Long–Term Maintenance of Taurocholate Uptake by Adult Rat Hepatocytes Co–Cultured With A Liver Epithelial Cell Line

Abstract

Taurocholate (TC) uptake by adult rat hepatocytes co–cultured with other rat liver epithelial cells (RLEC) was studied comparatively to hepatocytes in primary culture. Cells were cultured on Petri dishes for desired times prior to measuring their ability to transport TC. TC uptake was linear for 150 sec in both culture conditions. In hepatocytes cultured alone, the initial rate of TC uptake at an extracellular concentration of 100 μ M was 0.19 ± 0.02 nmole per min per 10⁶ cells after 48 hr of culture and decreased by 75% after 4 to 6 days. In hepatocytes co–cultured with RLEC, the rate of uptake at 48 hr (0.31 ± 0.01 nmole per min per 10⁶ cells) was significantly higher than in hepatocytes cultured alone (p < 0.01); in addition, TC uptake remained stable at an average rate of 0.17 ± 0.01 nmole per min per 10⁶ cells for up to 56 days. No detectable uptake was found in RLEC cultured alone. TC uptake exhibited both saturable (V_max = 0.30 ± 0.03 nmole per min per 10⁶ cells and Km = 42.6 ± 4.4 μM) and nonsaturable components. These kinetic parameters were similar to those previously reported in isolated hepatocytes and in short–term cultured hepatocytes. TC uptake exhibited sodium dependence and was significantly reduced when extracellular sodium was replaced by lithium and sucrose, or in the presence of 1 mM ouabain. After 18 days of co–culture, TC uptake had qualitatively the same characteristics as at 48 hr, with a saturable and a nonsaturable component. These findings strongly support the view that the sodium–dependent bile acid uptake system present in normal hepatocytes was maintained in the co–culture system as previously reported for other liver–specific functions.