Fig. 5: The effect of MDK inhibition on the cell cycle and apoptotic cell death.

a Flow cytometric cell cycle analyses on control or anti-MDK (1 µg/ml)-treated NCI827 cells at 72 h are shown. The data are representative of three independent experiments. b Immunoblots for the indicated proteins upon treatment with the anti-MDK antibody for 0, 6 or 12 h in NCI827 (left) and NCI131 (right) cells are shown. The data are representative of three independent experiments. c Immunofluorescence images for EdU (red)- and/or Hoechst (blue)-stained NCI131 cells upon treatment with control IgG or the anti-MDK antibody (5 µg/ml) for 6 or 24 h are shown. The scale bar represents 100 µm. The data are representative of three independent experiments. d The relative survival of control cells and cells treated with the anti-MDK antibody (5 µg/ml) with or without CHIR-124 (Selleckchem) at the indicated doses is shown in the bar graphs. e An LDA plot and representative images of control spheres and spheres treated with anti-MDK with or without CHIR-124 are shown. f Flow cytometric analysis of Annexin V/PI-stained control (left) or anti-MDK (5 µg/ml, right)-treated NCI827 cells is shown. The data are representative of three independent experiments. g Quantification of data in immunofluorescence images for detecting caspase 3/7-positive control or anti-MDK antibody-treated NCI827 cells treated at the indicated doses is shown in the bar graph. h The percentage of caspase 3/7-positive control (black bar) or recombinant MDK (10 ng/ml, white bar)-treated cells with or without TBHP (50 µM) is shown in the bar graph. i The relative intensities of the indicated proteins in the apoptosis array upon treatment with IgG (black) or the anti-MDK antibody (red, 5 µg/ml, 12 h) are shown in the bar graph. *p < 0.05, **p < 0.01, and ***p < 0.001.