Shimada A, Tamai T, Oida K, Takahashi S, Suzuki J, Nakai T, Miyabo S
Third Department of Internal Medicine, Fukui Medical School, Japan.
Biochim Biophys Acta. 1994 Nov 17;1215(1-2):126-32. doi: 10.1016/0005-2760(94)90101-5.
The metabolism of high-density lipoprotein-associated cholesteryl esters (HDL-CE) in liver cells is not well understood. We studied the possible role of lysosomal and extralysosomal pathways on such metabolism by measuring the uptake and hydrolysis of HDL-CE in H-35 rat hepatoma cells. Incubation of cells with [3H]cholesteryl ester-labeled HDL led to the intracellular accumulation of both 3H-free cholesterol and [3H]cholesteryl ester. The ratio of 3H-free cholesterol/[3H]cholesteryl ester increased with an increase in incubation time even in the presence of chloroquine. Because chloroquine did not inhibit the conversion of cholesteryl ester to free cholesterol, the hydrolysis of HDL-CE may have been catalyzed by an extralysosomal enzyme, perhaps by neutral cholesteryl ester hydrolase (NCEH). When we incubated cells with increasing concentrations of HDL, NCEH activity increased. This increase in enzyme activity was not inhibited by the addition of chloroquine. A complex of dimyristoylphosphatidylcholine (DMPC)/apo HDL/cholesteryl ester enhanced the activity as well as native HDL. Neither the DMPC/apo HDL nor the DMPC/cholesteryl ester complex affected the activity, suggesting that apo HDL may be required for the uptake of HDL-CE. The present study demonstrated that the extralysosomal hydrolysis by NCEH is operating in the metabolism of HDL-CE in hepatoma cells.
肝细胞中高密度脂蛋白相关胆固醇酯（HDL-CE）的代谢尚未完全明确。我们通过测量H-35大鼠肝癌细胞中HDL-CE的摄取和水解，研究了溶酶体和溶酶体外途径在这种代谢中的可能作用。用[3H]胆固醇酯标记的HDL孵育细胞导致细胞内游离3H胆固醇和[3H]胆固醇酯均积累。即使在存在氯喹的情况下，游离3H胆固醇/[3H]胆固醇酯的比例也随着孵育时间的增加而增加。由于氯喹不抑制胆固醇酯向游离胆固醇的转化，HDL-CE的水解可能由溶酶体外酶催化，可能是由中性胆固醇酯水解酶（NCEH）催化。当我们用浓度不断增加的HDL孵育细胞时，NCEH活性增加。这种酶活性的增加不受氯喹添加的抑制。二肉豆蔻酰磷脂酰胆碱（DMPC）/载脂蛋白HDL/胆固醇酯复合物以及天然HDL均增强了活性。DMPC/载脂蛋白HDL复合物和DMPC/胆固醇酯复合物均不影响活性，这表明载脂蛋白HDL可能是HDL-CE摄取所必需的。本研究表明，NCEH介导的溶酶体外水解作用参与了肝癌细胞中HDL-CE的代谢。
