Kazukiyo Kobayashi

Receptor-mediated formation of multilayer aggregates of primary cultured adult rat hepatocytes on lactose-substituted polystyrene.

We used a lactose-substituted polystyrene, poly-N-p-vinylbenzyl-D-lactonamide (PVLA), as a substratum for adult rat hepatocytes in primary culture. Spherical-shaped hepatocytes attached on PVLA substratum formed stable multilayer aggregates anchored on substratum through the stimulation of epidermal growth factor (EGF). The cells required calcium ion essentially to form the aggregates. The formation of multilayer aggregates was inhibited by colchicine, but not by cytochalasin B. The inhibition was also observed by added PVLA molecules in the culture medium and by treating surfaces of PVLA-coated dishes with allo A lectin. It was suggested that adult rat hepatocytes attached on PVLA substratum required the specific interaction between asialoglycoprotein receptors on the cell surface and PVLA substratum to form anchored multilayer aggregates.

Receptor-mediated regulation of differentiation and proliferation of hepatocytes by synthetic polymer model of asialoglycoprotein.

Morphology and responses of hepatocytes are investigated using an artificial asialoglycoprotein model polymer--lactose-carrying polystyrene (PVLA) as a culture substratum, especially in focusing on the effect of the surface density of the PVLA substratum. The surface density of PVLA on polystyrene dishes was determined using fluorescein-labeled PVLA as a probe under a fluorescence laser microscope. PVLA-coated surfaces were observed by scanning electron microscope and atomic force microscopies under air and water, which showed that PVLA molecules were adsorbed patchily on low density surfaces and uniformly concentrated all over the dish on high density surfaces. It is suggested from the requirement of the Ca2+ ion, inhibition of galactosyl substances, and localization of receptors that the adhesion of hepatocytes to both low and high PVLA-density surfaces is mediated by galactose-specific interactions between PVLA and asialoglycoprotein receptors. At low PVLA densities (0.07 micrograms cm-2), the hepatocytes were flat and expressed high levels of 3H-thymidine uptake and low levels of bile acid secretion. Contrastingly, at high PVLA densities (1.08 micrograms cm-2), they were round and expressed a low level of 3H-thymidine uptake and a high level of bile acid secretion. The shapes, proliferation, and differentiation of hepatocytes could be regulated by varying the densities of PVLA adsorbed to polystyrene dishes. We assume that there are two recognition mechanisms operating between PVLA and hepatocytes: (1) adhesion through highly concentrated or clustered galactose-specific interaction; and (2) responses in shape, proliferation, and differentiation by PVLA-coating densities.

Chemo-Enzymatic Synthesis of a Glycopolymer Carrying Clustered-N-ACETYL-β-lactosamine Moieties

Abstract A polyacrylamide derivative having a β-linked N-acetyllactosamine moiety, a major component of oligosaccharide chains of glycoproteins, on each repeating unit was synthesized via a chemo-enzymatic process. p-Nitrophenyl N-acetyl-β-lactosaminide was prepared by an one-step enzymatic synthesis, using a β-galactosidase as catalyst, from p-nitrophenyl N-acetyl-β-d-glucosaminide as the glycosyl acceptor and lactose as the glycosyl donor. The nitro group was reduced to an amino function, which was then allowed to react with either acryloyl chloride or acrylic acid. The resulting p-acryloylaminophenyl N-acetyl-β-lactosaminide was polymerized with azobisisobutyronitrile as initiator in dimethyl sulfoxide to give a homopolymer with a number-average molecular weight ( M n ) of 3.2 × 105. High molecular-weight polymers carrying α-lactose, N-acetyl-β-d-glucosamine, and α-d-glucopyranose were also synthesized as reference polymers in similar manners. Solution properties of these poly-p-acryloylaminophenyl der...

Investigation of Dynamic Behaviors of Asialoglycoprotein Receptors of Hepatocytes Using a Latex Probe Whose Surface Was Coated with Lactose-Carrying Polystyrene

Characteristic properties of oligosaccharide-substituted poly styrenes as a hepatocyte-specific culture substratum were investigated from the viewpoint of adsorption and recognition. X-ray photoelectron spectra sug gest that the adsorption of the polymer onto a polystyrene film occurred in near mono-molecular layer thickness of 10-25 Å in globular conformations. Distri bution and migration of asialoglycoprotein receptors on the surface of a hepato cyte was followed under a scanning electron microscope using a latex probe whose surface was coated with lactose-substituted polystyrene.

Biological and Related Polymers

Abstract In this years meetings, 131 and 75 papers were presented at the spring and fall meetings of 1990, respectively. The numbers of papers increased remarkably as compared with 77 and 61 papers presented in 1989. This review highlights five topics related to proteins, nucleic acids, and lipids.