Bachorik P S, Franklin F A, Virgil D G, Kwiterovich P O
Arteriosclerosis. 1985 Mar-Apr;5(2):142-52. doi: 10.1161/01.atv.5.2.142.
We examined the high affinity binding, uptake, and degradation of apo E-free 125I-high density lipoprotein (HDL) in cultured pig hepatocytes. At steady state, the cells degraded 9.4% of cell-associated 125I-HDL/hour, compared with 41.7%/hour for 125I-LDL. Pulse-chase experiments at 4 degrees C revealed that high affinity 125I-HDL binding was reversible. Similar experiments at 37 degrees C revealed that about 70% of the cell-associated 125I-HDL was released as a macromolecule; the remainder was degraded to acid-soluble products. In contrast, over 75% of the 125I-LDL that was released had been degraded to acid soluble products. The amount of macromolecular 125I-HDL released at 37 degrees C was similar to the amount that was bound to the cell surface, as estimated from measurements of trypsin-releasable radioactivity. Density gradient ultracentrifugation and SDS-polyacrylamide gel electrophoretic analysis of macromolecular 125I-HDL released to the medium revealed an increase in density, and the apparent partial proteolysis of apo A-I (Mr 25,000) to products of apparent Mr 12,000-14,000. The findings suggest that high affinity 125I-HDL uptake had a reversible component in which HDL was concentrated temporarily at the cell surface, modified, and then released as a somewhat denser lipoprotein particle. Measurement of 125I-HDL and 125I-LDL degradation in cell homogenates revealed no difference in the inherent susceptibility of the two lipoproteins to proteolysis by lysosomal enzymes. The overall slower rate of degradation of 125I-HDL compared to 125I-LDL was therefore due in part to the smaller fraction of HDL that was committed to irreversible catabolism. The rate of catabolism of this fraction, however, was considerable. Cells pulsed at 4 degrees C and subsequently warmed to 37 degrees C released one-half the acid-soluble products from 125I-HDL within about 4 hours, compared with 2 hours for cells pulsed with 125I-LDL. These findings indicate that HDL was internalized, transported to lysosomes, and degraded at about one-half the rate of LDL.
我们研究了培养的猪肝细胞中无载脂蛋白E的125I-高密度脂蛋白（HDL）的高亲和力结合、摄取和降解情况。在稳态下，细胞每小时降解细胞相关的125I-HDL的9.4%，而125I-LDL的降解率为每小时41.7%。在4℃下进行的脉冲追踪实验表明，高亲和力的125I-HDL结合是可逆的。在37℃下进行的类似实验表明，约70%的细胞相关125I-HDL以大分子形式释放；其余部分则降解为酸溶性产物。相比之下，释放的125I-LDL中超过75%已降解为酸溶性产物。根据胰蛋白酶可释放放射性的测量估计，在37℃下释放的大分子125I-HDL的量与结合到细胞表面的量相似。对释放到培养基中的大分子125I-HDL进行密度梯度超速离心和SDS-聚丙烯酰胺凝胶电泳分析，结果显示密度增加，载脂蛋白A-I（Mr 25,000）明显部分水解为表观Mr 12,000 - 14,000的产物。这些发现表明，高亲和力的125I-HDL摄取有一个可逆成分，其中HDL暂时在细胞表面浓缩，发生修饰，然后以密度稍高的脂蛋白颗粒形式释放。对细胞匀浆中125I-HDL和125I-LDL降解的测量表明，两种脂蛋白对溶酶体酶蛋白水解的内在敏感性没有差异。因此，与125I-LDL相比，125I-HDL总体降解速度较慢部分是由于HDL中致力于不可逆分解代谢的部分较小。然而，这部分的分解代谢速度相当可观。在4℃下脉冲处理后随后升温至37℃的细胞，在约4小时内从125I-HDL中释放出一半的酸溶性产物，而用125I-LDL脉冲处理的细胞则为2小时。这些发现表明，HDL被内化，转运到溶酶体，并以约为LDL一半的速度降解。
