Figure 1: H. pylori induces CagA-dependent EMT in gastric cancer cells.

(a) E-cadherin expressions in MKN28 gastric cancer cells infected with CagA-positive 60190 (left panels) or transfection of CagA expression vector (middle panels) were assessed by indirect immunofluorescence with anti-E-cadherin antibody (green) and 4',6-diamidino-2-phenylindole (DAPI) nuclear stain (blue). Following infection or transfection, the protein abundances of CagA, phospho-Tyr (pTyr) and E-cadherin were assessed by immunoblot analysis (right panels). Tubulin was used as the loading control; relative abundance of each loading control is denoted below the blot. (b) Confluent AGS cells were wounded and then infected with 60190 or isogenic CagA deletion mutant strain for 6 h (left panels). Protein lysates from AGS cells of the migration assay were analysed by immunoblotting for CagA translocation and pTyr (middle panels). Distance between the cell borders was measured from five independent sites after 6 h infection (right panel, **P<0.01 by Student’s t-test). Error bars indicate s.d. Scale bar, 100 μm. (c) The non-invasive MCF-7 cells were transiently co-transfected with either a control or CagA with GFP expression vector. The cells were cultured atop the embryonic chick chorioallantoic membrane (CAM) for 3 days and the fixed tissue sections then were examined by fluorescence microscopy. The upper face of the CAM is indicated by dashed white lines; invasive cancer cells are denoted by arrows. Scale bars, 300 μm. The number of GFP-positive invasive cells under the basement membrane were quantitated and statistically analysed by Student’s t-test (right panel). Error bars indicate s.d.