Figure 4.

AEG-1−/− hepatocytes are pro-senescent and susceptible to DEN-induced DNA damage. AEG-1+/+ and AEG-1−/− hepatocytes, isolated from AEG-1^fl/fl and AEG-1^ΔHEP mice, respectively, were cultured for 3 days and senescence was determined by Senescence-associated β-galactosidase (SA-β-Gal) assay (A) and immunofluorescence (IF) staining for γ-H2AX (B-C). For B, magnification: 630x. D. Proliferation of AEG-1+/+ and AEG-1−/− hepatocytes, isolated from AEG-1^fl/fl and AEG-1^ΔHEP mice, respectively, was analyzed by MTT assay 48 h after DEN (25 ng/mL) treatment. E. Western blot for the indicated proteins in AEG-1+/+ and AEG-1−/− hepatocytes, isolated from AEG-1^fl/fl and AEG-1^ΔHEP mice, respectively, treated or not with DEN (25 ng/mL) for 8 h. GAPDH was used as loading control and one representative image for GAPDH levels is shown. F. AEG-1^fl/fl, AEG-1^ΔHEP and AEG-1^ΔMAC mice were treated with DEN (10 μg/gm) and apoptosis was determined by TUNEL assay (left panel) and proliferation was determined by BrdU incorporation assay (right panel). For A and C, data represent mean ± SEM. *: p<0.01 vs hepatocytes isolated from AEG-1^fl/fl mice. For D, data represent mean ± SEM. *: p<0.01 vs corresponding No Rx, #: p<0.01 vs hepatocytes isolated from AEG-1^fl/fl mice. For F, data represent mean ± SEM. *: p<0.01 vs AEG-1^fl/fl and AEG-1^ΔMAC.