Inhibition of glutathione-conjugate secretion from isolated hepatocytes by dipolar bile acids and other organic anions.
The effect of a spectrum of organic compounds on the secretion of a model organic anion, dinitrophenylglutathione (GS-DNP), by hepatocytes was tested. Previous experiments have demonstrated that the secretion of GS-DNP from isolated rat hepatocytes is predominantly mediated by a canalicular transport system for this compound. Preincubation of isolated rat hepatocytes with the bile acids cholic acid (C), taurocholic acid (TC), tauroursodeoxycholic acid (TUDC) and glyco- or tauro-lithocholic acid (GLC or TLC) had no effect on the initial efflux rate of GS-DNP. In contrast, the 3-sulphates of GLC (SGLC) and TLC (STLC) did inhibit GS-DNP efflux; half-maximal inhibition with SGLC was reached with 10 microM. The 3-O-glucuronides of both cholate and lithocholate (GlucLC) were even more potent inhibitors of transport; 10 microM-GlucLC inhibited GS-DNP transport by 89%. Other cholephilic organic anions also inhibited GS-DNP secretion, albeit at higher concentrations; at 100 microM, bilirubin ditaurate, an analogue of bilirubin diglucuronide, inhibited transport by 48%. On the other hand, a number of cholephilic cationic and neutral compounds had no effect on GS-DNP efflux. The hepatobiliary secretion of oxidized glutathione (GSSG) was also investigated. In normal isolated perfused rat liver, extensive biliary secretion of GSSG was observed upon intracellular oxidation of reduced glutathione (GSH). GSSG was also actively secreted from isolated normal hepatocytes, and this secretion could be inhibited by 95% by incubation of the cells with 100 microM-SGLC. In contrast, biliary secretion was absent in the isolated perfused liver and in isolated hepatocytes from TR- mutant rats with a hereditary conjugated hyperbilirubinaemia. These results show that the canalicular efflux of GSSG and GS conjugates can be inhibited by a wide variety of polyvalent organic anions, but not by cations, neutral compounds and unianionic bile acids. This suggests that a multispecific organic-anion transporter is responsible for transport of these polyvalent anions, which is in close agreement with the fact that the biliary transport of all these compounds is defective in the mutant TR4 rat.
Oude Elferink
R P
RP
Division of Gastrointestinal and Liver Diseases, Academic Medical Centre, Amsterdam, The Netherlands.
Ottenhoff
R
R
Radominska
A
A
Hofmann
A F
AF
Kuipers
F
F
Jansen
P L
PL
D000818:Animals; D001646:Bile / Q000378:metabolism*; D001647:Bile Acids and Salts / Q000494:pharmacology*; D002478:Cells, Cultured; D005965:Glucuronates / Q000494:pharmacology; D005978:Glutathione / Q000031:analogs & derivatives* / Q000378:metabolism; D007477:Ions; D007565:Jaundice / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D010042:Ouabain / Q000494:pharmacology; D010477:Perfusion; D051381:Rats; D011919:Rats, Inbred Strains; D012016:Reference Values; D013464:Sulfuric Acids / Q000494:pharmacology
D001647:Bile Acids and Salts; D005965:Glucuronates; D007477:Ions; D013464:Sulfuric Acids; C030190:S-(2,4-dinitrophenyl)glutathione; D010042:Ouabain; D005978:Glutathione