Unbound ligand drives hepatocyte taurocholate and BSP uptake at physiological albumin concentration

Abstract

We have recently shown (D. Sorrentino, R.B. Robinson, C.-L. Kiang, and P.D. Berk. J. Clin. Invest. 84: 1325-1333, 1989) that, in a variety of isolated cell types, the uptake of oleate at physiological albumin concentrations is consistent with traditional pharmacokinetic theory (i.e., driven by unbound ligand). Lower albumin concentrations were associated with a deviant uptake pattern for which alternative theories have been proposed. Whether other classes of organic anions exhibit similar behavior is unknown. Therefore, we examined the effect of albumin on uptake of two widely studied organic anions, sulfobromophthalein (BSP) and taurocholate. Initial uptake velocity of [35S]BSP and [3H]taurocholate by isolated hepatocytes was studied employing a fixed albumin concentration and ligand-to-albumin molar ratios from 0.01:1 to 2:1 for taurocholate and 0.031:1 to 0.75:1 for BSP. In other experiments, albumin and ligand were altered in parallel, keeping their molar ratio constant. Unbound taurocholate concentrations were measured directly by equilibrium dialysis; unbound BSP concentrations were calculated from published data (K.J. Baker and S.E. Bradley. J. Clin. Invest. 45: 281-287, 1966). At 600 microM albumin, uptake of both ligands was a function of the unbound ligand concentration. At low ligand-to-albumin molar ratios and consequent unbound ligand concentrations this relationship was linear; over the entire range of unbound ligand concentrations studied, both ligands exhibited Michaelis-Menten kinetics, with definable maximal velocity and Michaelis constant values. At low albumin concentrations, the relationships between uptake and unbound ligand were unchanged for taurocholate; however, BSP exhibited altered kinetics similar to those observed with oleate. Nontraditional uptake kinetics at low albumin concentrations appear to correlate with very high affinity for albumin.