Metabolic and efflux properties of Caco-2 cells stably transfected with nuclear receptors.
OBJECTIVE
To characterise in detail the patterns of expression and functional activities of CYP and efflux pump genes in Caco-2 cells stably transfected with human Pregnane X Receptor or murine Constitutive Androstane Receptor.


METHODS
Cell lines transfected with nuclear receptors were treated with established ligands, and gene expression of CYP and efflux pump genes were quantified by qRT-PCR and Western blot. P-glycoprotein activity was assessed by measuring calcein-AM accumulation and bidirectional permeability coefficients of digoxin and quinidine. CYP activities were measured with both fluorescent and non-fluorescent substrates.


RESULTS
hPXR and mCAR upregulated some CYP and efflux pump genes ligand dependently. P-glycoprotein level was increased, but CYP3A4 protein remained below the limit of detection. P-glycoprotein activity was markedly elevated in Caco/mCAR cells and more modestly in Caco/hPXR cells. CYP3A4 activity remained lower than that in vitamin D-treated Caco-2 cells.


CONCLUSIONS
Nuclear receptors can modulate the expression of metabolic genes in Caco-2 cells, but the overall level of metabolism could not be efficiently controlled. P-glycoprotein activity increased, but CYP activities remained very low.
Korjamo
Timo
T
Department of Pharmaceutics, University of Kuopio, PO B 1627 , FIN-70211, Kuopio, Finland. Timo.Korjamo@uku.fi
Mönkkönen
Jukka
J
Uusitalo
Jouko
J
Turpeinen
Miia
M
Pelkonen
Olavi
O
Honkakoski
Paavo
P
D018435:ATP Binding Cassette Transporter, Subfamily B / Q000096:biosynthesis / Q000235:genetics; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D015153:Blotting, Western; D018938:Caco-2 Cells; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000235:genetics; D005452:Fluoresceins / Q000378:metabolism; D006801:Humans; D008024:Ligands; D010539:Permeability; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics* / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D005452:Fluoresceins; D008024:Ligands; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; C085925:calcein AM; D003577:Cytochrome P-450 Enzyme System; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human