Evidence for an intracellular barrier to cadmium transport through Caco-2 cell monolayers.
109Cd transport was studied in the highly differentiated TC7 clone of the enterocytic-like Caco-2 cells grown on filters. Accumulation curves for 0.3 microM 109Cd over 12 h from the apical (AP) or the basal (BL) sides revealed a three-step mechanism involving: 1) a zero-time accumulation Ao; 2) a fast process Af(t1/2 < or = 10 min); and 3) a slow process of uptake As (5 h < or = t1/2 < or = 10 h) responsible for the major cellular levels of 109Cd. The relative contribution of adsorption to total accumulation is greater for short exposure times (< or = 35%), but is no longer significant after the exposure times needed to reach equilibrium. Transepithelial transport was less 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 (Km = 2.5+/-0.5 and 5.4+/-0.4 microM) but distinct capacities (Vmax = 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 sides simultaneously failed to demonstrate any competition for cellular accumulation: uptake was additive relative to AP and BL uptake values. These data suggest that Af leads to an accumulation of loosely bound Cd, whereas As represents irreversible intracellular binding processes. We conclude that Cd transport occurs exclusively by a transcellular route and that saturation of the intracellular high-capacity binding sites is the rate-limiting step in Cd absorption.
Jumarie
C
C
Department of Biological Sciences, University of Quebec at Montreal, Canada. jumarie.catherine@uqam.ca
Campbell
P G
PG
Houde
M
M
Denizeau
F
F
D000255:Adenosine Triphosphate / Q000378:metabolism; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D018938:Caco-2 Cells / Q000166:cytology / Q000201:enzymology*; D002104:Cadmium / Q000493:pharmacokinetics*; D018929:Cell Culture Techniques / Q000379:methods; D002455:Cell Division / Q000502:physiology; D002462:Cell Membrane / Q000378:metabolism; D002614:Chelating Agents / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D011485:Protein Binding / Q000502:physiology; D013393:Sucrase / Q000032:analysis
D002614:Chelating Agents; D002104:Cadmium; D000255:Adenosine Triphosphate; D004492:Edetic Acid; D013393:Sucrase