Evidence for an intracellular barrier to cadmium transport through Caco-2 cell monolayers

Cd-109 transport was studied in the highly differentiated TC7 clone of the enterocytic-like Caco-2 cells grown on filters. Accumulation curves for 0.3 mu M Cd-109 over 12 h from the apical (AP) or the basal (BL) sides reveale d a three-step mechanism involving: 1) a zero-time accumulation A(O); 2) a fast process A(f)(t(1/2) less than or equal to 10 min); and 3) a slow proce ss of uptake A(S) (5 h less than or equal to t(1/2) less than or equal to 1 0 h) responsible for the major cellular levels of Cd-109. The relative cont ribution of adsorption to total accumulation is greater for short exposure times (less than or equal to 35%), but is no longer significant after the e xposure times needed to reach equilibrium. Transepithelial transport was le ss than 4% of the cellular level at 12 h. A negligible but specific binding onto the BL surface of the filters was characterized. Saturable systems of accumulation with comparable affinities (K-m = 2.5 +/- 0.5 and 5.4 +/- 0.4 mu M) but distinct capacities (V-max = 8.9 +/- 1.2 and 312 +/- 22 pmol/min /mg protein) were identified at the AP and BL cell membranes, respectively. Efflux studies revealed that Cd accumulation is only partially reversible, with an exclusive metal release at the same side. A 2-h exposure on both s ides simultaneously failed to demonstrate any competition for cellular accu mulation: uptake was additive relative to AP and BL uptake values. These da ta suggest that A(f) leads to an accumulation of loosely bound Cd, whereas A(S) represents irreversible intracellular binding processes. We conclude t hat Cd transport occurs exclusively by a transcellular route and that satur ation of the intracellular high-capacity binding sites is the rate-limiting step in Cd absorption. (C) 1999 Wiley-Liss, Inc.