Increased functional expression of P-glycoprotein in Caco-2 TC7 cells exposed long-term to cadmium.
The objective of this study was to investigate whether P-glycoprotein (P-gp) functional expression in intestinal cells is modified after long-term exposure to the food contaminant cadmium (Cd). The Caco-2 cell line, clone TC7, was first validated as a cellular model for long-term exposure to cadmium. Cytotoxicity tests after acute exposure of 24 h showed a significant concentration-dependent decrease in cellular viability at cadmium levels higher than 10 microM and led us to select the cadmium ranges for long-term exposure: 1, 5, and 10 microM. Intestinal cells were exposed to these cadmium concentrations for four consecutive weeks without inducing DNA condensation or fragmentation. In the second part of this work, we studied the functional expression of the drug efflux pump multidrug resistance P-glycoprotein after long-term exposure to cadmium by immunoblotting with the monoclonal antibody F4 and measurement of calcein-AM+/-the P-gp inhibitor verapamil. Western blot analysis with the F4 antibody detected a single band of 170 to 180 kDa which is the size previously reported for P-gp. Calcein-AM assay showed that four weeks exposure of intestinal cells to 1, 5, and 10 microM Cd increased P-gp functional expression in proportion to the Cd concentration.
Huynh-Delerme
Céline
C
Laboratoire de Pharmacie-Toxicologie, Department of Pharmacy and Toxicology, Ecole Nationale Vétérinaire d'Alfort, 7 avenue du Général de Gaulle, 94704 Maisons-Alfort, France. chuynh-delerme@vet-alfort.fr
Huet
Hélène
H
Noël
Laurent
L
Frigieri
Amanda
A
Kolf-Clauw
Martine
M
D018435:ATP Binding Cassette Transporter, Subfamily B / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D001562:Benzimidazoles; D015153:Blotting, Western; D018938:Caco-2 Cells; D002105:Cadmium Poisoning / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D004247:DNA / Q000096:biosynthesis / Q000235:genetics; D018432:Drug Resistance, Multiple; D004587:Electrophoresis, Agar Gel; D005452:Fluoresceins / Q000378:metabolism; D005456:Fluorescent Dyes; D006801:Humans; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D009499:Neutral Red; D013778:Tetrazolium Salts; D013844:Thiazoles; D014700:Verapamil / Q000494:pharmacology
D018435:ATP Binding Cassette Transporter, Subfamily B; D001562:Benzimidazoles; D002121:Calcium Channel Blockers; D005452:Fluoresceins; D005456:Fluorescent Dyes; D013778:Tetrazolium Salts; D013844:Thiazoles; C085925:calcein AM; D009499:Neutral Red; D004247:DNA; D014700:Verapamil; D007770:L-Lactate Dehydrogenase; C022616:thiazolyl blue; C017807:bisbenzimide ethoxide trihydrochloride