Triacylglycerol-rich lipoprotein cholesterol is derived from the plasma membrane in CaCo-2 cells.

: The source for triacylglycerol-rich lipoprotein cholesterol was investigated in CaCo-2 cells grown on filters separating an upper and a lower well. Oleic acid, a fatty acid that promotes triacylglycerol-rich lipoprotein synthesis and secretion in CaCo-2 cells, increased the vesicular-mediated influx of plasma membrane cholesterol to the endoplasmic reticulum. Unesterified and esterified cholesterol derived from the plasma membrane were increased in triacylglycerol-rich lipoproteins secreted by cells incubated with oleic acid. Fatty acids, which increased the number of lipoprotein particles secreted (increased apoB secretion), increased plasma membrane cholesterol influx and secretion. Oleic acid caused a modest increase in the synthesis of cholesterol and a two-fold increase in cholesteryl esters. The amount of newly synthesized cholesterol secreted in lipoproteins of density < 1.006 g/ml represented a small fraction of that present within the cell; however, oleic acid did increase the amount of both newly synthesized cholesterol and cholesteryl esters in triacylglycerol-rich lipoproteins. Oleic acid did not affect the fraction of newly synthesized cholesterol trafficking to the plasma membrane. Compared to cholesterol delivered to cells in micelles, plasma membrane cholesterol was the much preferred substrate for acyl-CoA:cholesterol acyltransferase. Micellar cholesterol displaced cholesterol from the plasma membrane causing more of it to influx intracellularly for esterification and secretion. We propose that plasma membrane cholesterol is the major source for triacylglycerol-rich lipoprotein cholesterol in CaCo-2 cells. Micellar cholesterol and newly synthesized cholesterol replenish the plasma membrane cholesterol that is being used for the transport of lipids.