Effects of gamma-tototrienol on ApoB synthesis, degradation, and secretionin HepG2 cells

gamma-Tocotrienol (gamma-T3), a naturally occurring analog of tocopherol (v itamin E), has been shown to have a hypocholesterolemic effect in animals a nd humans. Unlike tocopherol, it has also been shown to reduce plasma apoB levels in hypercholesterolemic subjects. The aim of this study was to defin e the mechanism of action of gamma-T3 on hepatic modulation of apoB product ion using cultured HepG2 cells as the model system. HepG2 cells preincubate d with gamma-T3 were initially shown to inhibit the rate of incorporation o f [C-14]acetate into cholesterol in a concentration- and time-dependent man ner, with a maximum 86 +/- 3% inhibition at 50 mu mol/L observed within 6 h ours. gamma-T3, on the other hand, had no significant effect on the uptake of [C-14]glycerol into pools of cellular triacylglycerol and phospholipid r elative to untreated control. The rate of apoB synthesis and secretion was then studied by an [S-35]methionine pulse-labeling experiment and quantifie d by immunoprecipitating apoB on chasing up to 3 hours. An average reductio n of 24 +/- 3% in labeled apoB in the media was apparent with gamma-T3 desp ite a 60 +/- 2% increase in apoB synthesis. Fractionation of secreted apoB revealed a relatively denser lipoprotein particle, suggesting a less stable particle. Using a digitonin-permeabilized HepG2 cell system, the effects o f gamma-T3 on apoB translocation and degradation in the endoplasmic reticul um were further investigated. The generation of a specific N-terminal 70-kD a proteolytic fragment proved to be a sensitive measure of the rate of apoB translocation and degradation The abundance of this fragment increased sig nificantly in gamma-T3-treated cells relative to untreated control cells (5 0 +/- 21%) after 2 hours of chase. In addition, the presence of gamma-T3 re sulted in an average decrease of 64 +/- 8% in intact apoB. Taken together, the data suggest that gamma-T3 stimulates apoB degradation possibly as the result of decreased apoB translocation into the endoplasmic reticulum lumen . It is speculated that the lack of cholesterol availability reduces the nu mber of secreted apoB-containing lipoprotein particles by limiting transloc ation of apoB into the endoplasmic reticulum lumen.