Recovery of mRNA Expression of Tryptophan 2,3-Dioxygenase and Serine Dehydratase in Long-Term Cultures of Primary Rat Hepatocytes

• Published in: Journal of biochemistry (Tokyo) Vol. 120; no. 3; pp. 511 - 517
• Main Authors: Mizuguchi, Toru, Mitaka, Toshihiro, Hirata, Koichi, Nakamura, Toshikazu, Mochizuki, Yohichi
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 01.09.1996
• Subjects: 3-dioxygenase; Animals; Blotting, Northern; Cells, Cultured; Dexamethasone - pharmacology; dimethyl sulfoxide; Dimethyl Sulfoxide - pharmacology; Glucagon - pharmacology; Kinetics; L-Serine Dehydratase - biosynthesis; Liver - drug effects; Liver - enzymology; Male; primary rat hepatocytes; Rats; Rats, Sprague-Dawley; re-expression; RNA, Messenger - biosynthesis; serine dehydratase; Serum Albumin - biosynthesis; Time Factors; Transcription, Genetic - drug effects; tryptophan 2; Tryptophan Oxygenase - biosynthesis
• ISSN: 0021-924X
• DOI: 10.1093/oxfordjournals.jbchem.a021443

Expression of tryptophan 2,3-dioxygenase (TO) and serine dehydratase (SDH) has not previously been maintained or re-induced in long-term cultured hepatocytes. In the present study, we succeeded in inducing expression of TO and SDH mRNAs in adult rat hepatocytes cultured in serum-free L-15 medium supplemented with epidermal growth factor and 2% dimethyl sulfoxide (DMSO). After the start of culture, the expression of TO mRNA rapidly disappeared and at 96 h it was less than 10% of that at isolation. However, after the addition of 2% DMSO from 96 h, the transcript level gradually increased and reached about 40% of that of the isolated cells at day 14. In addition, the expression of TO mRNA was enhanced in cells treated with both 10-5 M dexamethasone and 10-7 M glucagon. In contrast, the expression of SDH mRNA decreased very rapidly and we could not detect it after 24 h of culture. Furthermore, 2% DMSO failed to induce it. However, when both 10-5 M dexameth asone and 10-7 M glucagon were added to the culture medium at day 9, we observed dramatic induction of SDH mRNA 24 h later. Primary hepatocytes cultured by this method could express and maintain highly differentiated hepatic functions for a long time. Thus, this in vitro system is suitable for the investigation of hepatic functions.