Dexamethasone regulation of P-glycoprotein activity in an immortalized rat brain endothelial cell line, GPNT.
The blood-brain barrier (BBB) plays an important role in controlling the passage of molecules from the blood to the extracellular fluid environment of the brain. The multidrug efflux pump P-glycoprotein (P-gp) is highly expressed in the luminal membrane of brain capillary endothelial cells, thus forming a functional barrier to lipid-soluble drugs, notably, antitumor agents. It is of interest to develop an in vitro BBB model that stably expresses P-gp to investigate the mechanisms of regulation in expression and activity. The rat brain endothelial cell line, GPNT, was derived from a previously characterized rat brain endothelial cell line. A strong expression of P-gp was found in GPNT monocultures, whereas the multidrug resistance-associated pump Mrp1 was not expressed. The transendothelial permeability coefficient of the P-gp substrate vincristine across GPNT monolayers was close to the permeability coefficient of bovine brain endothelial cells cocultured with astrocytes, a previously documented in vitro BBB model. Furthermore, the P-gp blocker cyclosporin A induced a large increase in apical to basal permeability of vincristine. Thus, P-gp is highly functional in GPNT cells. A 1-h treatment of GPNT cells with dexamethasone resulted in decreased uptake of vincristine without any increase in P-gp expression. This effect could be mimicked by protein kinase C (PKC) activation and prevented by PKC inhibition, strongly suggesting that activation of P-gp function may involve a PKC-dependent pathway. These results document the GPNT cell line as a valuable in vitro model for studying drug transport and P-gp function at the BBB and suggest that activation of P-gp activity at the BBB might be considered in chemotherapeutic treatment of cancer patients.
Régina
A
A
INSERM U. 26, Hôpital Fernand Widal, Paris, France.
Romero
I A
IA
Greenwood
J
J
Adamson
P
P
Bourre
J M
JM
Couraud
P O
PO
Roux
F
F
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D001253:Astrocytes / Q000378:metabolism; D001812:Blood-Brain Barrier / Q000502:physiology; D001921:Brain / Q000098:blood supply*; D002417:Cattle; D002461:Cell Line, Transformed; D002463:Cell Membrane Permeability; D018920:Coculture Techniques; D003078:Colchicine / Q000378:metabolism; D016572:Cyclosporine / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000032:analysis / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000378:metabolism*; D005938:Glucocorticoids / Q000494:pharmacology*; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D000074081:MutS Homolog 3 Protein; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014750:Vincristine / Q000378:metabolism
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D003078:Colchicine; D016572:Cyclosporine; D004268:DNA-Binding Proteins; D003907:Dexamethasone; D005938:Glucocorticoids; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D000074081:MutS Homolog 3 Protein; D011493:Protein Kinase C; D014750:Vincristine; C078444:MSH3 protein, human; C421109:multidrug resistance-associated protein 1