Cholesteryl ester metabolism in tissue culture cells: II. Source of accumulated esterified cholesterol in Fu5AH rat hepatoma cells

Abstract

Previous investigations had demonstrated that Fu5AH rat hepatoma cells accumulated large quantities of esterified cholesterol when grown in hyperlipemic rabbit serum. The present investigation has determined the sources of the cellular esterified cholesterol when the cells were grown in hyperlipemic serum. Cellular esterification of endogenous and exogenous free cholesterol contributed 10% and 30%, respectively. The remaining 60% of the accumulated cellular esterfied cholesterol was derived from exogenous (serum) cholesteryl esters. Evidence for the hydrolysis of a portion of the incorporated esterified cholesterol is presented. A stimulation of free cholesterol incorporation and cellular esterification is elicited by hyperlipemic serum and serum lipoproteins when compared to normolipemic serum present at equivalent exogenous cholesterol concentrations. The effect of hyperlipemic serum is reduced by Tween-80 and Triton WR-1339. Comparative data on esterified cholesterol accumulation, free cholesterol incorporation, and cellular cholesterol esterification in Fu5-5 rat hepatoma cells, L-cells, and rabbit aortic medial cells are presented.

Cellular cholesterol ester accumulation induced by free cholesterol-rich lipid dispersions.

“…Cholesterol poor dispersions (FC/P less than one) were prepared with 40 mg of lecithin and less than 20 mg of cholesterol. Albumin (5 mg/ml) was added to the sonicated dispersions and the mixture was centrifuged at 26,000 X g for 30 (12), and the radioactivity in the separated bands was measured by liquid scintillation techniques (11,12). of FC per ml.…”

“…EC accumulation in patients with cholesteryl ester storage disease results from a block in the hydrolysis of EC (28), and a similar mechanism has been proposed in atherosclerosis (29). Alternatively, studies on both aorta (25,26) and cells in culture (12,30) have demonstrated that at least a portion of the EC accumulated in these experimental systems is derived from an increased esterification of FC. The present experiments illustrate that cholesterol-rich dispersions promote enhanced cellular FC esterification in Fu5AH cells and that this accelerated esterification leads to EC accumulation comparable to that observed when these cells are grown in hyperlipemic sera (I1, 12).…”

“…These two sets of observations appear to relate to the studies of Friedman and Byers (7)(8), and others (9,10) We sought to determine if variations in the exogenous FC/P composition could influence the accumulation of EC by tissue culture cells. The Fu5AH rat hepatoma cell line was selected because these cells are capable of accumulating large quantities of EC when grown in the presence of hyperlipemic animal sera (11), and because the accumulation of EC correlates closely with the extent of esterification of FC (12). Thus, this cell, with its high capacity for cholesterol esterification and EC accumulation, can serve as a sensitive experimental tool for the quantitation of cholesterol flux between the cells and the culture medium.…”

Source of the cholesterol ester accumulated in monkey arterial smooth muscle cells grown in hyperlipemic serum.

“…Clair et al, 1977;Rothblat et al, 1976;Mahley et al, 1977;Pearson, 1976). These results are qualitatively similar to those seen in the normal aorta when exposed to hyperlipemic serum in vitro using organ cultures (St. Clair and Harpold, 1975) or in vivo by dietary manipulations (Vesselinovitch et al, 1976;Armstrong and Megan, 1972).…”

Dietary fat saturation and hepatic acylcoenzyme A:cholesterol acyltransferase activity. Effect of n-3 polyunsaturated and long-chain saturated fat.

“…ACAT activity was assayed isotopically using [1][2][3][4][5][6][7][8][9][10][11][12][13][14] C palmitoyl CoA as the radioactive substrate. at 260 nm.…”