Fig. 3.

Akt phosphorylation was suppressed in CYP2E1-HepG2 cells compared with the NC-HepG2 cells. Monoclonal cell lines stably expressing CYP2E1 (CYP2E1-HepG2) and the control HepG2 (NC-HepG2) were established as described in the materials and methods sections. CYP2E1-HepG2 and NC-HepG2 were exposed to different doses of ethanol for 5 d, and the cellular TG levels and protein levels of Akt, p-Akt^ser473, p-Akt^thr308, GSK3β and p-GSK3β^ser9 were determined by western blotting. (a) Immunohistochemical staining and western blotting analysis showed that CYP2E1 was expressed in CYP2E1-HepG2 cells, but not in NC-HepG2 cells; (b) ethanol exposure led to increased protein levels of CYP2E1; (c) TG levels in CYP2E1-HepG2 were much higher than those in NC-HepG2 cells; (d)–(g) CYP2E1 expression suppressed the phosphorylation of Akt at Ser473 and Thr308, and inhibited the GSK3β at Ser9; (h) the cellular MDA levels in NC-HepG2 and CYP2E1-HepG2 cells exposed to ethanol (200 Mm) for 5 d; (i) CMZ treatment led to increased phosphorylation of Akt in CYP2E1-HepG2 cells. Data were presented as mean ± SD from at least 3 independent experiments, and expressed as the percentage of the control. ^*P < 0.05, ^**P < 0.01, compared with the control NC-HepG2 cells (0 mM group); ^#P < 0.05, ^##P < 0.01, compared with the control CYP2E1-HepG2 cells (0 mM group).