Cellular disposition of arabinogalactan in primary cultured rat hepatocytes

• Published in: European journal of pharmaceutical sciences 2004-08, Vol.22 (5), p.435-444
• Main Authors: Tanaka, Tetsuro, Fujishima, Yuko, Hamano, Shinya, Kaneo, Yoshiharu
• Format: Article
• Language: English
• Subjects: Animals; Arabinogalactan; Asialoglycoprotein receptor; Binding affinity; Biological and medical sciences; Cell Membrane - metabolism; Cells, Cultured; Dose-Response Relationship, Drug; Drug delivery system; Galactans - metabolism; General pharmacology; Hepatocytes - metabolism; Internalization; Male; Medical sciences; Pharmaceutical technology. Pharmaceutical industry; Pharmacology. Drug treatments; Protein Binding - physiology; Rats; Rats, Wistar; Receptor mediated endocytosis
• ISSN: 0928-0987; 1879-0720
• DOI: 10.1016/j.ejps.2004.04.014

To characterize a targeting property of arabinogalactan (AG) as a carrier to the liver, we examined cellular disposition, such as binding and internalization in primary cultured rat hepatocytes, comparing them to those of asialofetuin (AF). A tyramine derivative of AG was synthesized to allow labeling with 125 I . Binding of AG to the cells was concentration-dependent and saturable. The number of binding sites ( n) of AG on the cell surface was 4.0 × 10 5 ± 0.1 × 10 5 sites per cell which was about similar to that of AF. The value of K a of AG was 2.2 × 10 8 ± 0.1 × 10 8 M −1 being seven-fold higher than that of AF. The binding of AG was competitively inhibited by AF and was decreased by calcium depletion. These results indicate that AG can bind strongly to hepatocytes probably through the recognition by the asialoglycoprotein receptor (ASGP-R). Both 125 I -labeled AG and fluorescein-labeled AG were internalized into the cells. The rate of internalization of AG was faster than that of AF, indicating that AG is effectively endocytosed. Microscopic observations showed that FITC labeled AG accumulated in granules within the primary cultured rat hepatocytes. Subcellular fractionation indicated that the internalized AG was mainly associated with the lysosomal fraction. However, the internalized AG seemed to remain intact in the hepatocytes. In conclusion, we found that AG is effectively internalized in primary cultured rat hepatocytes. Although AG seems a good candidate for targeting to the liver due to its high affinity binding and rapid internalization, it remains to be established whether the apparent lack of biodegradation will result in cytotoxic effects at chronic administration in vivo.