Small dense low density lipoprotein has increased affinity for LDL receptor-independent cell surface binding sites: a potential mechanism for increased atherogenicity.

Small dense low density lipoprotein (LDL) particles have altered apolipoprotein (apo) B conformation and lowered affinity for the LDL receptor (J. Biol. Chem. 1994. 269: 511-519). Herein, we examine the interaction of small dense LDL with cell LDL receptor-independent binding sites. Compared to normal LDL, at low LDL cell media concentrations (<10 microg/ml), small dense LDL had decreased specific binding to the LDL receptor on normal fibroblasts at 4 degrees C, but a 2-fold increased binding to LDL receptor-independent cell sites. At higher LDL concentration (100 microg/ ml), LDL receptor-independent binding of small dense LDL was 4.5-fold that of normal LDL in normal fibroblasts, but greater (2- to 14- fold) in LDL receptor-negative fibroblasts. In LDL receptor-negative fibroblasts at 37 degrees C, small dense LDL had higher (3-fold) cell association than normal size LDL but no effective LDL degradation. At high LDL concentrations (> or =100 microg/ml), LDL binding to normal or LDL receptor-negative fibroblasts was not affected by several anti-apoB monoclonal antibodies or by cell pretreatment with proteases, chondroitinase, or neuraminidase. In contrast, pretreating normal and receptor-negative fibroblasts with heparinase and heparitinase decreased LDL cell binding by 35% and 50%, respectively. Similarly, preincubation of receptor-negative fibroblasts with sodium chlorate, an inhibitor of proteoglycan sulfation, decreased LDL binding by about 45%. We hypothesize that small dense LDL might be more atherogenic than normal size LDL due to decreased hepatic clearance by the LDL receptor, and enhanced anchoring to LDL receptor-independent binding sites in extrahepatic tissues (e.g., the arterial wall), a process mediated, in part, by cell surface proteoglycans.

Identification and characterization of the immunoprecipitating medium- and low-density tryptic fragments of bovine nasal cartilage proteoglycan.

The fragments responsible for the immunodiffusion reactivity of middle- and low-density fractions of trypsin-digested bovine nasal cartilage proteoglycan have been identified and obtained in relatively homogeneous fractions. Glycosaminoglycan-bearing tryptic fragments were isolated from 4 M guanidinium chloride extracts of cartilage by ion-exchange chromatography and fractionated by dissociative equilibrium density gradient ultracentrifugation at a starting density of 1.50. Fragments in the middle fractions of the density gradient were digested with chondroitinase ABC and subfractionated by Sepharose 6B column chromatography. Middle-density subfractions contained fragments which were chemically and immunologically identical to those in high-density fragment subfractions of similar elution from Sepharose 6B. The middle-density subfractions contained two additional immunoprecipitating fragments. One, with alanine as N-terminal amino acid, was isolated by virtue of its retention by a column of concanavalin A-Sepharose 4B and its resistance to digestion with keratanase; the second was concentrated in a subfraction whose elution from concanavalin A-Sepharose 4B was retarded. The gradient fraction of lowest density contained fragments with the properties of the major tryptic fragments of the hyaluronic acid-binding segment of the proteoglycan monomer and the link proteins. These were recovered as a complex in the void volume upon Sepharose gel chromatography in saline-buffer and were resolved into relatively homogeneous fractions by column chromotography on CL-Sepharose 6B in 4 M guanidinium chloride. In all, tryptic digests of cartilage proteoglycan contain at least seven different immunoprecipitating fragments, some of which may not have been correctly identified previously.