DNA damage and micronucleus induction in human leukocytes after acute in vitro exposure to a 1.9 GHz continuous-wave radiofrequency field

Human blood cultures were exposed to a 1.9 GHz continuous-wave (CW) radiofrequency (RF) field for 2 h using a series of six circularly polarized, cylindrical waveguides. Mean specific absorption rates (SARs) of 0.0, 0.1, 0.26, 0.92, 2.4 and 10 W/kg were achieved, and the temperature within the cultures during a 2-h exposure was maintained at 37.0 +/- 0.5 degrees C. Concurrent negative (incubator) and positive (1.5 Gy (137)Cs gamma radiation) control cultures were run for each experiment. DNA damage was quantified immediately after RF-field exposure using the alkaline comet assay, and four parameters (tail ratio, tail moment, comet length and tail length) were used to assess DNA damage for each comet. No evidence of increased primary DNA damage was detected by any parameter for RF-field-exposed cultures at any SAR tested. The formation of micronuclei in the RF-field-exposed blood cell cultures was assessed using the cytokinesis-block micronucleus assay. There was no significant difference in the binucleated cell frequency, incidence of micronucleated binucleated cells, or total incidence of micronuclei between any of the RF-field-exposed cultures and the sham-exposed controls at any SAR tested. These results do not support the hypothesis that acute, nonthermalizing 1.9 GHz CW RF-field exposure causes DNA damage in cultured human leukocytes.     

Microarray gene expression profiling of a human glioblastoma cell line exposed in vitro to a 1.9 GHz pulse-modulated radiofrequency field

The widespread use of mobile phones has led to public concerns about the health effects associated with exposure to radiofrequency (RF) fields. The paramount concern of most persons relates to the potential of these fields to cause cancer. Unlike ionizing radiation, RF fields used for mobile telecommunications (800-1900 MHz) do not possess sufficient energy to directly damage DNA. Most rodent bioassay and in vitro genotoxicity/mutation studies have reported that RF fields at non-thermal levels have no direct mutagenic, genotoxic or carcinogenic effects. However, some evidence has suggested that RF fields may cause detectable postexposure changes in gene expression. Therefore, the purpose of this study was to assess the ability of exposure to a 1.9 GHz pulse-modulated RF field for 4 h at specific absorption rates (SARs) of 0.1, 1.0 and 10.0 W/kg to affect global gene expression in U87MG glioblastoma cells. We found no evidence that non-thermal RF fields can affect gene expression in cultured U87MG cells relative to the nonirradiated control groups, whereas exposure to heat shock at 43 degrees C for 1 h up-regulated a number of typical stress-responsive genes in the positive control group. Future studies will assess the effect of RF fields on other cell lines and on gene expression in the mouse brain after in vivo exposure.     

Analysis of gene expression in two human-derived cell lines exposed in vitro to a 1.9 GHz pulse-modulated radiofrequency field

There is considerable controversy surrounding the biological effects of radiofrequency (RF) fields, as emitted by mobile phones. Previous work from our laboratory has shown no effect related to the exposure of 1.9 GHz pulse-modulated RF fields on the expression of 22,000 genes in a human glioblastoma-derived cell-line (U87MG) at 6 h following a 4 h RF field exposure period. As a follow-up to this study, we have now examined the effect of RF field exposure on the possible expression of late onset genes in U87MG cells after a 24 h RF exposure period. In addition, a human monocyte-derived cell-line (Mono-Mac-6, MM6) was exposed to intermittent (5 min ON, 10 min OFF) RF fields for 6 h and then gene expression was assessed immediately after exposure and at 18 h postexposure. Both cell lines were exposed to 1.9 GHz pulse-modulated RF fields for 6 or 24 h at specific absorption rates (SARs) of 0.1-10.0 W/kg. In support of our previous results, we found no evidence that nonthermal RF field exposure could alter gene expression in either cultured U87MG or MM6 cells, relative to nonirradiated control groups. However, exposure of both cell-lines to heat-shock conditions (43 degrees C for 1 h) caused an alteration in the expression of a number of well-characterized heat-shock proteins.     

No evidence for genotoxic effects from 24 h exposure of human leukocytes to 1.9 GHz radiofrequency fields

The current study extends our previous investigations of 2-h radiofrequency (RF)-field exposures on genotoxicity in human blood cell cultures by examining the effect of 24-h continuous-wave (CW) and pulsed-wave (PW) 1.9 GHz RF-field exposures on both primary DNA damage and micronucleus induction in human leukocyte cultures. Mean specific absorption rates (SARs) ranged from 0 to 10 W/kg, and the temperature within the cultures was maintained at 37.0 +/- 1.0 degrees C for the duration of the 24-h exposure period. No significant differences in primary DNA damage were observed between the sham-treated controls and any of the CW or PW 1.9 GHz RF-field-exposed cultures when processed immediately after the exposure period by the alkaline comet assay. Similarly, no significant differences were observed in the incidence of micronuclei, incidence of micronucleated binucleated cells, frequency of binucleated cells, or proliferation index between the sham-treated controls and any of the CW or PW 1.9 GHz RF-field-exposed cultures. In conclusion, the current study found no evidence of 1.9 GHz RF-field-induced genotoxicity in human blood cell cultures after a 24-h exposure period.     

Mitogen responsiveness after exposure of influenza virus-infected human mononuclear leukocytes to continuous or pulse-modulated radiofrequency radiation

Data are available regarding interactions of radiofrequency radiation (RFR) with normal human mononuclear leukocytes. However, no data have emerged regarding effects of RFR on human leukocytes already challenged by a commonly encountered alternate agent, such as a virus. Therefore, in these studies, uninfected (control) and in vitro influenza virus-infected human mononuclear leukocytes were exposed to 2450 MHz RFR as continuous waves or pulse-modulated at 60 or 16 Hz, at a specific absorption rate of 4 mW/ml. Such exposures produced no significant effects on leukocyte viability or on mitogen-stimulated DNA synthesis by either uninfected or influenza virus-infected leukocytes when compared to sham-RFR-exposed of cells.     

Evaluation of genotoxic effects in human leukocytes after in vitro exposure to 1950 MHz UMTS radiofrequency field

In the present study the third generation wireless technology of the Universal Mobile Telecommunication System (UMTS) signal was investigated for the induction of genotoxic effects in human leukocytes. Peripheral blood from six healthy donors was used and, for each donor, intermittent exposures (6 min RF on, 2 h RF off) at the frequency of 1950 MHz were conducted at a specific absorption rate of 2.2 W/kg. The exposures were performed in a transverse electro magnetic (TEM) cell hosted in an incubator under strictly controlled conditions of temperature and dosimetry. Following long duration intermittent RF exposures (from 24 to 68 h) in different stages of the cell cycle, micronucleus formation was evaluated by applying the cytokinesis block micronucleus assay, which also provides information on cell division kinetics. Primary DNA damage (strand breaks/alkali labile sites) was also investigated following 24 h of intermittent RF exposures, by applying the alkaline single cell gel electrophoresis (SCG)/comet assay. Positive controls were included by treating cell cultures with Mitomycin-C and methylmethanesulfonate for micronucleus and comet assays, respectively. The results obtained indicate that intermittent exposures of human lymphocytes in different stages of cell cycle do not induce either an increase in micronucleated cells, or change in cell cycle kinetics; moreover, 24 h intermittent exposures also fail to affect DNA structure of human leukocytes soon after the exposures, likely indicating that repairable DNA damage was not induced.     

Evaluating the biological effects of intermittent 1.9 GHz pulse-modulated radiofrequency fields in a series of human-derived cell lines

Several recent studies have suggested that radiofrequency (RF) fields may cause changes in a variety of cellular functions that may eventually lead to potential long-term health effects. In the present study, we have assessed the ability of non-thermal RF-field exposure to affect a variety of biological processes (including apoptosis, cell cycle progression, viability and cytokine production) in a series of human-derived cell lines (TK6, HL60 and Mono-Mac-6). Exponentially growing cells were exposed to intermittent (5 min on, 10 min off) 1.9 GHz pulse-modulated RF fields for 6 h at mean specific absorption rates (SARs) of 0, 1 and 10 W/kg. Concurrent negative (incubator) and positive (heat shock for 1 h at 43 degrees C) controls were included in each experiment. Immediately after the 6-h exposure period and 18 h after exposure, cell pellets were collected and analyzed for cell viability, the incidence of apoptosis, and alterations in cell cycle kinetics. The cell culture supernatants were assessed for the presence of a series of human inflammatory cytokines (TNFA, IL1B, IL6, IL8, IL10, IL12) using a cytometric bead array assay. No detectable changes in cell viability, cell cycle kinetics, incidence of apoptosis, or cytokine expression were observed in any of RF-field-exposed groups in any of the cell lines tested, relative to the sham controls. However, the positive (heat-shock) control samples displayed a significant decrease in cell viability, increase in apoptosis, and alteration in cell cycle kinetics (G(2)/M block). Overall, we found no evidence that non-thermal RF-field exposure could elicit any detectable biological effect in three human-derived cell lines.     

DNA damage in frog erythrocytes after in vitro exposure to a high peak-power pulsed electromagnetic field

Till the present time, the genotoxic effects of high peak-power pulsed electromagnetic fields (HPPP EMF) on cultured cells have not been studied. We investigated possible genotoxic effects of HPPP EMF (8.8 GHz, 180 ns pulse width, peak power 65 kW, repetition rate 50 Hz) on erythrocytes of the frog Xenopus laevis. We used the alkaline comet assay, which is a highly sensitive method to assess DNA single-strand breaks and alkali-labile lesions. Blood samples were exposed to HPPP EMF for 40 min in rectangular wave guide. The specific absorption rate (SAR) calculated from temperature kinetics was about 1.6 kW/kg (peak SAR was about 300 MW/kg). The temperature rise in the blood samples at steady state was 3.5 +/- 0.1 degrees C. The data show that the increase in DNA damage after exposure of erythrocytes to HPPP EMF was induced by the rise in temperature in the exposed cell suspension. This was confirmed in experiments in which cells were incubated for 40 min under the corresponding temperature conditions. The results allow us to conclude that HPPP EMF-exposure at the given modality did not cause any a-thermal genotoxic effect on frog erythrocytes in vitro.     

Lack of direct DNA damage in human blood leukocytes and lymphocytes after in vitro exposure to high power microwave pulses

Currently, the potential genotoxicity of high power microwave pulses (HPMP) is not clear. Using the alkaline single cell gel electrophoresis assay, also known as the alkaline comet assay, we studied the effects of HPMP (8.8 GHz, 180 ns pulse width, peak power 65 kW, pulse repetition frequency 50 Hz) on DNA of human whole-blood leukocytes and isolated lymphocytes. The cell suspensions were exposed to HPMP for 40 min in a rectangular waveguide. The average SAR calculated from the temperature kinetics was about 1.6 kW/kg (peak SAR was about 300 MW/kg). The steady-state temperature rise in the 50 microl samples exposed to HPMP was 3.5 +/- 0.1 degrees C. In independent experiments, we did not find any statistically significant DNA damage manifested immediately after in vitro HPMP exposure of human blood leukocytes or lymphocytes or after HPMP exposure of leukocytes subsequently incubated at 37 degrees C for 30 min. Our results indicate that HPMP under the given exposure conditions did not induce DNA strand breaks, alkali-labile sites, and incomplete excision repair sites, which could be detected by the alkaline comet assay.     

Measurement of DNA damage and apoptosis in Molt-4 cells after in vitro exposure to radiofrequency radiation

To determine whether exposure to radiofrequency (RF) radiation can induce DNA damage or apoptosis, Molt-4 T lymphoblastoid cells were exposed with RF fields at frequencies and modulations of the type used by wireless communication devices. Four types of frequency/modulation forms were studied: 847.74 MHz code-division multiple-access (CDMA), 835.62 MHz frequency-division multiple-access (FDMA), 813.56 MHz iDEN(R) (iDEN), and 836.55 MHz time-division multiple-access (TDMA). Exponentially growing cells were exposed to RF radiation for periods up to 24 h using a radial transmission line (RTL) exposure system. The specific absorption rates used were 3.2 W/kg for CDMA and FDMA, 2.4 or 24 mW/kg for iDEN, and 2.6 or 26 mW/kg for TDMA. The temperature in the RTLs was maintained at 37 degrees C +/- 0.3 degrees C. DNA damage was measured using the single-cell gel electrophoresis assay. The annexin V affinity assay was used to detect apoptosis. No statistically significant difference in the level of DNA damage or apoptosis was observed between sham-treated cells and cells exposed to RF radiation for any frequency, modulation or exposure time. Our results show that exposure of Molt-4 cells to CDMA, FDMA, iDEN or TDMA modulated RF radiation does not induce alterations in level of DNA damage or induce apoptosis.     

Evaluation of genotoxic effects in human peripheral blood leukocytes following an acute in vitro exposure to 900 MHz radiofrequency fields

Human peripheral blood leukocytes from healthy volunteers have been employed to investigate the induction of genotoxic effects following 2 h exposure to 900 MHz radiofrequency radiation. The GSM signal has been studied at specific absorption rates (SAR) of 0.3 and 1 W/kg. The exposures were carried out in a waveguide system under strictly controlled conditions of both dosimetry and temperature. The same temperature conditions (37.0 +/- 0.1 degrees C) were realized in a second waveguide, employed to perform sham exposures. The induction of DNA damage was evaluated in leukocytes by applying the alkaline single cell gel electrophoresis (SCGE)/comet assay, while structural chromosome aberrations and sister chromatid exchanges were evaluated in lymphocytes stimulated with phytohemagglutinin. Alterations in kinetics of cell proliferation were determined by calculating the mitotic index. Positive controls were also provided by using methyl methanesulfonate (MMS) for comet assay and mitomycin-C (MMC), for chromosome aberration, or sister chromatid exchange tests. No statistically significant differences were detected in exposed samples in comparison with sham exposed ones for all the parameters investigated. On the contrary, the positive controls gave a statistically significant increase in DNA damage in all cases, as expected. Thus the results obtained in our experimental conditions do not support the hypothesis that 900 MHz radiofrequency field exposure induces DNA damage in human peripheral blood leukocytes in this range of SAR.     

DNA damage in human leukocytes after ischemia/reperfusion injury

Leukocytes have been shown to play an important role in the development of tissue injury after ischemia and reperfusion (I/R). In the present study, the effects of tourniquet-ischemia on induction of DNA damage in peripheral leukocytes and on respiratory burst of neutrophils in humans were examined. The DNA damage was measured as increased migration of DNA using the single-cell gel-electrophoresis technique (comet assay). Intracellular production of reactive oxygen species by neutrophils was measured flow-cytometrically using dihydrorhodamine 123 as indicator. Postischemic, significantly increased migration of DNA was found in leukocytes of 20 patients (tourniquet-ischemia of the lower limb for 65-130 min, anterior-cruciate-ligament-reconstruction) and in 10 experiments (1 volunteer, repeated tourniquet-ischemia of the upper limb for 60 min, no operation). DNA effects were most pronounced 5-30 min after tourniquet release, and then declined over a 2 h period, but did not return to preischemic baseline values. A similar time course showed the oxidative status of unstimulated granulocytes during reperfusion. Simultaneously, opposing changes were measured in formyl peptide (f-MLP)- or phorbol ester (PMA)-stimulated granulocytes, which showed a significantly declined respiratory burst reaction after tourniquet-release indicating preactivation of neutrophils by IR. Our data suggest that IR induces genotoxic effects in human leukocytes presumably in response to oxidative stress during reperfusion.     

Effects of 1.8 GHz radiofrequency field on DNA damage and expression of heat shock protein 70 in human lens epithelial cells

To investigate the DNA damage, expression of heat shock protein 70 (Hsp70) and cell proliferation of human lens epithelial cells (hLEC) after exposure to the 1.8 GHz radiofrequency field (RF) of a global system for mobile communications (GSM). An Xc-1800 RF exposure system was used to employ a GSM signal at 1.8 GHz (217 Hz amplitude-modulated) with the output power in the specific absorption rate (SAR) of 1, 2 and 3 W/kg. After 2 h exposure to RF, the DNA damage of hLEC was accessed by comet assay at five different incubation times: 0, 30, 60, 120 and 240 min, respectively. Western blot and RT-PCR were used to determine the expression of Hsp70 in hLECs after RF exposure. The proliferation rate of cells was evaluated by bromodeoxyuridine incorporation on days 0, 1 and 4 after exposure. The results show that the difference of DNA-breaks between the exposed and sham-exposed (control) groups induced by 1 and 2 W/kg irradiation were not significant at any incubation time point (P > 0.05). The DNA damage caused by 3 W/kg irradiation was significantly increased at the times of 0 and 30 min after exposure (P < 0.05), a phenomenon that could not be seen at the time points of 60, 120 or 240 min (P > 0.05). Detectable mRNA as well as protein expression of Hsp70 was found in all groups. Exposure at SARs of 2 and 3 W/kg for 2 h exhibited significantly increased Hsp70 protein expression (P < 0.05), while no change in Hsp70 mRNA expression could be found in any of the groups (P > 0.05). No difference of the cell proliferation rate between the sham-exposed and exposed cells was found at any exposure dose tested (P > 0.05). The results indicate that exposure to non-thermal dosages of RF for wireless communications can induce no or repairable DNA damage and the increased Hsp70 protein expression in hLECs occurred without change in the cell proliferation rate. The non-thermal stress response of Hsp70 protein increase to RF exposure might be involved in protecting hLEC from DNA damage and maintaining the cellular capacity for proliferation.     

Primary DNA damage in human blood lymphocytes exposed in vitro to 2450 MHz radiofrequency radiation

Human peripheral blood samples collected from three healthy human volunteers were exposed in vitro to pulsed-wave 2450 MHz radiofrequency (RF) radiation for 2 h. The RF radiation was generated with a net forward power of 21 W and transmitted from a standard gain rectangular antenna horn in a vertically downward direction. The average power density at the position of the cells in the flask was 5 mW/cm(2). The mean specific absorption rate, calculated by finite difference time domain analysis, was 2.135 (+/-0.005 SE) W/kg. Aliquots of whole blood that were sham-exposed or exposed in vitro to 50 cGy of ionizing radiation from a (137)Cs gamma-ray source were used as controls. The lymphocytes were examined to determine the extent of primary DNA damage (single-strand breaks and alkali-labile lesions) using the alkaline comet assay with three different slide-processing schedules. The assay was performed on the cells immediately after the exposures and at 4 h after incubation of the exposed blood at 37 +/- 1 degrees C to allow time for rejoining of any strand breaks present immediately after exposure, i.e. to assess the capacity of the lymphocytes to repair this type of DNA damage. At either time, the data indicated no significant differences between RF-radiation- and sham-exposed lymphocytes with respect to the comet tail length, fluorescence intensity of the migrated DNA in the tail, and tail moment. The conclusions were similar for each of the three different comet assay slide-processing schedules examined. In contrast, the response of lymphocytes exposed to ionizing radiation was significantly different from RF-radiation- and sham-exposed cells. Thus, under the experimental conditions tested, there is no evidence for induction of DNA single-strand breaks and alkali-labile lesions in human blood lymphocytes exposed in vitro to pulsed-wave 2450 MHz radiofrequency radiation, either immediately or at 4 h after exposure.     

Cylindrical waveguide applicator for in vitro exposure of cell culture samples to 1.9-GHz radiofrequency fields

An applicator for in vitro cell culture exposure was developed based on a circularly polarized, cylindrical waveguide for the 1.9-GHz frequency band used by Personal Communications Services (PCS) in Canada. The applicator consists of two coaxial Petri dishes that sit on the open end of the cylindrical waveguide. The inner 60-mm Petri dish contains the cell culture while the outer 150-mm dish contains coolant water, which is circulated from a pump. A dosimetric evaluation was made using thermometric and E-field probe techniques. The latter allowed the entire inner dish to be scanned to determine the range of specific absorption rates (SARs) pertinent to the expected position of the cells. A representative SAR rate (SAR per unit of input power) of 8.6 +/- 2.1 W/kg/W (95th percentile) was determined 1 mm from the bottom, for a 10 ml sample volume of standard medium. Evaluation of the cooling system demonstrated that following an initial 0.3 degrees C temperature increase, a constant temperature was maintained for 24 h when the waveguide was energized to achieve an average sample SAR of 10 W/kg. These properties enable both acute and sub-acute in vitro bio-effect studies to be performed on a variety of cell culture samples.     

Cytogenetic studies in human blood lymphocytes exposed in vitro to 2.45 GHz or 8.2 GHz radiofrequency radiation

Peripheral blood samples collected from healthy human volunteers were exposed in vitro to 2.45 GHz or 8.2 GHz pulsed-wave radiofrequency (RF) radiation. The net forward power, average power density, mean specific absorption rate, and the temperature maintained during the 2-h exposure of the cells to 2.45 GHz or 8.2 GHz were, respectively, 21 W or 60 W, 5 mW/cm(2) or 10 mW/cm(2), 2.13 W/kg or 20.71 W/kg, and 36.9 +/- 0.1 degrees C or 37.5 +/- 0.2 degrees C. Aliquots of the same blood samples that were either sham-exposed or exposed in vitro to an acute dose of 1.5 Gy gamma radiation were used as unexposed and positive controls, respectively. Cultured lymphocytes were examined to determine the extent of cytogenetic damage assessed from the incidence of chromosomal aberrations and micronuclei. Under the conditions used to perform the experiments, the levels of damage in RF-radiation-exposed and sham-exposed lymphocytes were not significantly different. Also, there were no significant differences in the response of unstimulated lymphocytes and lymphocytes stimulated with phytohemagglutinin when exposed to 8.2 GHz RF radiation. In contrast, the positive control cells that had been subjected to gamma irradiation exhibited significantly more damage than RF-radiation- and sham-exposed lymphocytes.     

Studying the synergistic damage effects induced by 1.8 GHz radiofrequency field radiation (RFR) with four chemical mutagens on human lymphocyte DNA using comet assay in vitro

The aim of this investigation was to study the synergistic DNA damage effects in human lymphocytes induced by 1.8 GHz radiofrequency field radiation (RFR, SAR of 3 W/kg) with four chemical mutagens, i.e. mitomycin C (MMC, DNA crosslinker), bleomycin (BLM, radiomimetic agent), methyl methanesulfonate (MMS, alkylating agent), and 4-nitroquinoline-1-oxide (4NQO, UV-mimetic agent). The DNA damage of lymphocytes exposed to RFR and/or with chemical mutagens was detected at two incubation time (0 or 21 h) after treatment with comet assay in vitro. Three combinative exposure ways were used. Cells were exposed to RFR and chemical mutagens for 2 and 3h, respectively. Tail length (TL) and tail moment (TM) were utilized as DNA damage indexes. The results showed no difference of DNA damage indexes between RFR group and control group at 0 and 21 h incubation after exposure (P>0.05). There were significant difference of DNA damage indexes between MMC group and RFR+MMC co-exposure group at 0 and 21 h incubation after treatment (P<0.01). Also the significant difference of DNA damage indexes between 4NQO group and RFR+4NQO co-exposure group at 0 and 21 h incubation after treatment was observed (P<0.05 or P<0.01). The DNA damage in RFR+BLM co-exposure groups and RFR+MMS co-exposure groups was not significantly increased, as compared with corresponding BLM and MMS groups (P>0.05). The experimental results indicated 1.8 GHz RFR (SAR, 3 W/kg) for 2h did not induce the human lymphocyte DNA damage effects in vitro, but could enhance the human lymphocyte DNA damage effects induced by MMC and 4NQO. The synergistic DNA damage effects of 1.8 GHz RFR with BLM or MMS were not obvious.     

DNA damage and apoptosis in the immature mouse cerebellum after acute exposure to a 1 mT, 60 Hz magnetic field.

Several recent studies have reported that whole-body exposure of rodents to power frequency magnetic fields (MFs) can result in DNA single- and double-strand breaks in the brains of these animals. The current study was undertaken to investigate whether an acute 2h exposure of a 1 mT, 60 Hz MF could elicit DNA damage, and subsequently apoptosis, in the brains of immature (10-day-old) mice. DNA damage was quantitated at 0, 2, 4, and 24h after exposure using the alkaline comet assay. Apoptosis was quantitated in the external granule cell layer (EGCL) of the immature mouse cerebellum at 0 and 24h after exposure to MF by the TdT-mediated dUTP nick-end labeling (TUNEL) assay. Four parameters (tail ratio, tail moment, comet length and tail length) were used to assess DNA damage for each comet. While increased DNA damage was detected by tail ratio at 2h after MF exposure, no supporting evidence of increased DNA damage was detected by the other parameters. In addition, no similar differences were observed using these parameters at any of the other post-exposure times. No increase in apoptosis was observed in the EGCL of MF-exposed mice, when compared to sham mice. Taken together, these results do not support the hypothesis that acute MF exposure causes DNA damage in the cerebellums of immature mice.     

Influence of iron-overload on DNA damage and its repair in human leukocytes in vitro

Iron is an important element that modulates the production of reactive oxygen species, which are thought to play a causative role in biological processes such as mutagenesis and carcinogenesis. The potential genotoxicity of dietary iron has been seldom studied in human leukocyte and only few reports have investigated in human colon tumor cells. Therefore, DNA damage and repair capacity of human leukocytes were examined using comet assay for screening the potential toxicity of various iron-overloads such as ferric-nitrilotriacetate (Fe-NTA), FeSO(4), hemoglobin and myoglobin, and compared with 200μM of H(2)O(2) and HNE. The iron-overloads tested were not cytotoxic in the range of 10-1000 microM by trypan blue exclusion assay. The exposure of leukocytes to Fe-NTA (500 and 1000 microM), FeSO(4) (250-1000 microM), hemoglobin (10 microM) and myoglobin (250 microM) for 30 min induced significantly higher DNA damage than NC. Treatment with 500 and 1000 microM of Fe-NTA showed a similar genotoxic effect to H(2)O(2), and a significant higher genotoxic effect than HNE. The genotoxicity of FeSO(4) (250-1000 microM), hemoglobin (10 microM) and myoglobin (250 microM) was not significantly different from that of H(2)O(2) and HNE. Iron-overloads generated DNA strand break were rejoined from the first 1h. Their genotoxic effect was not observed at 24h. These data from this study provide additional information on the genotoxicity of iron-overloads and self-repair capacity in human leukocytes.