Rifampin and digoxin induction of MDR1 expression and function in human intestinal (T84) epithelial cells.
OBJECTIVE
Oral drug bioavailability is limited by intestinal expression of P-glycoprotein (MDR1, Pgp, ABCB1) whose capacity is regulated via nuclear receptors e.g. the pregnane X receptor (PXR, SXR, NR1I2). In order to study dynamic regulation of MDR1 transport capacity we have identified the T84 epithelial cell-line as a model for human intestine co-expressing MDR1 with PXR. The ability of rifampin, a known PXR agonist and digoxin, a model MDR1 substrate, to regulate MDR1 expression and transport activity has been tested, in these T84 cells.


METHODS
Transport was assayed by bi-directional [(3)H]-digoxin transepithelial fluxes across epithelial layers of T84 cells seeded onto permeable filter supports following pre-exposure to rifampin and digoxin. Quantitative real-time PCR, Western blotting and immunocytochemistry were used to correlate induction of MDR1 transcript and protein levels with transport activity.


RESULTS
Rifampin exposure (10 microM, 72 hours) increased MDR1 transcript levels (3.4 fold), MDR1 total protein levels (4.4 fold), apical MDR1 protein (2.7 fold) and functional activity of MDR1 (1.2 fold). Pre-incubation with digoxin (1 microM, 72 hours) potently induced MDR1 transcript levels (92 fold), total protein (7 fold), apical MDR1 protein (4.7 fold) and functional activity (1.75 fold). Whereas PXR expression was increased by rifampin incubation (2 fold), digoxin reduced PXR expression (0.3 fold).


CONCLUSIONS
Chronic digoxin pre-treatment markedly upregulates MDR1 expression and secretory capacity of T84 epithelia. Digoxin-induced changes in MDR1 levels are distinct from PXR-mediated changes resulting from rifampin exposure.
Haslam
I S
IS
Epithelial Research Group, Institute for Cell and Molecular Biosciences, University of Newcastle Upon Tyne, Medical School, Newcastle Upon Tyne, UK.
Jones
K
K
Coleman
T
T
Simmons
N L
NL
D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000096:biosynthesis* / Q000235:genetics; D018528:ATP-Binding Cassette Transporters / Q000096:biosynthesis; D000199:Actins / Q000096:biosynthesis / Q000235:genetics; D000995:Antitubercular Agents / Q000494:pharmacology*; D001693:Biological Transport, Active / Q000187:drug effects; D015153:Blotting, Western; D018938:Caco-2 Cells; D002460:Cell Line; D000090702:Constitutive Androstane Receptor; D004077:Digoxin / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004847:Epithelial Cells / Q000187:drug effects*; D006801:Humans; D007150:Immunohistochemistry; D007413:Intestinal Mucosa / Q000166:cytology* / Q000187:drug effects*; D000077297:Pregnane X Receptor; D012313:RNA / Q000096:biosynthesis; D018160:Receptors, Cytoplasmic and Nuclear / Q000187:drug effects; D011987:Receptors, Steroid / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012293:Rifampin / Q000494:pharmacology*; D014157:Transcription Factors / Q000187:drug effects
C513055:ABCB1 protein, human; D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D018528:ATP-Binding Cassette Transporters; D000199:Actins; D000995:Antitubercular Agents; D000090702:Constitutive Androstane Receptor; D004791:Enzyme Inhibitors; C000625852:NR1I2 protein, human; D000077297:Pregnane X Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D011987:Receptors, Steroid; D014157:Transcription Factors; D012313:RNA; D004077:Digoxin; D012293:Rifampin