title: High-affinity uptake and degradation of apolipoprotein E free high-density lipoprotein and low-density lipoprotein in cultured porcine hepatocytes.
abstract: Isolated pig liver membranes contain a specific "lipoprotein binding site" that recognizes low-density lipoproteins (LDL) and apolipoprotein E (apoE) free high-density lipoprotein (HDL) [Bachorik, P. S., Kwiterovich, P. O., & Cooke, J. (1978) Biochemistry 17, 5287-5299]. We report here that a similar site exists in cultured porcine hepatocytes and that it mediates the uptake and degradation of apoE-free HDL. The binding of 125I-labeled HDL and 125I-labeled LDL (125I-HDL and 125I-LDL, respectively) at 4 degrees C and the uptake and degradation of the lipoproteins at 37 degrees C were time dependent and saturable and were not inhibited by unrelated proteins. Chloroquine (6 x 10(-5)M) inhibited the degradation of 125I-HDL by 76% and of 125I-LDL by greater than 99%; leupeptin inhibited the degradation of both lipoproteins by about 25%. 125I-HDL binding (4 degrees C), uptake, and degradation (37 degrees C) were inhibited by LDL, methyl-LDL, and methyl-HDL about as well as by unlabeled HDL but were unaltered in Pronase-treated cells or in cells that were cultured for 24 h in either lipoprotein-free medium containing HDL or LDL (200 micrograms/mL). In contrast, these conditions affected the uptake and degradation of 125I-LDL disproportionately. HDL and methyl-LDL inhibited 125I-LDL uptake by 50% or more but had little effect on degradation. 125I-LDL binding was reduced by 12% and degradation by 57% in Pronase-treated cells. Preincubation of the cells with LDL (200 micrograms/mL) reduced uptake by 35% and degradation by 68%. Similar preincubation with HDL (200 micrograms/mL) increased 125I-LDL degradation by 60% but did not affect 125I-LDL uptake. The findings indicated the presence in porcine hepatocytes of at least two distinct sites for lipoproteins. One site resembled the LDL receptor and mediated 125I-LDL degradation. A second, Pronase-insensitive site recognized both HDL and LDL. This site mediated almost all of the degradation of 125I-HDL but little if any degradation of 125I-LDL.
mesh_terms: D000818:Animals; D001053:Apolipoproteins / Q000378:metabolism*; D001057:Apolipoproteins E; D002478:Cells, Cultured; D002738:Chloroquine / Q000494:pharmacology; D003470:Culture Media; D007976:Leupeptins / Q000494:pharmacology; D008075:Lipoproteins, HDL / Q000378:metabolism*; D008077:Lipoproteins, LDL / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D011402:Pronase / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000378:metabolism; D011973:Receptors, LDL; D013552:Swine
chemical_list: D001053:Apolipoproteins; D001057:Apolipoproteins E; D003470:Culture Media; D007976:Leupeptins; D008075:Lipoproteins, HDL; D008077:Lipoproteins, LDL; D011956:Receptors, Cell Surface; D011973:Receptors, LDL; D002738:Chloroquine; D011402:Pronase