Evidence for receptor-mediated hepatic uptake of pullulan in rats

Kaneo, Yoshiharu; Tanaka, Tetsuro; Nakano, Takayuki; Yamaguchi, Yasunori

doi:10.1016/s0168-3659(00)00368-0

Cited by 163 publications

(81 citation statements)

References 22 publications

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“…Sugars, such as galactose, glucose, and mannose in polysaccharides direct intracellular disposition. After intravenous injection, many polysaccharides accumulate in the liver, partially due to the 3,10) In cultured rat parenchymal cells, pullulan specifically was bound and internalized and the uptake of pullulan was inhibited by asialofetuin, in contrast to dextran, suggesting that the RME via ASGPR participates in intracellular disposition of pullulan and that fluid phase endocytosis participates in that of dextran. Stereochemical configuration resulting from branching and hydroxyl group positions is a key factor in intracellular disposition.…”

Section: Discussionmentioning

confidence: 99%

Uptake of Pullulan in Cultured Rat Liver Parenchymal Cells

Tanaka

Hamano

Fujishima

et al. 2005

Biological & Pharmaceutical Bulletin

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Polysaccharides are an attractive drug carrier directed to the liver because they are distributed in the liver after intravenous injection. Pullulan is a polysaccharide consisting of three a-1,4-linked glucose molecules, polymerized by a-1,6-linkages to the terminal glucose, and has been employed as a carrier of human interferon-b and plasmid DNA to the liver. 1,2) We previously investigated the biodisposition of pullulan in rats and demonstrated that the asialoglycoprotein receptor (ASGPR) contributes to hepatic distribution.3) Furthermore, intracellular disposition of polysaccharides has been examined in rat liver parenchymal and nonparenchymal cells and studies have shown that binding and internalization to parenchymal cells was inhibited by asialofetuin, indicating that ASGPR is involved in the intracellular disposition of pullulan. 4)In this study, we examined the uptake of pullulan, including the binding process followed by internalization in cultured rat liver parenchymal cells, and discuss the possibility of pullulan as a passive or active drug carrier via receptor mediated endocytosis (RME). MATERIALS AND METHODSMaterials Pullulan (MW 58200) was supplied by Kuraray (Japan). Arabinogalactan and asialofetuin were purchased from Sigma (St. Louis, MO. U.S.A.). Fluorescein isothiocyanate was obtained from Wako Pure Chemical (Osaka, Japan). All other reagents were the highest grade commercially available.Radio-Labeling of Pullulan The tyramine derivative of pullulan was prepared according to a previous report. 4)Briefly, pullulan was dissolved in 10% acetone in water and cooled in an ice bath. Then, 10% CNBr in acetone was added and the mixture stirred for 2 min. Tetraethylamine in acetone was added to the solution. After 2 min, tyramine in 0.1 M NaHCO 3 containing 0.5 M NaCl was added and the reaction mixture stirred overnight at 4°C. The mixture was dialyzed against water and the macromolecular fraction obtained as a white powder. Animals Wistar male rats (6 weeks old) were purchased from Japan SLC, Inc. (Shizuoka, Japan). The rats were maintained on a commercially balanced stock diet (Oriental Yeast Co. Ltd., Tokyo, Japan) with water ad libitum. Before experimentation, each rat was anesthetized with pentobarbital. All aspects of the study were performed according to the Fukuyama University guidelines for animal experiments.Isolation and Culture of Rat Liver Parenchymal Cells Hepatocytes were isolated from male Wistar rats (7 weeks old) by the collagenase perfusion method.6) The hepatocytes were centrifuged at 50ϫg and the resulting pellet washed with Hanks' medium containing 2.4 mM CaCl 2 and 10 mM HEPES (pH 7.4). The cells were fractionated on Percoll density gradients, which resulted in a greater than 98% viability, as verified by a trypan blue exclusion test. The cells were then resuspended in William's Medium E containing dexamethasone (1 µM), insulin (0.1 µM), and 10% FBS. The cells (1ϫ10 6 cells) were plated in a 35-mm collagen type I coated culture dish (Iwaki, Funabashi, Japan) and incubated in hu...

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Section: Discussionmentioning

confidence: 99%

Uptake of Pullulan in Cultured Rat Liver Parenchymal Cells

Tanaka

Hamano

Fujishima

et al. 2005

Biological & Pharmaceutical Bulletin

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“…92 Mediating through interaction between polysaccharide backbone of pullulan and ASGPR, the pullulan-coated drug carrier significantly inhibited hepatoma cell proliferation and migration, as well as tumor growth and angiogenesis. 93 …”

Section: Retinoic Acid Biotinmentioning

confidence: 95%

Ligand-based targeted therapy: a novel strategy for hepatocellular carcinoma

Li¹,

Zhang²,

Wang³

et al. 2016

IJN

118

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“…24 FITC-CHSP was prepared by the method of Kaneo et al 30 Briefly, CHSP (1 g) was dissolved in dimethyl sulfoxide (DMSO; Sigma-Aldrich, St Louis, MO, USA) (10 mL) containing a few drops of pyridine. FITC (Sigma-Aldrich; 100 mg) was added, followed by dibutyltin dilaurate (SigmaAldrich; 20 mg), and the mixture was heated for 2 hours at 95°C.…”

Section: Preparation and Characterization Of Fitc-chspmentioning

confidence: 99%

Cellular uptake mechanism and intracellular fate of hydrophobically modified pullulan nanoparticles

Jiang¹,

Li²,

Liu³

et al. 2013

IJN

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Abstract:The cellular uptake mechanism and intracellular fate of self-assembled nanoparticles (NPs) of cholesterol-modified pullulan (CHSP) by human hepatocellular carcinoma (HepG2) cells were investigated. Covalent conjugation with fluorescein isothiocyanate (FITC) yielded stably labeled CHSP (FITC-CHSP), which was successfully formulated into NPs (mean particle size 63.0 ± 1.9 nm) by dialysis. A cytotoxicity assay clearly indicated that the CHSP NPs did not show significant toxicity in HepG2 cells. The effects of NP concentration, incubation time, and temperature on the cellular uptake of the NPs were systematically evaluated by fluorometry, and the results suggested that cellular uptake of the NPs was concentration-, time-, and temperature-dependent. In vitro experiments with endocytic inhibitors revealed that clathrin-mediated endocytosis and macropinocytosis were involved in the internalization of CHSP NPs. The intracellular trafficking study demonstrated that CHSP NPs were entrapped in the lysosomes at 1 hour after incubation; colocalization of NPs with either the Golgi apparatus or the endoplasmic reticula was not observed during the entire course of the study. These results suggested that the CHSP NPs may serve as a versatile carrier for intracellular delivery of therapeutic agents.

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Evidence for receptor-mediated hepatic uptake of pullulan in rats

Cited by 163 publications

References 22 publications

Uptake of Pullulan in Cultured Rat Liver Parenchymal Cells

Uptake of Pullulan in Cultured Rat Liver Parenchymal Cells

Ligand-based targeted therapy: a novel strategy for hepatocellular carcinoma

Cellular uptake mechanism and intracellular fate of hydrophobically modified pullulan nanoparticles

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