RECONSTITUTION OF BILE-ACID TRANSPORT IN THE RAT HEPATOMA MCARDLE RH-7777 CELL-LINE

The Liver recovers bile acids from the portal circulation primarily vi a an active process that is dependent on sodium ions, Hepatocytes lose the ability to transport bile acids in culture, and, in liver-derived permanent cell lines, this ability is severely reduced or absent, To study the importance of bile acids in regulating Liver-specific functi ons (e.g., cellular bile acid and cholesterol metabolism), we have re- established active bile acid transport in cultured cells, The compleme ntary DNA (cDNA) encoding the rat sodium/taurocholate cotransporting p olypeptide (ntcp) was placed under the control of a cytomegalovirus pr omoter and transfected into the rat hepatoma cell line, McArdle RH-777 7. Transfected cells were screened for the ability to take up [H-3]-ta urocholate, Clones that displayed the ability to take up taurocholate were expanded (designated McNtcp) and further characterized, The appar ent Michaelis constant (K-m) for taurocholate uptake was similar among the different clones, The observed maximum velocity (V-max), however, differed and was positively correlated with the abundance of recombin ant ntcp messenger RNA (mRNA), The highest level of taurocholate uptak e activity observed in McNtcp cells was comparable with that of freshl y isolated hepatocytes, Efflux of accumulated taurocholate from McNtcp cells proceeded in a manner similar to primary hepatocytes, indicatin g that McArdle RH-7777 cells have retained the ability to secrete bile acids, Moreover, taurocholate uptake in McNtcp cells was inhibited by other bile acid species, Based on the observed kinetic parameters, th e reconstituted McArdle RH-7777 cells mimic the ability of primary hep atocytes to transport bile acids.