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Fan J, Liu S, Du Y, Morrison J, Shipman R, Pang KS: Up-regulation of transporters and enzymes by the vitamin D receptor ligands, 1alpha,25-dihydroxyvitamin D3 and vitamin D analogs, in the Caco-2 cell monolayer. J Pharmacol Exp Ther. 2009 Aug;330(2):389-402. Epub 2009 May 4.
The effects of 1alpha,25-dihydroxyvitamin D (3) [1,25 (OH)(2) D (3)] on gene expression and function were studied in Caco-2 cells. Microarray analyses, real-time quantitative polymerase chain reactions, and Western blotting were used to determine the mRNA and protein expression of transporters and enzymes after 1,25 (OH)(2) D (3) or vehicle (0.1% ethanol) treatment for 1, 3, 6, and 10 days. The mRNA and protein expressions of the apical sodium-dependent bile acid transporter, oligopeptide transporter 1, multidrug resistance-associated protein (MRP) 3, and sulfotransferase 1E1 remained unchanged with 1,25 (OH)(2) D (3) treatment, whereas those for CYP3A4, multidrug resistance protein 1, and MRP2 were significantly increased (P < 0.05). 1,25 (OH)(2) D (3) treatment significantly enhanced MRP4 protein expression by increasing protein stability without affecting mRNA expression, as confirmed in cycloheximide experiments. Marked increase in 6beta-hydroxylation of testosterone by CYP3A4 was also observed in the 6-day 1,25 (OH)(2) D (3)-treated (100 nM) cell lysate. The transport of [(3) H] digoxin, the P-glycoprotein (P-gp) substrate, after treatment with 100 nM 1,25 (OH)(2) D (3) for 3 days revealed a higher apparent permeability (P (app)) value in the basal (B)-to-apical (A) direction over that of vehicle treatment (15.1 +/- 0.53 x 10 (-6) versus 11.8 +/- 0.58 x 10 (-6) cm/s; P < 0.05), whereas the P (app) in the A-to-B direction was unchanged; the efflux ratio was increased (from 5.8 to 8.0). Reduced cellular retention of 5-(and-6)-carboxy-2',7'-dichlorofluorescein, suggestive of higher MRP2 activity, was observed in the 3-day 100 nM 1,25 (OH)(2) D (3)-treated cells over controls. Higher protein expression of CYP3A4, MRP2, P-gp, and MRP4 was also observed after a 6-day treatment with other vitamin D analogs (100 nM 1alpha-hydroxyvitamin D (3),1alpha-hydroxyvitamin D (2) or Hectorol, and 25-hydroxyvitamin D (3)) in Caco-2 cells, suggesting a role of 1,25 (OH)(2) D (3) and analogs in the activation of enzymes and transporters via the vitamin D receptor. |
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