Caco-2 Cell Line Used as an In Vitro Model to Study Cadmium Accumulation in Intestinal Epithelial Cells

Abstract

109Cd uptake was studied using the highly differentiated TC7 clone of Caco-2 cells as a model of human enterocyte function. Intracellular accumulation of 0.3 microM 109Cd involved a rapid and a slow uptake phase, which resulted in complete equilibration (t(1/2) = 17.3 +/- 1.3 min) with an apparent in-to-out distribution ratio (alphae) of 11.6 +/- 0.8. The amplitude of the rapid phase (U0) and the rate of the slow phase (V) were similarly reduced in the less differentiated PF11 clone, but comparable alphae values were observed at equilibrium. In both clones, the t(1/2) and alphae values increased and decreased, respectively, upon addition of unlabeled Cd to the uptake media. In TC7 cells, 109Cd uptake at 1 min (U1) was unaffected by Ca concentrations four order of magnitude in excess, but both U0 and V demonstrated similar sensitivities to unlabeled Cd, Zn and sulfhydryl-reactive agents. Only U0 disappeared when EDTA was present in the wash solutions. U1 showed saturation kinetics and the data were found compatible with a model assuming rapid initial Cd binding and transport through a unique transport protein (Km = 3.8 +/- 0.7 microM). Cd efflux kinetics demonstrated partial reversibility in EDTA-containing solutions, suggesting that the taken up Cd might be both tightly and loosely bound to intracellular binding sites. However, the displacement of 109Cd measured at 65 min failed to reveal this heterogeneity: the data were found compatible with a model equation assuming the presence of one class of high-capacity high-affinity binding sites. We conclude that a slow-transport fast-intracellular binding mechanism of Cd uptake best accounts for these results and that Cd transport most likely involves a carrier-type of protein unrelated to Ca absorption.

Coupling the use of computer chemical speciation models and culture techniques in laboratory investigations of trace metal toxicity

“…However, we often neglect to consider that metal uptake is generally partially reversible, and that efflux may occur very rapidly, being significant within a few minutes. Indeed, in human intestinal cells Caco-2 Cd efflux was observed to proceed rapidly (Jumarie et al, 1997), this efflux not being exclusively related to desorption (Jumarie et al, 1999). This result demonstrates that metal taken up into the cells may be rapidly lost during the washing procedure, especially in the presence of EDTA/EGTA used as a chelator.…”

“…For example, in Cd uptake studies performed on human intestinal cells, Caco-2, we used a serum-free minimal uptake medium containing NaCl, KCl, CaCl 2 , MgSO 4 and D-glucose, plus HEPES as buffer (Jumarie et al, 1997). Note that the cells should be thoroughly washed before being exposed to the metal, in order to remove the remaining serum and to ensure that uptake / toxicity experiments are performed under well-controlled conditions.…”

Identification of mouse SLC39A8 as the transporter responsible for cadmium-induced toxicity in the testis

“…Cd uptake was performed by control cell lines with 109 Cd [3.64 mCi͞mg (1 Ci ϭ 37 GBq) in 0.5 M HCl; NEZ058; PerkinElmer] as described in ref. 36.…”

Effect of DMT1 knockdown on iron, cadmium, and lead uptake in Caco-2 cells

“…When confluent, the cells differentiate to form a distinct apical and basolateral surface, the former of which is enriched in DMT1 (15,30), along with other markers of differentiation (9,19). These cells have become useful tools for the study of uptake and transport of nutrients (21,24), including iron (30) and cadmium (17). The involvement of DMT1 in metal transport has been, until now, based on overexpression of DMT1 by transfection (32) and control of expression using iron treatment (30).…”