Additional file 1: Figure S1. of A system biology approach to understanding the molecular mechanisms of Gubentongluo decoction acting on IgA Nephropathy

Overview of data generation, processing and analysis. (JPEG 118 kb

RF-EMF-induced γH2AX foci formation does not change cell cycle distribution in HSF cells.

<p>Histograms show the percentages of HSF cells in different phases of the cell cycle at 0 (left), 6 (middle), and 12 h (right) after 24 h exposure to 1800 MHz RF-EMF at 3.0 W/kg. Values represent mean ± SEM of 5 independent experiments. *<i>p</i><0.05 compared with sham-exposed sample (Student’s t-test).</p

1800 MHz RF-EMF induces cell type-dependent DSBs as evaluated by the γH2AX foci formation assay.

<p>(A) Representative images of γH2AX immunofluorescent staining of CHL, astrocytes, FL, HUVEC, HLEC, and HSF exposed to radiation at 3.0 W/kg for either 1 h or 24 h. Red dots indicate γH2AX foci; nuclei are stained blue with DAPI. Scale bar, 10 µm. (B) Histograms showing the average numbers of γH2AX foci per cell by scoring ∼200 cells per sample. Values are mean ± SEM of at least 4 independent experiments. *<i>p</i><0.05 compared with the sham-exposed sample.</p

Cell Type-Dependent Induction of DNA Damage by 1800 MHz Radiofrequency Electromagnetic Fields Does Not Result in Significant Cellular Dysfunctions

<div><h3>Background</h3><p>Although IARC clarifies radiofrequency electromagnetic fields (RF-EMF) as possible human carcinogen, the debate on its health impact continues due to the inconsistent results. Genotoxic effect has been considered as a golden standard to determine if an environmental factor is a carcinogen, but the currently available data for RF-EMF remain controversial. As an environmental stimulus, the effect of RF-EMF on cellular DNA may be subtle. Therefore, more sensitive method and systematic research strategy are warranted to evaluate its genotoxicity.</p> <h3>Objectives</h3><p>To determine whether RF-EMF does induce DNA damage and if the effect is cell-type dependent by adopting a more sensitive method γH2AX foci formation; and to investigate the biological consequences if RF-EMF does increase γH2AX foci formation.</p> <h3>Methods</h3><p>Six different types of cells were intermittently exposed to GSM 1800 MHz RF-EMF at a specific absorption rate of 3.0 W/kg for 1 h or 24 h, then subjected to immunostaining with anti-γH2AX antibody. The biological consequences in γH2AX-elevated cell type were further explored with comet and TUNEL assays, flow cytometry, and cell growth assay.</p> <h3>Results</h3><p>Exposure to RF-EMF for 24 h significantly induced γH2AX foci formation in Chinese hamster lung cells and Human skin fibroblasts (HSFs), but not the other cells. However, RF-EMF-elevated γH2AX foci formation in HSF cells did not result in detectable DNA fragmentation, sustainable cell cycle arrest, cell proliferation or viability change. RF-EMF exposure slightly but not significantly increased the cellular ROS level.</p> <h3>Conclusions</h3><p>RF-EMF induces DNA damage in a cell type-dependent manner, but the elevated γH2AX foci formation in HSF cells does not result in significant cellular dysfunctions.</p> </div

RF-EMF-induced γH2AX foci formation does not affect cell proliferation and viability in HSF cells.

<p>(A) HSF cell numbers at 0, 12, 24, and 48 h after 24 h exposure to 1800 MHz RF-EMF at 3.0 W/kg. (B) HSF cell viability at 0, 1, 2, 3, and 4 days after re-seeding at 1000 cells/well (B, left) and 2000 cells/well (B, right) immediately after 24 h exposure. 1 µM 4NQO treatment for 1 h serves as positive control. Values represent mean ± SEM of 3 independent experiments. *<i>p</i><0.05 and **<i>p</i><0.01 compared with sham-exposed sample (Student’s t-test).</p

RF-EMF-induced γH2AX foci formation does not result in more DNA nicks in HSF cells.

<p>Histogram of DNA fragment levels in HSF cells. The background fluorescence value of the cells (NC) was determined without adding rTdT, and 1 µM 4NQO treatment for 1 h serves as positive control. Values represent mean ± SEM of 3 independent experiments. *<i>p</i><0.05 compared with sham-exposed sample (Student’s t-test).</p

Effect of RE-EMF exposure on ROS production in HSF cells.

<p>Histogram of ROS levels in HSF cells. The intracellular ROS level of 24 h exposed cells was measured by flow cytometry using DCFH-DA. The background fluorescence value of the cells (NC) was determined without adding DCFH-DA, and 1 µM 4NQO treatment for 1 h serves as positive control. Values represent mean ± SEM of 7 independent experiments. **<i>p</i><0.01 compared with sham-exposed sample (Student’s t-test).</p

Associations^a of sleep duration^b with various electromagnetic exposures.

a<p>Adjusted for age, gender, cigarette smoking, tea drinking, BMI, and work pressure.</p>b<p>Medium sleep indicates that sleep duration is between 7 and 8 hours, short sleep is less than 7 hours, and long sleep is more than 8 hours.</p>c<p>The cut points of tertiles of daily occupational exposure time (DOET) were ≤1.5 hours/day for T1, 1.5 4 hours/day for T3.</p><p>Associations^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110825#nt105" target="_blank">a</a>} of sleep duration^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110825#nt106" target="_blank">b</a>} with various electromagnetic exposures.</p

Demographic characteristics of study subjects stratified by sleep quality.

a<p>The unmarried include those who are single, divorced or widowed.</p>b<p>Comparison between groups was tested used Student’s <i>t</i>-test for the continuous variables and Pearson’s χ² test for the categorical variables.</p><p>Demographic characteristics of study subjects stratified by sleep quality.</p

The associations^a of daily occupational exposure time (DOET)^b with sleep quality stratified by occupational exposure duration^c.

a<p>Adjusted for age, gender, cigarette smoking, tea drinking, BMI, and work pressure.</p>b<p>The cut points of tertiles of daily occupational exposure time (DOET) were ≤1.5 hours/day for T1, 1.5 4 hours/day for T3.</p>c<p>Short occupational exposure duration means less than or equals 23 years, while long occupational exposure duration means more than 23 years.</p><p>The associations^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110825#nt108" target="_blank">a</a>} of daily occupational exposure time (DOET)^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110825#nt109" target="_blank">b</a>} with sleep quality stratified by occupational exposure duration^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110825#nt110" target="_blank">c</a>}.</p