jcmm12071-sup-0001-FigS1.tifimage/tif71KFigure S1. H2O2 causes transcriptional upregulation of genes of diverse signal transduction pathways in TE7 cells. Differential gene expression profile was detected by cDNA microarray analysis. Cells were treated with 250 μM H2O2 or were left untreated (co) and grown for 30 min. Subsequently, the RNA was isolated and reverse transcription was performed. The cDNA was hybridized with the fixed cDNA samples of the ‘GEArray Q Series Human Signal Transduction PathwayFinder Gene Array’. The hybridization extent was measured by chemiluminescence and evaluated by densitometry. Blank value was subtracted from gene raw values. The received gene values were adjusted with the housekeeping gene β-actin value by calculating the ratio.
jcmm12071-sup-0002-FigS2.tifimage/tif38KFigure S2. H2O2 decreases proliferation and increases apoptosis. (A) 5-bromo-2′-deoxyuridine (BrdU) proliferation ELISA showed reduced proliferation of H2O2-treated TE7 cells, as well as abolished proliferation of CDNB-treated TE7 cells. Cells were treated with 250 μM H2O2 or 10 μM CDNB or were left untreated or treated with ethanol, and grown for 24 hrs. BrdU assay was performed as described in Materials and methods by measuring the absorbance of BrdU-positive cells at 450 nm following treatment. Data are mean ± SD. (B) H2O2 causes apoptosis and necrosis in a dose-dependent manner. Apoptotic, necrotic and live cell populations were measured after 24 hrs using Annexin-V and PI staining. Mean values are shown ± SD.
jcmm12071-sup-0003-FigS3.tifimage/tif45KFigure S3. p21WAF1 is induced via the JNK-ATF2-pathway after H2O2 treatment (250 μM) in TE7 cells. Inhibition of JNK activity using SP600125 revealed decreased activation of ATF2 and c-Jun, therefore causing reduced p21WAF1 expression. Cells were treated with H2O2 and grown for 1, 3, 6, 12, and 24 hrs following pre-incubation for 1 hr with 10 μM of JNK inhibitor SP600125. Lysates were immunoblotted for total ATF2 and c-Jun, as well as for their activated forms p-ATF2Thr69/71 and p-c-JunSer73, and for p21WAF1. β-actin was used as loading control. Fold expression changes are given below the blots.
jcmm12071-sup-0004-TableS1-S2.docxWord document18K

Table S1. Differential gene expression profile of H2O2-treated TE7 cells detected by cDNA microarray analysis (Figure S1).

Table S2 PubMed search for potential target genes in oesophageal cancer. State 01/30/13.

jcmm12071-sup-0005-SupportingInformation.docWord document34KData S1. Material and methods.

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