Figure 6. Effect of FoxO1 on VLDL-TG production in HepG2 cells.

HepG2 cells in 6-well plates were transduced with either control or FoxO1-ADA vector at an MOI of 100 pfu/cell. (A) After 24-h incubation, conditioned media were collected for the determination of TG levels. Cells were collected for the determination of apoB100 mRNA abundance by real-time quantitative RT-PCR assay using β-actin mRNA as control (B), and for determination of apoB100 protein levels by semiquantitative immunoblot assay using anti-apoB100 and anti-actin antibodies (C). In addition, HepG2 cells pretransduced with Adv–FoxO1-ADA and Adv-null vectors were pulse-labeled with 200 μCi/ml of L-[³⁵S]-methionine for 1 h, followed by a 30-min chase with 10 mM unlabeled methionine. Conditioned medium and cell lysates were subjected to electrophoresis on 5% SDS-polyacrylamide gels, which were dried prior to autoradiography. Gel slices corresponding to apoB100 protein bands were excised from gels with the aid of autoradiogram for the determination of radioactive content of apoB100 proteins that were secreted into conditioned medium (D) and present in cells (E). Data were from 3 experiments. *P < 0.05; **P < 0.001 versus control.