Vinita Chauhan

Microarray Gene Expression Profiling of a Human Glioblastoma Cell Line Exposed In Vitro to a 1.9 GHz Pulse-Modulated Radiofrequency Field

Abstract Qutob, S. S., Chauhan, V., Bellier, P. V., Yauk, C. L., Douglas, G. R., Berndt, L., Williams, A., Gajda, G. B., Lemay, E., Thansandote, A. and McNamee, J. P. Microarray Gene Expression Profiling of a Human Glioblastoma Cell Line Exposed In Vitro to a 1.9 GHz Pulse-Modulated Radiofrequency Field. Radiat. Res. 165, 636–644 (2006). 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°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.

Development of Laser Desorption Imaging Mass Spectrometry Methods to Investigate the Molecular Composition of Latent Fingermarks

AbstractFor a century, fingermark analysis has been one of the most important and common methods in forensic investigations. Modern chemical analysis technologies have added the potential to determine the molecular composition of fingermarks and possibly identify chemicals a suspect may have come into contact with. Improvements in analytical detection of the molecular composition of fingermarks is therefore of great importance. In this regard, matrix-assisted laser desorption ionization (MALDI) and laser desorption ionization (LDI) imaging mass spectrometry (IMS) have proven to be useful technologies for fingermark analysis. In these analyses, the choice of ionizing agent and its mode of deposition are critical steps for the identification of molecular markers. Here we propose two novel and complementary IMS approaches for endogenous and exogenous substance detection in fingermarks: sublimation of 2-mercaptobenzothiazol (2-MBT) matrix and silver sputtering. Graphical Abstractᅟ

Gene Expression Analysis of a Human Lymphoblastoma Cell Line Exposed In Vitro to an Intermittent 1.9 GHz Pulse-Modulated Radiofrequency Field

Abstract Chauhan, V., Mariampillai, A., Bellier, P. V., Qutob, S. S., Gajda, G. B., Lemay, E., Thansandote, A. and McNamee, J. P. Gene Expression Analysis of a Human Lymphoblastoma Cell Line Exposed In Vitro to an Intermittent 1.9 GHz Pulse-Modulated Radiofrequency Field. Radiat. Res. 165, 424–429 (2006). This study was designed to determine whether radiofrequency (RF) fields of the type used for wireless communications could elicit a cellular stress response. As general indicators of a cellular stress response, we monitored changes in proto-oncogene and heat-shock protein expression. Exponentially growing human lymphoblastoma cells (TK6) were exposed to 1.9 GHz pulse-modulated RF fields at average specific absorption rates (SARs) of 1 and 10 W/kg. Perturbations in the expression levels of the proto-oncogenes FOS, JUN and MYC after exposure to sham and RF fields were assessed by real-time RT-PCR. In addition, the transcript levels of the cellular stress proteins HSP27 and inducible HSP70 were also monitored. We demonstrated that transcript levels of these genes in RF-field-exposed cells showed no significant difference in relation to the sham treatment group. However, concurrent positive (heat-shock) control samples displayed a significant elevation in the expression of HSP27, HSP70, FOS and JUN. Conversely, the levels of MYC mRNA were found to decline in the positive (heat-shock) control. In conclusion, our study found no evidence that the 1.9 GHz RF-field exposure caused a general stress response in TK6 cells under our experimental conditions.

Analysis of proto-oncogene and heat-shock protein gene expression in human derived cell-lines exposed in vitro to an intermittent 1.9 GHz pulse-modulated radiofrequency field.

Purpose: Several studies have reported that radiofrequency (RF) fields, as emitted by mobile phones, may cause changes in gene expression in cultured human cell-lines. The current study was undertaken to evaluate this possibility in two human-derived immune cell-lines. Materials and methods: HL-60 and Mono-Mac-6 (MM6) cells were individually exposed to intermittent (5 min on, 10 min off) 1.9 GHz pulse-modulated RF fields at a average specific absorption rate (SAR) of 1 and 10 W/kg at 37 ± 0.5°C for 6 h. Concurrent negative and positive (heat-shock for 1 h at 43°C) controls were conducted with each experiment. Immediately following RF field exposure (T = 6 h) and 18 h post-exposure (T = 24 h), cell pellets were collected from each of the culture dishes and analyzed for transcript levels of proto-oncogenes (c-jun, c-myc and c-fos) and the stress-related genes (heat shock proteins (HSP) HSP27 and HSP70B) by quantitative reverse transcriptase polymerase chain reaction (RT-PCR). Results: No significant effects were observed in mRNA expression of HSP27, HSP70, c-jun, c-myc or c-fos between the sham and RF-exposed groups, in either of the two cell-lines. However, the positive (heat-shock) control group displayed a significant elevation in the expression of HSP27, HSP70, c-fos and c-jun in both cell-lines at T = 6 and 24 h, relative to the sham and negative control groups. Conclusion: This study found no evidence that exposure of cells to non-thermalizing levels of 1.9 GHz pulse-modulated RF fields can cause any detectable change in stress-related gene expression.

Evaluating the Biological Effects of Intermittent 1.9 GHz Pulse-Modulated Radiofrequency Fields in a Series of Human-Derived Cell Lines

Abstract Chauhan, V., Mariampillai, A., Kutzner, B., Wilkins, R. C., Ferrarotto, C., Bellier, P. V., Marro, L., Gajda, G. B., Lemay, E., Thansandote, A. and McNamee, J. P. Evaluating the Biological Effects of Intermittent 1.9 GHz Pulse-Modulated Radiofrequency Fields in a Series of Human-Derived Cell Lines. Radiat. Res. 167, 87–93 (2007). 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°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 (G2/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.

Effects of ambient air particles on the endothelin system in human pulmonary epithelial cells (A549)

Inhalation of urban particles results in higher circulating levels of the vasoconstrictor peptide endothelin-1 (ET-1), which may account for the adverse cardiovascular impacts associated with air pollution. The objective of this study was to examine the direct effects of urban particles on the production of ET-1 by human epithelial cells (A549). A549 cells were exposed to TiO2, SiO2, Ottawa urban particulate matter EHC-93, and fractions of the urban particles. The levels of ET-1, interleukin-8 (IL-8) and vascular endothelial growth factor (VEGF) in the culture medium were detected by ELISA. The mRNA levels of preproET-1, endothelin converting enzyme (ECE-1), ETa receptor and ETb receptor, matrix metalloproteinase (MMP-2), tissue inhibitor of MMP (TIMP-2), and heat shock protein (HSP-70) were determined by quantitative real-time RT-PCR. Cluster analysis of the variables identified similarities in the patterns of effects. Cluster I comprised variables that were primarily inhibited by particles: ET-1 and MMP-2 mRNAs, ET-1 and bigET-1 peptides, and cell viability. Clusters II and III comprised variables that were either inhibited or induced, depending on the test material: HSP-70, ETaR and ECE mRNAs, and IL-8 and VEGF proteins. Cluster IV comprised variables that were mainly induced by particle preparations: ETbR and TIMP-2 mRNAs. The decreased expression of preproET-1 in A549 cells suggests that epithelial cells may not be the source of higher pulmonary ET-1 spillover in the circulation measured in vivo in response to inhaled urban particles. However, higher ECE-1 in A549 cells after exposure to particles suggests an increased ability to process bigET-1 into the mature ET-1 peptide, while increased receptor expression implies higher responsiveness. The increased release of IL-8 and VEGF by epithelial cells in response to particles could possibly upregulate ET-1 production in the adjacent pulmonary capillary endothelial cells, with concomitant increased ET-1 spillover in the systemic circulation.

Evaluating DNA Damage in Rodent Brain after Acute 60 Hz Magnetic-Field Exposure

Abstract McNamee, J. P., Bellier, P. V., Chauhan, V., Gajda, G. B., Lemay, E. and Thansandote, A. Evaluating DNA Damage in Rodent Brain after Acute 60 Hz Magnetic-Field Exposure. Radiat. Res. 164, 791–797 (2005). In recent years, numerous studies have reported a weak association between 60 Hz magnetic-field exposure and the incidence of certain cancers. To date, no mechanism to explain these findings has been identified. The objective of the current study was to investigate whether acute magnetic-field exposure could elicit DNA damage within brain cells from both whole brain and cerebellar homogenates from adult rats, adult mice and immature mice. Rodents were exposed to a 60 Hz magnetic field (0, 0.1, 1 or 2 mT) for 2 h. Then, at 0, 2 and 4 h after exposure, animals were killed humanely, their brains were rapidly removed and homogenized, and cells were cast into agarose gels for processing by the alkaline comet assay. Four parameters (tail ratio, tail moment, comet length and tail length) were used to assess DNA damage for each comet. For each species, a significant increase in DNA damage was detected by each of the four parameters in the positive control (2 Gy X rays) relative to the concurrent nonirradiated negative and sham controls. However, none of the four parameters detected a significant increase in DNA damage in brain cell homogenates from any magnetic-field exposure (0– 2 mT) at any time after exposure. The dose–response and time-course data from the multiple animal groups tested in this study provide no evidence of magnetic-field-induced DNA damage.

Identification of gene-based responses in human blood cells exposed to alpha particle radiation

BackgroundThe threat of a terrorist-precipitated nuclear event places humans at danger for radiological exposures. Isotopes which emit alpha (α)-particle radiation pose the highest risk. Currently, gene expression signatures are being developed for radiation biodosimetry and triage with respect to ionizing photon radiation. This study was designed to determine if similar gene expression profiles are obtained after exposures involving α-particles.MethodsPeripheral blood mononuclear cells (PBMCs) were used to identify sensitive and robust gene-based biomarkers of α-particle radiation exposure. Cells were isolated from healthy individuals and were irradiated at doses ranging from 0-1.5 Gy. Microarray technology was employed to identify transcripts that were differentially expressed relative to unirradiated cells 24 hours post-exposure. Statistical analysis identified modulated genes at each of the individual doses.ResultsTwenty-nine genes were common to all doses with expression levels ranging from 2-10 fold relative to control treatment group. This subset of genes was further assessed in independent complete white blood cell (WBC) populations exposed to either α-particles or X-rays using quantitative real-time PCR. This 29 gene panel was responsive in the α-particle exposed WBCs and was shown to exhibit differential fold-changes compared to X-irradiated cells, though no α-particle specific transcripts were identified.ConclusionCurrent gene panels for photon radiation may also be applicable for use in α-particle radiation biodosimetry.

Development and characterization of an in vitro alpha radiation exposure system

A simple in vitro alpha radiation exposure system (ARES) was designed to study the biological effects of alpha particle radiation. The ARES consists of six (241)Am electroplated stainless steel discs with activities averaging 66 kBq and Mylar-based culture dishes to allow the transmission of alpha particles. The dosimetry of the exposure system was calculated using the GEANT4 Monte Carlo simulation toolkit with the source code adapted from the open-source Microbeam example. The average dose rate and linear energy transfer of the system was simulated to be 0.98 ± 0.01 (statistical)(+0.18)( - 0.00) (systematic) Gy h(-1) and 127.4 ± 0.4 (statistical)(+23)( - 0) (systematic) keV µm(-1), respectively. The system was characterized by a comparison of the survival curves of gamma and alpha irradiated cell lines which showed a relative biological effectiveness of 6.3. This is in good agreement with values obtained using other published alpha particle exposure systems. Results show that the ARES provides a simple, cost-effective exposure platform for research into the biological effects of alpha particle radiation using in vitro modelling of cell cultures.

Effects of ambient air particles on nitric oxide production in macrophage cell lines

We assessed thein vitro toxicity of various particles on three murine macrophage cell lines, (J774A.1, WR19M.1, RAW264.7). The cells were exposed to aqueous suspensions (0–100 μg/30 mm2 well) of urban particulate matter (SRM-1648, SRM-1649, EHC-93), fine particulate matter (PM2.5), titanium dioxide (SRM-154b), and respirable cristobalite (SRM-1879) for 2 h and were then stimulated with lipopolysaccharide (LPS, 100 ng/ml) and recombinant interferon-gamma (IFN, 100 U/ml). After overnight incubation with the particles and LPS/IFN, nitric oxide production was estimated from culture supernatant nitrite. Cell viability was determined by monitoring the rate of AlamarBlue™ reduction. The dose-effect relationships for nitrite and viability were modeled as a power function (Fold change=[Dose+1]β), where β represents the slope of the dose-response curve. Potency was defined as the rate of change in nitrite production corrected for cell viability (βPOTENCY = βNITRITE − βVIABILITY). Overall, the urban particles decreased nitric oxide production (βPOTENCY < 0), while exposure of the cells to fine particulate matter or cristobalite increased the production of nitric oxide (βPOTENCY > 0). Titanium dioxide (TiO2) was essentially inactive (βPOTENCY ≈ 0). The decrease in nitric oxide production seen in cells exposed to the urban particles was directly correlated to a decrease in the expression of inducible nitric oxide (iNOS) as determined by Western blot analysis. The results indicate that particles are modulators of nitric oxide production in murine macrophages and may directly disrupt expression of iNOS during concomitant pathogen exposure. Pathways leading to enhanced NO production causing cell injury, and to decreased NO release resulting in lower bacterial clearance, may both be relevant to the health effects of ambient particles.