Functional expression of the P-glycoprotein mdr in primary cultures of bovine cerebral capillary endothelial cells.
The P-glycoprotein mdr is expressed not only in tumoral cells, but also in nontransformed cells, including the specialized endothelial cells of brain capillaries which build up the blood-brain barrier. Since all previously identified blood-brain barrier markers are rapidly lost when cerebral capillary endothelial cells are maintained in primary culture, we have investigated whether P-glycoprotein (P-gp) would follow the same rule, in order to address the influence of the cerebral environment on the specific P-gp expression in the brain endothelium. As compared to freshly isolated purified cerebral capillaries, P-glycoprotein was detected by immunochemistry at a high level in 5-7 day primary cultures. In our culture conditions, P-glycoprotein was immunodetected at a lower molecular weight than that found in freshly isolated capillaries. Enzymatic deglycosylation led to the same 130 kDa protein for both fresh and cultured samples, suggesting that P-gp post-translational modifications were altered in primary cultures. However, studies on the uptake and efflux of the P-gp substrate [3H]vinblastine, and on the effect of various mdr reversing agents on the uptake and efflux, clearly indicated that the efflux pump function of the P-glycoprotein was maintained in primary cultures of bovine cerebral capillary endothelial cells. P-Glycoprotein may thus represent the first blood-brain barrier marker which is maintained in cerebral endothelial cells cultured in the absence of factors originating from the brain parenchyma.
Lechardeur
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UMR 133 CNRS/Rhône-Poulenc Rorer, Centre de recherche de Vitry-Alfortville, Vitry/Seine, France.
Scherman
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D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000032:analysis / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D000818:Animals; D015415:Biomarkers; D001812:Blood-Brain Barrier / Q000502:physiology*; D002196:Capillaries / Q000166:cytology; D002417:Cattle; D002478:Cells, Cultured / Q000737:chemistry / Q000502:physiology; D002540:Cerebral Cortex / Q000098:blood supply; D004730:Endothelium, Vascular / Q000737:chemistry* / Q000201:enzymology / Q000502:physiology*; D007150:Immunohistochemistry; D014316:Tritium / Q000493:pharmacokinetics; D014747:Vinblastine / Q000493:pharmacokinetics*; D005723:gamma-Glutamyltransferase / Q000378:metabolism
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D015415:Biomarkers; D014316:Tritium; D014747:Vinblastine; D005723:gamma-Glutamyltransferase