnormal attractor states; large enough environmental perturbations can force the biological system to leave normal evolutionarily optimized attractors (corresponding to normal cell phenotypes) and migrate to less stable variant attractors. We discuss experimental approaches that can be coupled with theoretical systems biology such as testable predictions, derived from the theory and experimental methods, that can be used for measuring the state of the complex biological system. We also review potentially informative studies and make recommendations for further studies.
Findings		Effects
Status		Completed With Publication
Principal Investigator		University of Kuopio, Finland - juutilainen@uku.fi
Funding Agency		Nat'l Res Prog, Finland, MMF, GSM Association, TEKES, HERMO
Country		FINLAND
References		Luukkonen, J et al. Mut Res., (2009) 662:54-58 Hoyto, A et al. Radiation Research, (2008) 170:235-243 Markkanen , A et al. Bioelectromagnetics, (2004) 25:127-133 Herrala, M et al. Int J Radiat Biol., (2018) 94:883-889 Naarala, J et al. J. Genes., (2019) 10:479- Luukkonen, J et al. Bioelectromagnetics, (2010) 31:417-424
Comments		Such a modulation-specific effect, if real, might imply that the effect is non-thermal in nature. However, the fact that the observations were not consistent between cell lines and that related endpoints were not affected in a concomitant way adds some question to the findings. In the human cell line study, total glutathione was not measured in GSM + t-BOOH treated SH-SY5Y cells to corroborate the lipid peroxidation observation, and DNA fragmentation was not affected in GSM + menadion treated L929 cells to corroborate the increase in apoptosis that might follow from increased caspase-3 activity. The effect of non-RF temperature elevation on the oxidative endpoints in these two cell lines was not determined. In the yeast study, no dose response was reported, which should have existed with 0.5 and 3.0 W/kg exposures.