Fig. 4

Immunofluorescence of proteins involved in cell cycle progression and apoptosis. Microphotographs taken stitching 4 images of HepG2 cells cultured on a glass coverslip of a PFA-fixed immunofluorescence, representative for 3 experiments (scale bar = 50 μm). Without APAP (panels A, D, G, J): no staining (Null), control cells (CTRL), TEV- and EEV-treated cells. With APAP (panels B,E,H,K): vehicle-treated cells (DMSO), control cells (CTRL), TEV- and EEV-treated cells. A-B Ki67 staining (green) and DAPI counterstaining (blue) of HepG2 cells. C Diagram showing number of Ki67 positive cells in CTRL, TEV- and EEV-treated cells with or without 24 h APAP (or vehicle) treatment. Counting of Ki67 positive nuclei was performed in 3 different fields of view after reaching 200 counted nuclei. D-E p-p53 staining (green), p53 staining (red) and DAPI counterstaining (blue) of HepG2 cells. F Diagram showing the ratio between p-p53 and p53 positive cells. Counting of p-p53 or p53 positive nuclei was performed in 3 different fields of view after reaching 200 counted nuclei. G-H p21 staining (green) and DAPI counterstaining (blue) of HepG2 cells. I Diagram showing number of p21 positive cells. J-K p27 staining (red) and DAPI counterstaining (blue) of HepG2 cells. L Diagram showing number of p27 positive cells. For all panels, data are mean ± SD with n = 3 per group. *, p < 0.05 vs CTRL, Φ p < 0.05 vs APAP treatment, #, p < 0.05 vs vehicle (DMSO)