Efflux of bis-carboxyethyl-carboxyfluorescein (BCECF) by a novel ATP-dependent transport mechanism in epithelial cells

doi:10.1016/S0006-291X(05)80203-7

Biochemical and Biophysical Research Communications

Volume 172, Issue 1, 15 October 1990, Pages 262-267

https://doi.org/10.1016/S0006-291X(05)80203-7 Get rights and content

The efflux of the intracellular pH fluorochrome 2′, 7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF) was quantified in four cultured epithelial cell lines; HCT-8, T84, HGT-1 and MDCK. BCECF efflux was time-dependent, and after 5 h 45–91% of the initial BCECF loaded was extracellular, efflux being greatest in MDCK cells. Depletion of cellular ATP approximately halved BCECF efflux. BCECF efflux was inhibited by indomethacin, vinblastine and verapamil, but not by nifedipine or reserpine. Certain features of BCECF efflux resemble drug efflux in multidrug resistant cells, but inhibition of efflux displays a distinct pharmacological profile suggesting BCECF is a substrate for a novel ATP-dependent transport system.

References (21)

ChenC. et al.
Cell
(1986)
FooteS.J. et al.
Cell
(1989)
InabaM. et al.
Biochem. Pharmacol.
(1981)
BrownC.D.A. et al.
Biochim. Biophys. Acta
(1982)
BeckW.T.
Biochem. Pharmacol.
(1987)
HigginsC.F. et al.
Nature
(1986)
HigginsC.F.
Nature
(1989)
HydeS.C. et al.
Nature
(1990)
RiordanJ.R. et al.
Science
(1989)
TsurioT. et al.
Cancer Res.
(1982)

There are more references available in the full text version of this article.

Cited by (32)

Control of Intracellular pH
2013, Seldin and Geibisch's The Kidney
Control of Intracellular pH
2012, Seldin and Giebisch's The Kidney: Physiology and Pathophysiology
Control of Intracellular pH
2008, Seldin and Giebisch's The Kidney
Control of Intracellular pH
2007, Seldin and Giebisch's The Kidney: Physiology & Pathophysiology 1-2
Drug Efflux Transport Properties of 2′,7′ -Bis(2-carboxyethyl)-5(6)-carboxyfluorescein Acetoxymethyl Ester (BCECF-AM) and Its Fluorescent Free Acid, BCECF
2004, Journal of Pharmaceutical Sciences
Citation Excerpt :
The lipophilic properties of BCECF-AM suggest it could be a substrate for P-gp and/or BCRP. However, evidence of BCECF interactions with P-gp is inconclusive, with some studies reporting a P-gp-mediated response31,33 and others showing no P-gp response.29,34 Additionally, Olson et al.34 concluded that BCECF-AM retention is not affected by MRP1 overexpression, suggesting that BCECF-AM is not a substrate for MRP1.
2′,7′-Bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF) is a fluorescent probe used to examine multidrug resistance-associated protein (MRP) transporter activity in cells. BCECF is introduced into the cell as the nonfluorescent membrane permeable acetoxymethyl ester, BCECF-AM, where it is hydrolyzed to the membrane impermeable BCECF. The lipophilic nature of BCECF-AM suggests it may be a substrate for other drug efflux transporters such as P-glycoprotein (P-gp) and the breast cancer resistance protein (BCRP). To assess the drug efflux transporter interactions of BCECF-AM and BCECF, accumulation studies were examined in various drug efflux-expressing cells. Inhibition of P-gp, BCRP, and/or MRP produced distinct changes in the time-dependent accumulation of BCECF in the cells. Treatment with GF120918 produced an immediate and sustained effect throughout the entire time course examined. Fumitremorgin C only affected BCECF accumulation at the early time points, whereas the impact of indomethacin on BCECF accumulation was observed only at the latter time points. Permeability studies in bovine brain microvessel endothelial cells indicated an increased basolateral-to-apical transport of BCECF, which could be reduced in the presence of either indomethacin or GF120918. These results indicate that the intracellular accumulation and transcellular permeability of BCECF are sensitive to a variety of drug efflux interactions. These results likely reflect an interaction of the ester form with P-gp and BCRP during the initial accumulation process, and an interaction of the free acid form with MRP after hydrolysis in the cell.
Evidence for a multidrug resistance-associated protein 1 (MRP1)-related transport system in cultured rat liver biliary epithelial cells
1999, Life Sciences
Cellular accumulation and efflux of the anionic fluorescent dye carboxy-2′,7′-dichlorofluorescein (CF) were studied in rat liver SDVI cells thought to derive from primitive bile ductules, in order to characterize carrier-related membrane transport of organic anions in epithelial cells. Probenecid, a common blocker of anion transport, was found to strongly enhance CF levels in SDVI cells in a dose-dependent manner through inhibition of dye efflux. Such an outwardly-directed transport was demonstrated to be temperature-dependent and down-regulated by various metabolic inhibitors, therefore outlining its requirement for energy; it was shown to be Na⁺- and membrane potential-independent and inhibited by anionic drugs such as indomethacin, indoprofen and rifamycin B. These functional features are closed to those described for multidrug resistance-associated protein l (MRP1) that was furthermore demonstrated, in contrast to P-glycoprotein, to be expressed in SDVI cells and to lower CF accumulation in MRP1-overexpressing drug-resistant tumor cells. These data therefore suggest that active membrane transport of organic anions such as CF occurs in epithelial cells like cultured liver biliary SDVI cells through a MRP1-related efflux system.

View all citing articles on Scopus

View full text

Efflux of bis-carboxyethyl-carboxyfluorescein (BCECF) by a novel ATP-dependent transport mechanism in epithelial cells

Cell

Cell

Biochem. Pharmacol.

Biochim. Biophys. Acta

Biochem. Pharmacol.

Nature

Nature

Nature

Science

Cancer Res.