Influence of 864 MHz electromagnetic field on growth kinetics of established cell line

Considering often contradictory data on biological effects of mobile phones frequencies on established cell culture lines, our study aimed at evaluating the influence of 864 MHz electromagnetic field on proliferation, colony forming ability and viability of Chinese hamster lung cells continuous line V79. Prior to exposure for 1, 2 and 3 hours in transversal electromagnetic mode cell (TEM-cell) equipped by Philips PM 5508 signal generator cell samples were sub-cultivated for one day. Cell samples were exposed to 864 MHz continuous wave at an average specific absorption rate (SAR) of 0.08 W/kg. To determine cell growth, V79 cells were plated in concentration of 1 × 10⁴ cells per milliliter of nutrient medium RPMI 1640, and raised in a humified atmosphere at 37°C in 5% CO₂. Cell proliferation was determined by cell counts for each hour of exposure on post-exposure day 1, 2, 3, 4 and 5. To identify colony-forming ability, cells were cultivated in concentration of 40 cells/mL of RPMI 1640 and incubated according to the deliberated experimental protocol. Colony forming ability for each hour of exposure was defined by colony counts on experimental day 7. Trypan blue exclusion test was used to determine viability of cells. In comparison to sham-exposed cells, growth curve of irradiated cell samples showed significant decrease (p < 0.05) after 2 and 3 hours of exposure on experimental day 3, respectively. Both, the colony forming ability and viability of irradiated cells did not significantly differ from exposed “mock” condition. Under strictly controlled laboratory conditions, applied radiofrequency microwaves (RF/MW) irradiation significantly affected cell proliferation kinetics but not viability or ability of V79 cells to form colonies. Sophisticated mechanism of action is intending to be elucidated in the further research which will include insight into the RF/MW related event at the subcellular level.     

The effect of chronic exposure to 835.62 MHz FDMA or 847.74 MHz CDMA radiofrequency radiation on the incidence of spontaneous tumors in rats

This study was designed to determine whether chronic exposure to radiofrequency (RF) radiation from cellular phones increased the incidence of spontaneous tumors in F344 rats. Eighty male and 80 female rats were randomly placed in each of three irradiation groups. The sham group received no irradiation; the Frequency Division Multiple Access (FDMA) group was exposed to 835.62 MHz FDMA RF radiation; and the Code Division Multiple Access (CDMA) group was exposed to 847.74 MHz CDMA RF radiation. Rats were irradiated 4 h per day, 5 days per week over 2 years. The nominal time-averaged specific absorption rate (SAR) in the brain for the irradiated animals was 0.85 +/- 0.34 W/kg (mean +/- SD) per time-averaged watt of antenna power. Antennas were driven with a time-averaged power of 1.50 +/- 0.25 W (range). That is, the nominal time-averaged brain SAR was 1.3 +/- 0.5 W/kg (mean +/- SD). This number was an average from several measurement locations inside the brain, and it takes into account changes in animal weight and head position during irradiation. All major organs were evaluated grossly and histologically. The number of tumors, tumor types and incidence of hyperplasia for each organ were recorded. There were no significant differences among final body weights or survival days for either males or females in any group. No significant differences were found between treated and sham-exposed animals for any tumor in any organ. We conclude that chronic exposure to 835.62 MHz FDMA or 847.74 MHz CDMA RF radiation had no significant effect on the incidence of spontaneous tumors in F344 rats.     

Effect of 2,450 MHz microwave radiation on the development of the rat brain

Male Sprague-Dawley rats were exposed to 2,450 MHz microwave radiation at an incident power density of 10 mW/cm2 daily for 3 hours from day 4 of pregnancy (in utero exposure) through day 40 postpartum, except for 2 days at the perinatal period. The animals were killed, and the brains removed, weighed, measured, and histologically examined at 15, 20, 30, and 40 days of age. The histologic parameters examined included the cortical architecture of the cerebral cortex, the decline of the germinal layer along the lateral ventricles, the myelination of the corpus callosum, and the decline of the external germinal layer of the cerebellar cortex. In 40-day-old rats, quantitative measurements of neurons were also made. The spine density of the pyramidal cells in layer III of the somatosensory cortex, and the density of basal dendritic trees of the pyramidal cells in layer V were measured in Golgi-Cox impregnated specimens. In addition, the density of Purkinje cells and the extent of the Purkinje cell layer in each lobule were measured in midsagittal sections of the cerebellum stained with thionin. There were no remarkable differences between microwave-exposed and control (sham-irradiated) groups for any of the histologic or quantitative parameters examined; however, the findings provide important information on quantitative measurements of the brain. The data from this study failed to demonstrate that there is a significant effect on rat brain development due to microwave exposure (10 mW/cm2) during the embryonic, fetal, and postnatal periods.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of 2450 MHz microwave radiation on hematopoiesis of pregnant mice

In this study, the influence of 2450 MHz CW microwave radiation on hematopoiesis in pregnant mice was examined. Dams (mice CD-1 strain) were irradiated during Days 1-6 or 6-15 of pregnancy. The animals were irradiated for a total of 8 hr per day (two 4-hr exposures in 9 hr) at an average power density of 30 mW/cm2. Peripheral blood and bone marrow samples were obtained on Day 18 of pregnancy. The total leukocyte and differential leukocyte counts of peripheral blood samples were not affected by either exposure regimen. In addition, no effects were noted in either the erythroid or myeloid mitotic indices of bone marrow samples. Exposure of pregnant mice to microwave radiation under the conditions of these experiments had no effects on the investigated aspects of hematopoiesis.     

The effects of 860 MHz radiofrequency radiation on the induction or promotion of brain tumors and other neoplasms in rats

Sprague-Dawley rats were irradiated with a continuous- wave (CW) or a pulsed-wave (P) radiofrequency (RF) for 6 h/day, 5 days/week from 2 up to 24 months of age. The RFs emanated from dipole antennas (1 W average output) 2.0 +/- 0.5 cm from the tip of each rat's nose. The RFs had an 860 MHz frequency, and the specific absorption rate was 1.0 W/ kg averaged over the brain. Fifteen groups of 60 rats (900 total) were formed from offspring of females injected i.v. with 0 (groups 1, 2, 9, 10, 13), 2.5 (groups 5, 6, 7, 8, 11, 12, 14) or 10 mg/kg (groups 3, 4, 15) ethylnitrosourea (ENU) to induce brain tumors. Groups 1, 3, 5 and 7 received the PRF, and groups 9 and 11 the CWRF; groups 2, 4, 6, 8, 10 and 12 were sham-irradiated, and groups 13-15 were cage controls. All rats but 2, totaling 898, were necropsied, and major tissues were studied histopathologically. There was no statistically significant evidence that the PRF or CWRF induced neoplasia in any tissues. Additionally, there was no significant evidence of promotion of cranial or spinal nerve or spinal cord tumors. The PRF or CWRF had no statistically significant effect on the number, volume, location, multiplicity, histological type, malignancy or fatality of brain tumors. There was a trend for the group that received a high dose of ENU and was exposed to the PRF to develop fatal brain tumors at a higher rate than its sham group; however, the result was not significant using the log-rank test (P = 0.14, 2-tailed). No statistically significant differences were related to the PRF or CWRF compared to controls in the low- or zero-dose groups regarding tumors of any kind.     

The effect of 2450 MHz microwave radiation on the ultrastructure of snail neurons

An electron microscopical study of snail neurons was undertaken to verify whether any ultrastructural alterations accompany microwave-induced electrophysiological changes observed in these neurons. Subesophageal ganglia from Helix aspersa snails were exposed to 2450 MHz microwave radiation in vitro at SAR 12.9 mW/g for 60 minutes. It was found that exposure at 21 degrees C causes minor changes in Golgi complexes and slight swelling of the endoplasmic reticulum.     

Effects of 460 MHz microwave radiation on Drosophila embryos under raised temperature

Combined effect of 460-MHz microwave irradiation and increased (up to 40 degrees C) temperature on Drosophila embryos of definite age was studied. It was demonstrated that the effect of 5-min exposures to non-modulated microwaves with 6 W/kg SAR accompanied with heating is only a little stronger than at normal temperature (24.5 degrees C). Irradiation with pulse-modulated microwaves with pulse repetition rates of 6, 10, 16, and 22 p.p.s. with average SAR of 0.12 W/kg (pulsed SAR 3 W/kg) combined with increased temperature caused some changes in PID dependent on the pulse rate. At 6 and 22 p.p.s, the increase in PID was close to that observed at normal temperature while at 10 and 16 p.p.s. the microwave irradiation did not produce any noticeable effect on development of the Drosophilas.     

Effect of 900 MHz radiofrequency radiation on oxidative stress in rat brain and serum

The increasing use of mobile telephones raises the question of possible adverse effects of the electromagnetic fields (EMF) that these phones produce. In this study, we examined the oxidative stress in the brain tissue and serum of rats that resulted from exposure to a 900-MHz EMF at a whole body average specific absorption rate (SAR) of 1.08 W/kg for 1 h/day for 3 weeks. We also examined the antioxidant effect of garlic powder (500 mg/kg/day) given orally to EMF-exposed rats. We found that malondialdehyde (MDA) (p < 0.001) and advanced oxidation protein product (AOPP) (p < 0.05) increased in rat brain tissue exposed to the EMF and that garlic reduced these effects (p < 0.05). There was no significant difference in the nitric oxide (NO) levels in the brain. Paraoxonase (PON) was not detected in the brain. There was a significant increase in the levels of NO (p < 0.001) detected in the serum after EMF exposure, and garlic intake did not affect this increase in NO. Our results suggest that there is a significant increase in brain lipid and protein oxidation after electromagnetic radiation (EMR) exposure and that garlic has a protective effect against this oxidative stress.     

Effect of ploidy on the response of V79 cells to ionizing radiation

The responses of diploid, tetraploid and near-hexaploid V79 cells to X-irradiation or DNA-associated 125I-decay were compared. When cell killing, following X-irradiation, was plotted against the induced level of DNA double-strand breakage (dsb) per unit length of DNA, there was no significant difference between the relationships for each cell line. This suggested that the number of X-ray-induced DNA dsb per cell required to produce a lethal lesion was proportional to ploidy. Consistent with the X-ray results, tetraploid cells required 121 +/- 4 and diploid cells 60 +/- 1 125I-decays to produce a lethal lesion. However, the hexaploid cells deviated from this relationship and required 137 +/- 5 decays. The relationship between relative elution and 125I decays/cell reflected cellular DNA content. It is concluded that current models of radiation action are unable to explain these findings satisfactorily.     

The effects of pulsed 860 MHz radiofrequency radiation on the promotion of neurogenic tumors in rats

In a previous study, this laboratory reported a statistically nonsignificant trend for shortened latency of ethylnitrosourea (ENU)-induced brain tumors in Sprague-Dawley rats exposed to an 860 MHz pulsed radiofrequency (RF) signal. The present study was designed to investigate further any promoting effect of the pulsed RF signal on latency and other characteristics of neurogenic tumors in the progeny of pregnant rats treated with 6.25 or 10 mg/kg ENU. The resulting 1080 offspring were randomized equally by number, sex and ENU dose into pulsed RF, sham and cage control groups. The rats were exposed to the pulsed RF signal 6 h per day 5 days per week; the sham-exposed group was similarly confined for the same periods, and the cage controls were housed in standard cages. An essentially equal number of rats from each group were killed humanely every 30 days between the ages of 171 and 325 days; 32 rats died and 225 rats were killed when they were moribund. Postmortem examinations on the 1080 rats revealed 38 spinal cord tumors, 191 spinal nerve tumors, 232 cranial nerve tumors, and 823 brain tumors. A methodical study of the tumor characteristics disclosed no evidence that exposure to the pulsed RF signal affected the incidence, malignancy, volume, multiplicity, latency or fatality associated with any kind of neurogenic tumor.     

Mutagenicity of N-nitrosodiethanolamine in a V79-derived cell line expressing two human biotransformation enzymes

N-Nitrosodiethanolamine (NDELA) has demonstrated carcinogenic activity in various rodent models. However, it is negative or only weakly active in standard in vitro genotoxicity assays. This poor response might be due to the requirement of specific enzymes for its activation. Previous work indicated that cytochrome P450 (CYP) 2E1, alcohol dehydrogenases and sulphotransferases (SULTs) can convert NDELA into reactive metabolites. We report here that NDELA induces concentration-dependent gene mutations (at the hprt locus) in V79-hCYP2E1-hSULT1A1 cells, engineered for expression of human CYP2E1 and human SULT1A1, but is inactive in parental V79 cells. Mutagenicity of NDELA in V79-hCYP2E1-hSULT1A1 cells was abolished by the CYP2E1 inhibitor 1-aminobenzotriazole, but unaffected by the SULT1A1 inhibitor pentachlorophenol. The efficiency and specificity of these inhibitors was demonstrated in gene mutation assays using SULT- and CYP2E1-dependent reference mutagens, 2-nitropropane and N-nitrosodimethylamine, respectively. In this study, it is documented for the first time that NDELA can induce gene mutations in mammalian cells. Whereas human CYP2E1 was required for its activation, human SULT1A1 was not involved either in its activation or its inactivation in our cell model.     

Effects of 2450 MHz microwave radiation during the gestational period on the postnatal hematology of rats

Pregnant Sprague-Dawley rats were exposed to 2.45 GHz CW radiation for 3 h daily from day 4 through day 20 of pregnancy at an average power density of 10.3 mW/cm2. At 2, 10, 20, and 30 d postpartum, several aspects of peripheral blood hematology of male and female pups were examined. No effects were noted on pregnancy rate or litter size; however, pup weight was reduced. Also, the white blood cell numbers were lower in male and female pups exposed to microwave radiation and the differential cell counts were altered in the female pups at 10 d postpartum. At 30 d of age, no differences were noted between the sham and exposed pups for any of the hematologic variables examined.     

Effect of amplitude-modulated 147 MHz radiofrequency radiation on calcium ion efflux from avian brain tissue

Cerebral cortex tissue slices and cerebral hemispheres prepared from Gallus domesticus chicks were exposed to 147 MHz radiofrequency radiation, amplitude modulated at 16 Hz and applied at a power density of 0.75 mW/cm2, to determine the effect of such exposure of 45Ca2+ efflux from the avian brain tissue. Statistical analysis of these data demonstrates that such exposure has no significant effect on 45Ca2+ efflux.     

The effect of microwave radiation on the cell genome

Cultured V79 Chinese hamster cells were exposed to continuous radiation, frequency 7.7 GHz, power density 30 mW/cm2 for 15, 30, and 60 min. The parameters investigated were the incorporation of [3H]thymidine and the frequency of chromosome aberrations. Data obtained by 2 methods (the incorporation of [3H]thymidine into DNA and autoradiography) showed that the inhibition of [3H]thymidine incorporation took place by complete prevention of DNA from entering into the S phase. The normal rate of incorporation of [3H]thymidine was recovered within 1 generation cycle of V79 cells. Mutagenic tests performed concurrently showed that even DNA macromolecules were involved in the process. In comparison with the control samples there was a higher frequency of specific chromosome lesions in cells that had been irradiated. Results discussed in this study suggest that microwave radiation causes changes in the synthesis as well as in the structure of DNA molecules.     

The effect of 835.62 MHz FDMA or 847.74 MHz CDMA modulated radiofrequency radiation on the induction of micronuclei in C3H 10T(1/2) cells

To determine if radiofrequency (RF) radiation induces the formation of micronuclei, C3H 10T(1/2) cells were exposed to 835.62 MHz frequency division multiple access (FDMA) or 847.74 MHz code division multiple access (CDMA) modulated RF radiation. After the exposure to RF radiation, the micronucleus assay was performed by the cytokinesis block method using cytochalasin B treatment. The micronuclei appearing after mitosis were scored in binucleated cells using acridine orange staining. The frequency of micronuclei was scored both as the percentage of binucleated cells with micronuclei and as the number of micronuclei per 100 binucleated cells. Treatment of cells with cytochalasin B at a concentration of 2 microg/ml for 22 h was found to yield the maximum number of binucleated cells in C3H 10T(1/2) cells. The method used for the micronucleus assay in the present study detected a highly significant dose response for both indices of micronucleus production in the dose range of 0.1-1.2 Gy and it was sensitive enough to detect a significant (P > 0.05) increase in micronuclei after doses of 0.3 Gy in exponentially growing cells and after 0.9 Gy in plateau-phase cells. Exponentially growing cells or plateau-phase cells were exposed to CDMA (3.2 or 4.8 W/kg) or FDMA (3.2 or 5.1 W/kg) RF radiation for 3, 8, 16 or 24 h. In three repeat experiments, no exposure condition was found by analysis of variance to result in a significant increase relative to sham-exposed cells either in the percentage of binucleated cells with micronuclei or in the number of micronuclei per 100 binucleated cells. In this study, data from cells exposed to different RF signals at two SARs were compared to a common sham-exposed sample. We used the Dunnett's test, which is specifically designed for this purpose, and found no significant exposure-related differences for either plateau-phase cells or exponentially growing cells. Thus the results of this study are not consistent with the possibility that these RF radiations induce micronuclei.     

Flow cytometric determination of cell cycle parameters of V79 cells by continuous labeling with bromodeoxyuridine

A simple method with which to determine the cell cycle parameters, TG1, TS and TG2M (the durations of the G1, S and G2 + M phases) is described. V79 Chinese hamster lung cells were used to evaluate the method. After continuous labeling with bromodeoxyuridine (BrdU), V79 cells were stained with anti BrdU-monoclonal antibody with FITC (fluorescein isothiocyanate) and with PI (propidium iodide). The individual cells were checked by flow cytometry for green and red fluorescences whose signal intensities corresponded to the BrdU and cellular DNA contents. The durations of G1, S and G2 + M phases of V79 cells were determined by measuring the cell fractions containing the nonlabeled G1, labeled S and nonlabeled G2 + M phases. The reliability of this method is discussed.     

RNA dependence in the cell cycle of V79 cells

The cell cycle of V79 Chinese hamster lung cells synchronized by hydroxyurea was investigated by flow cytometry. The metachromatic fluorochrome acridine orange was used to differentially stain DNA and RNA of V79 cells. Green and red fluorescence from individual cells, representing cellular DNA and RNA, respectively, was measured by flow cytometry. Periodic changes of cellular DNA and RNA contents were observed over nine cell cycles. The duration of G1, S, and G2 + M phases of synchronized V79 cells whose RNA content was close to that of the cells in balanced growth was 3, 4.5, and 1.5 hours, respectively. The duration of G1 and S phases of cells containing RNA above a certain threshold was inversely proportional to the RNA content. The RNA content of cells containing RNA above the normal level regressed to normal after a few generations. Coefficients of variation for RNA content were significantly larger than those for DNA. An explanation for the decay of synchrony in a synchronized cell population is proposed.     

The effect of 2450 MHz microwave radiation on histamine secretion by rat peritoneal mast cells

Isolated rat peritoneal mast cells actively secrete histamine in response to reaginic or chemical stimulation. Mast cells were irradiated in a waveguide microwave exposure chamber at 2450 MHz with power absorptions of 8.2 and 41.0 mW/g for periods up to 3 h. These levels of microwave absorption caused no change in the morphological characteristics or viability of the cells. Irradiated mast cells were stimulated with compound 48/80, a potent, noncytotoxic histamine releasing agent. The dose response curves showed that neither prior nor simultaneous irradiation of mast cells at 37°C affected 48/80-induced secretion. However, microwave power absorptions of 41.0 mW/g inhibited secretion at 44.0°C. Precise measurements of the effect of heat on secretion indicated that this level of inhibition could have been produced by a radiation induced increase in cell temperature between 0.4 and 0.9°C above ambient levels. Alternatively, the heat stress produced at 44°C may have sensitized the cells to the electromagnetic effects of the microwave radiation. Rat peritoneal mast cells can therefore be useful as a model for the study of functioning secretory cells during microwave irradiation and can also be used to monitor the synergistic effects of cell heating during in vitro exposure.