<i>H</i>. <i>pylori</i> infection promotes the genes and proteins expression in gastric cancer tissues.

<p>(A) Western blot analysis of the indicated proteins in the control cells (1), <i>H</i>.<i>pylori</i>-infected SGC-7901 cells (2) and <i>cagA</i>-overexpressed SGC-7901 cells (3). GAPDH served as the loading control. The data are representative of three independent experiments. (B) Quantitative RT-PCR analysis of the indicated genes in 30 gastric cancer tissues. Values are represented as average Ct fold compared to peri-cancerous tissues, where peri-cancerous tissues were set to 1. (C) Western blot analysis of the indicated proteins in 30 gastric cancer tissues. 200 mg tissues were homogenized and total proteins were collected. A total of 50 μg protein extracts were subjected to SDS-PAGE gel electrophoresis. GAPDH served as the loading control. (D) Detection of the <i>H</i>. <i>pylori</i> 16S <i>rRNA</i> gene and <i>cagA</i> gene in gastric cancer tissues by PCR (Upper). M represents the DNA molecular weight marker. Lane 1 is the positive control. Lane 2 is the negative control. Lanes 3, 4 and 6 are positive samples. Lanes 5 are negative samples. Quantitative RT-PCR analysis of the indicated genes in gastric cancer tissues with and without <i>H</i>. <i>pylori</i> infection (Lower). Values are presented as the average Ct fold compared to the group without <i>H</i>. <i>pylori</i> infection, which was set to 1. The figure presents the average of 30 samples. Data are presented as the means ± SD. Error bars represent standard deviations. LDH, L-lactate dehydrogenase. DLD, Dihydrolipoamide dehydrogenase. PRPF19, pre-mRNA processing factor 19 homolog. RanGAP, Ran-specific GTPase-activating protein. CaM, calmodulin. p64 CLCP, nuclear chloride ion channel protein. *, <i>P</i><0.05 compared to peri-cancerous tissue (B and C) and tissues without <i>H</i>. <i>pylori</i> infection (D).</p

Introduction of CagA into gastric cancer cells.

<p>(A and B) Western blot analysis of CagA and phosphorylated CagA in <i>H</i>. <i>pylori</i>-infected SGC-7901(A) and AGS (B) cells. The cells infected with the indicated ratio of cells to <i>H</i>. <i>pylori</i> for the indicated time were collected and lysed, and the proteins were separated by SDS-PAGE. Cells infected with <i>H</i>. <i>pylori</i> boiled for 15 min at a MOI of 1:1000 were used as a control. (C and D) Detection of CagA mRNA and protein in <i>cagA</i>-overexpressing SGC-7901 cells by RT-PCR (C) and western blot (D). GAPDH served as the loading control. The data are representative of three independent experiments. <i>Hp</i>, <i>H</i>. <i>pylori</i>; P-CagA, phosphorylated CagA; GAPDH, Glyceraldehyde-3-phosphate- dehydrogenase.</p

Representative 2-DE maps and magnified image of differential spots in three cell lines.

<p>SGC-7901 and AGS cells infected with <i>H</i>. <i>pylori</i> for 6 h at a MOI of 1:1000 (cell to <i>H</i>. <i>pylori</i>) and SGC-7901 cells transfected with pcDNA3.1/<i>cagA</i> for 48 h were collected and lysed, and the protein concentrations were determined using Bradford colorimetry. A total of 800 μg of protein was loaded for two-dimensional electrophoresis. Cells infected with boiled <i>H</i>. <i>pylori</i> or transfected with empty vector served as controls for the infected or transfected cells, respectively. (A) SGC-7901 cells infected with <i>H</i>. <i>pylori</i>. (B) AGS cells infected with <i>H</i>. <i>pylori</i>. (C) SGC-7901 cells transfected with the <i>cagA</i>-vector. (D) Magnified image of 10 differential spots. B4, B11, C6, C7, C8 and C9 spots were up-regulated, whereas D12, D16, E2 and E11 spots were down-regulated. These spots are identified in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0146521#pone.0146521.t003" target="_blank">Table 3</a>.</p