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Expression and characterization of a functional rat liver Na+ bile acid cotransport system in COS-7 cells

Boyer, J. L. and Ng, O. C. and Ananthanarayanan, M. and Hofmann, A. F. and Schteingart, C. D. and Hagenbuch, B. and Stieger, B. and Meier, P. J.. (1994) Expression and characterization of a functional rat liver Na+ bile acid cotransport system in COS-7 cells. American journal of physiology. Gastrointestinal and liver physiology, Vol. 266, H. 3 , G382-G387.

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Official URL: http://edoc.unibas.ch/dok/A5261751

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Abstract

A cDNA for the rat liver sodium-dependent bile acid cotransporter was expressed in COS-7 cells to study the functional properties of the translated protein in a mammalian cell line. A 1.2-kb insert was ligated into a pMAMneo vector and transiently transfected using electroporation. After optimal conditions were established, the transiently transfected COS cells were screened with fluorescent-conjugated labeled bile acids for evidence of expression of the cotransporter after 48 h. The uptake of [3H]taurocholate ([3H]TC) was then determined in cells transfected with or without the bile acid insert. Progressive uptake of [3H]TC (0.45 microM) was observed for 30 min in the presence of sodium. In contrast, no uptake of [3H]TC was observed in the absence of sodium, in nontransfected COS cells, or in COS cells transfected with the empty plasmid. Kinetic studies revealed a Michaelis constant (Km) of 29 microM, essentially identical to the Km of this cotransporter described in intact rat hepatocytes and membrane vesicles. Uptake of [3H]TC (5.0 microM) at 5 min (n = 3-6) was inhibited by 100 microM taurochenodeoxycholic acid (81%), tauroursodeoxycholic acid (77%), cholic acid (55%), chenodeoxycholic acid (74%), and ursodeoxycholic acid (56%) but not by 100 microM taurodehydrocholate, 1 mM probenecid, or 100 microM bilirubin. In contrast, bumetanide (500 microM) inhibited [3H]TC uptake by 52%. These studies indicate that the isolated cDNA codes for a physiological bile acid transporter present in rat hepatocytes and that posttranslational factors present in mammalian cells may not be as important in defining properties of this cotransport system.
Faculties and Departments:11 Rektorat und Verwaltung > Vizerektorat Forschung
UniBasel Contributors:Meier-Abt, Peter J.
Item Type:Article, refereed
Bibsysno:Link to catalogue
Publisher:American Physiological Society
ISSN:0002-9513
Note:Publication type according to Uni Basel Research Database: Journal article
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Last Modified:22 Mar 2012 14:21
Deposited On:22 Mar 2012 13:23

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