ENHANCED LIVER-SPECIFIC FUNCTIONS BY COCULTIVATION OF IMMORTALIZED HUMAN HEPATOCYTES AND LIVER FAT-STORING CELLS

Abstract

Purpose: Previously we have established a reversibly immortalized human hepatocyte cell line NKNT-3 to develop a bioartificial liver (BAL) (N. Kobayashi et al., Science 1252; 1246–1262, 2000). Preservation of differentiated hepatic phenotypes have an important application in BAL therapy. Hepatocyte viability and liver-specific functions have been shown to be stabilized for several weeks in vitro on cocultivation with liverfat storing cells (FSCs). Here we describe a method to expand the population of human FSCs for BAL treatment. Methods: Human fat-storing cells were transduced by a retroviral vector SSR#197 expressing the telomerase reverse transcriptase (hTERT) gene. One of immortalized FSCs with SSR#197, TWNT-1, was used in the present study based on the functional and proliferative characteristics. Differentiated liver functions were evaluated in a NKNT-3/TWNT-1 cell coculture system. Results: TWNT-1 cells showed the uptake of retinol and acetylated-LDL and synthesized collagen type 1. Cocultivation of immortalized hepatocytes NKNT-3 with TWNT-1 cells increased activities of P-450 isoenzymes 3A4 and 2C9 compared with a single culture of NKNT-3 cells. In addition, the amount of intracellular albumin and glycogen of NKNT-3 cells increased in a coculture condition. Conclusion: We have established highly differentiated immortalized human liver-fat storing TWNT-1 cells. Cocultivation of reversibly immortalized hepatocyte NKNT-3 and TWNT-1 cells could allow the development of a functional BAL.