RESUMO
We developed an in vitro screening system for antihyperlipidemic activity by measuring lipoprotein profiles secreted from human intestinal epithelium-like cells from the colon cancer cell line, Caco-2. Sodium (Na) butyrate at 5 mM differentiated Caco-2 cells into intestinal epithelium-like cells and numerous microvilli on the apical side of cells were observed under transmission electron microscopy. Real-time RT-PCR analysis revealed that Na butyrate stimulated expression levels of intestinal differentiation markers in Caco-2 cells in a dose-dependent manner and 5 mM Na butyrate up-regulated intestinal alkaline phosphatase, sucrase-isomaltase complex, and microsomal triglyceride transfer protein by 8.1-, 1.9-, and 2.1-fold that of non-treated cells, respectively. Lipoprotein secretions from differentiated Caco-2 cells were promoted by lysophosphatidyl choline and Na oleate, which are a stimulator of lipoprotein secretion and a substrate of triglycerides, respectively. We examined the effects of Pluronic L-81, a lipoprotein secretion inhibitor, on lipoprotein profiles of differentiated Caco-2 cells. Pluronic L-81 at 1.0 µg/ml inhibited TG contents in lipoprotein fractions from cells by 25.6 % and secretion was completely suppressed by the agent at 10 µg/ml.
RESUMO
We isolated a HepG2-derived sub-clone (HepG2-Lipo), which possessed an increased lipoprotein synthesizing ability. HepG2-Lipo cells could secrete triglycerides (TG) and cholesterol at rates 9.4- and 6-fold higher, respectively, when compared to HepG2 cells. Real-time RT-PCR analysis revealed that the expression levels of sterol regulatory element-binding protein-1c and -2 were 2.9- and 1.5-fold higher than in HepG2 cells. Furthermore, two apolipoprotein (apo) genes (apoA-1 and apoB-100) in HepG2-Lipo cells were expressed at 2.8- and 1.9-fold higher levels when compared to those in parental cells. We examined the effects of three antihyperlipidemic agents on the lipoprotein profiles of HepG2-Lipo cells. Simvastatin at 5 microM selectively suppressed cholesterol secretion from HepG2-Lipo cells, and 500 microM fenofibrate inhibited both TG and cholesterol secretion from the cells.