DIFFERENCES IN METABOLISM OF 3 LOW-DENSITY-LIPOPROTEIN SUBFRACTIONS IN HEP G2-CELLS

The metabolism of low density lipoprotein (LDL) subfractions was investigated in the human hepatoma cell line Hep G2. By means of a density gradient ultracentrifugation method three LDL subfractions were isolated from pooled serum of normolipidemic subjects: very light LDL-1A, light LDL-1B and heavy LDL-2, differing in size, relative lipid and protein content. Cell specific association, stimulation of cholesterol esterification and inhibition of sterol synthesis were determined in parallel after incubation of Hep G2 cells with increasing amounts of LDL-protein of the three LDL subfractions. These parallel experiments were repeated four times with freshly prepared LDL subfractions. Response curves were parametrized with the function y = a square-root x, depicting the relation between the cellular metabolic event (y) and the LDL-protein (x) or LDL-cholesterol (x) levels. An analysis of covariance model was used to test differences between parameters of the three LDL subfractions. When the results of all four experiments were taken into account, the cell specific association increased more with increasing LDL-protein concentration for LDL-1A than for LDL-2 (P < 0.05). At the LDL-protein level of 80 mu-g/ml the cell specific association for LDL-2 amounted to 85.5% of that for LDL-1A. Results for LDL-1B were intermediate between those for LDL-1A and LDL-2. The corresponding cholesteryl ester formation increased more with increasing LDL-protein concentration for LDL-1A than for LDL-1B (P < 0.001), and for LDL-1B more than for LDL-2 (P < 0.001). At the LDL-protein level of 70 mu-g/ml the cholesterol ester accumulation for LDL-2 and LDL-1B was 48.4% and 70.3%, respectively, of that for LDL-1A. These differences between LDL subfractions in cholesteryl esterification were independent of the cholesterol content of the subfractions. Consistent with these findings, the [C-14]acetate incorporation to sterols decreased more with increasing LDL-protein concentration for LDL-1A and LDL-1B than for LDL-2 (P < 0.05). At the LDL-protein concentration of 70 mu-g/ml the decrease in sterol synthesis after incubation with LDL-2 and LDL-1B was 79.2% and 98.0%, respectively, relative to that for LDL-1A. When adjusted for differences in cholesterol content of the LDL subfractions these differences with regard to the C-14-acetate incorporation were not significant. The metabolic differences between LDL subfractions in vitro may have implications for the metabolism and atherogenic potential of the distinct LDL subfractions in vivo. Thus, in the study of atherogenicity of lipoproteins, the existence of subfractions of LDL should be taken into account. Even for normolipidemic subjects the subfractions appear to have different metabolic characteristics.