The Human Hyaluronan Receptor for Endocytosis (HARE/Stabilin-2) Is a Systemic Clearance Receptor for Heparin

Abstract

Heparin (Hep)2 is the most anionic proteoglycan, due to the extensive sulfation of its glycosaminoglycan (GAG) chains, and contains many different sulfated disaccharide isomers of N-acetylgalactosamine and glucuronic acid or iduronic acid. Hep binds to many different soluble, matrix, and cell surface proteins and receptors, and has many functions, including its roles as an anti-coagulant and as a co-receptor for some growth factors (1, 2). Hep is a highly prescribed drug in surgical patients and those at risk for thromobosis. The genes and metabolic pathway for Hep biosynthesis in mast cells are understood reasonably well, and many of the biological and clinical activities of Hep have been well studied for several decades (3-6). In contrast, we know less about the catabolism of Hep and how total body homeostasis of this multifunctional proteoglycan is maintained. In particular, the mechanisms controlling systemic turnover of Hep, whether as endogenous proteoglycan or free chain drugs, are not known.Although receptors for Hep have been characterized on a variety of cell types (e.g. macrophages, vascular smooth muscle cells), none of these mediate substantial clearance of Hep (7-9). A few reports implicated a role for Kupffer cells in Hep clearance, but were not followed up (10, 11). The possible contribution of liver sinusoidal endothelial cells to Hep uptake in these primary cell preparations was not examined. Because Hep is a widely prescribed drug, it is even more important to understand the factors that control its clearance and, therefore, pharmokinetics. Animal studies of Hep clearance, except by renal function, have been difficult to perform, because Hep binds to so many soluble, cell surface or matrix proteins and become widely distributed after injection.Unfractioned Hep (3000 -30000 Da), low molecular mass Hep (300 -8000 Da), and the pentasaccharide, Fondaparinux, are the three classes of Hep drugs used to treat venous thrombosis, acute myocardial infarction, trauma, obesity, and coronary and peripheral vascular procedures; all situations wherein patients need immediate platelet anti-coagulation therapy (12). Following an intravenous bolus, unfractioned Hep has a halflife of ϳ1 h and is cleared from the circulation by the liver and kidney (13). Low molecular mass Hep and the pentasaccharide, subcutaneously injected, have half-lives of ϳ3-6 and ϳ17 h, respectively (14). Larger more structurally diverse Hep is more readily cleared than the lower molecular weight fragments. Although clinical handbooks declare that Hep is metabolized and cleared by the reticuloendothelial system, the mechanisms for Hep clearance are not known.The primary scavenger receptor for systemic turnover of HA and most types of chondroitin sulfate, but not HS, is HARE/ Stab-2, which mediates most of the total body HA turnover per day (15)(16)(17). HARE is found primarily in the sinusoidal endothelial cells of the lymph nodes, liver, and spleen (18 -21), and * This work was supported, in whole or in part, by National Institutes of Hea...

Probing Structural Selectivity of Synthetic Heparin Binding to Stabilin Protein Receptors

“…Although it is known that the size of the heparin chain plays a role in the rate of clearance in patients, the precise structural requirements for regulating clearance remain uncharacterized. Previous studies have demonstrated that a liver endothelial cell receptor, known as Stab, is primarily responsible for heparin clearance (5). By taking advantage of our recent success in synthesizing heparins (22,24), we investigated the contribution of the sulfation and the (19), and 19-mer (23) for 3 h. All cell lines were washed with Hanks' buffered saline solution followed by determination of cpm by scintillation counting and protein by the Bradford assay; mean Ϯ S.D., n ϭ 5.…”

“…The clearance of heparin is primarily mediated by liver sinusoidal endothelial cells (4). The primary receptor responsible for the systemic clearance of heparin is Stabilin-2/HARE (Stab2)(HARE, hyaluronic acid receptor for endocytosis) (5,6). Thus, the binding affinity of heparin to Stab2 likely relates to the clearance rate of heparin.…”

“…Stab1 binds to SPARC (secreted protein acidic and rich in cysteine) (12) and lactogen (13); Stab2 binds to collagen propeptides (6) in addition to glycosaminoglycans. The common ligands for both receptors are acetylated LDL (5,14), GDF-15 (15), phosphatidylserine (16 -18), and advanced glycation end products (19).…”

Pharmacology of Heparin and Related Drugs

“…Both UH and LMWHs are cleared in the liver, initially by binding to the hyaluronic acid receptor for endocytosis, also known as stabilin-2, then by endocytosis into lysosomes (Harris et al, 2008). The enzymes of lysosomal breakdown of HS, when dysfunctional, give rise to some of the group of conditions known as mucopolysaccharidoses (MPSs).…”

<em>In vivo</em> Liver Endocytosis Followed by Purification of Liver Cells by Liver Perfusion

“…The Stabilin-2/HARE receptor is the main clearance receptor that binds and internalizes heparin in SECs 18 . In our example here, we labeled heparin with a biotin tag on the carboxylate group of GlcNAc/NS.…”