Distribution of an asialoglycoprotein receptor on rat hepatocyte cell surface.

Abstract

Direct ferritin immunoelectron microscopy was applied to visualize the distribution of the hepatocyte cell surface of the asialoglycoprotein receptor which is responsible for the rapid clearance of serum glycoproteins and lysosomal catabolism. For this purpose, rabbit antibody against the purified hepatic binding protein specific for asialoglycoproteins was prepared and coupled to ferritin by glutaraldehyde. The specific antibody conjugates were incubated with the hepatocytes, which were isolated from rat liver homogenate after fixation by glutaraldehyde perfusion. These cells preserved well the original polygonal shape and polarity, and it was easy to identify the sinusoidal, lateral, and bile canalicular faces. The surface density of the ferritin particles bound to the sinusoidal face was about four times higher than that of particles bound to the lateral face, while the bile canalicular face was hardly labeled and almost at the control level. Using the surface area of hepatocyte measured by morphometrical analyses, it was estimated that ~90% of bound ferritin particles were at the sinusoidal face, ~10% at the lateral face, and ~1% at the bile canalicular face. Nonhepatic cells such as endothelial and Kupffer cells had no receptor specific for asialoglycoproteins.Ashwell and co-workers found that the asialoglycoproteins (ASGPs) produced by removal of the terminal sialic acid from serum glycoproteins are recognized by a hepatic receptor, rapidly removed from the circulation, and taken up by the hepatocytes to be catabolized in lysosomes (2,23).Ultrastructural analyses of the carbohydrate recognition system in rat liver have been extensively carried out by Hubbard et al. (9,10,32) by using E2~I-ASGPs and two electron microscopic tracers, asialoorosomucoid (ASOR) covalently coupled to horseradish peroxidase (ASOR-HRP) and lactosaminated ferritin. According to them, the tracers injected intravenously into rat were rapidly internalized into the hepatocytes via coated pits and coated vesicles, began to accumulate in a complex arrangement of larger smooth-surfaced vesicles, and tubular structures at the sinusoidal periphery of the cells (30 sec-2 min), appeared in Golgi-lysosome regions either in small vesicles <200 nm in diameter, larger irregular vesicles or tubules (5 min), and finally, some of these vesicles fused with lysosomes (15 min). Very recently, it has been further shown by Wall and Hubbard (33) that, at low temperature or after formaldehyde prefixation, ASGP binding sites are present over much of the sinusoidal cell surface but are concentrated heavily over coated pits. Electron microscopic observations on the endocytosis of ASGP-enzyme conjugates by hepatocytes also have been reported by Stockert et al. (30).We are interested in the distribution of the ASGP receptor on the cell surface of rat hepatocytes in situ. We asked whether the receptor exists homogeneously on all the cell surfaces or exclusively on the sinusoidal face, and, if the latter is true, whether it exists homogeneously on the s...

Expression of Asialoglycoprotein Receptor 1 in Human Hepatocellular Carcinoma

“…One important consideration for galactose-mediated targeting is that ASGPR1 expression exhibits polarity (Matsuura et al 1982) and zonality (Ise et al 2004). ASGPR1 is expressed high on the basolateral/sinusoidal membrane of rat hepatocytes and low in apical hepatocellular membrane (polarity); its expression is higher in the rat hepatocytes of centrolobular areas than in those of portalobular areas (zonality).…”

“…It plays a role in the clearance of desialylated proteins from the serum through endocytosis and lysosomal degradation. Mammalian hepatic ASGPRs mediate the binding, internalization, and degradation of extracellular glycoproteins with exposed terminal galactose, lactose or N-acetyl-galactosamine residues (Wall and Hubbard 1981;Matsuura et al 1982;Geuze et al 1982Geuze et al , 1983Spiess 1990). Natural ligands of ASGPR consist of multiple galactoses (Gal) and/or galactosamines (GalNAc), including asialoorosomucoid (ASOR, high affinity ligand of ASGPR with K i = 1.7 nM; 20 Gal), asialofetuin (17 nM; 12 Gal, 3 GalNAc), asialoceruloplasmin (86 nM; 12 Gal) and asialotransferrin (3300 nM; 5 Gal).…”

Selective Transport of Glycolipid Molecules Containing N-acetylgalactosamine to the Basolateral Domain of Mouse Hepatocyte Plasma Membrane.

“…As shown in the present results, however, glycolipids with terminal Ga1NAc were not concentrated in the lateral region of the basolateral domain but abundant in the sinusoidal region. Furthermore, asialoglycoprotein receptors are also concentrated in the sinusoidal region [10,19] . Thus, there may be an unknown mechanism to prevent the free floating of asialoglycoprotein receptors and the glycolipid molecules in the basolateral domain.…”

“…Thus, the sorting of apical proteins from basolateral proteins may occur in the basolateral domain. Asialoglycoprotein receptors, binding sites for Ga1NAc and Gal, are shown to be concentrated in the basolateral domain of the hepatocyte plasma membrane [10,19] . The glycolipid molecules with terminal Ga1NAc transported to the basolateral domain via the vesicular transport are possibly retained in the domain by binding of their terminal Ga1NAc to asialoglycoprotein receptors.…”

“…Hepatocytes maintain specialized plasma membrane domains with distinct protein and lipid compositions [19,21,23,28]. Carbohydrate-containing macromolecules such as glycoproteins and glycolipids in the plasma membrane play an important role in cell to cell interaction [1,6].…”