Oude Elferink R P, Ottenhoff R, Radominska A, Hofmann A F, Kuipers F, Jansen P L
Division of Gastrointestinal and Liver Diseases, Academic Medical Centre, Amsterdam, The Netherlands.
Biochem J. 1991 Feb 15;274 ( Pt 1)(Pt 1):281-6. doi: 10.1042/bj2740281.
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.
测试了一系列有机化合物对肝细胞分泌模型有机阴离子二硝基苯基谷胱甘肽（GS-DNP）的影响。先前的实验表明，分离的大鼠肝细胞分泌GS-DNP主要由该化合物的胆小管转运系统介导。将分离的大鼠肝细胞与胆汁酸胆酸（C）、牛磺胆酸（TC）、牛磺熊去氧胆酸（TUDC）以及甘氨胆酸或牛磺胆酸（GLC或TLC）预孵育，对GS-DNP的初始外排速率没有影响。相比之下，GLC（SGLC）和TLC（STLC）的3-硫酸盐确实抑制了GS-DNP外排；10μM的SGLC可达到半数最大抑制。胆酸和石胆酸的3-O-葡萄糖醛酸苷（GlucLC）是更有效的转运抑制剂；10μM的GlucLC抑制GS-DNP转运达89%。其他亲胆有机阴离子也抑制GS-DNP分泌，尽管需要更高的浓度；100μM时，胆红素二牛磺酸盐（胆红素二葡萄糖醛酸苷的类似物）抑制转运达48%。另一方面，一些亲胆阳离子和中性化合物对GS-DNP外排没有影响。还研究了氧化型谷胱甘肽（GSSG）的肝胆分泌。在正常分离灌注的大鼠肝脏中，当还原型谷胱甘肽（GSH）在细胞内氧化时，观察到大量的GSSG胆汁分泌。GSSG也从分离的正常肝细胞中主动分泌，用100μM的SGLC孵育细胞可使这种分泌受到95%的抑制。相比之下，在遗传性结合型高胆红素血症的TR-突变大鼠的分离灌注肝脏和分离肝细胞中没有胆汁分泌。这些结果表明，GSSG和GS缀合物的胆小管外排可被多种多价有机阴离子抑制，但不能被阳离子、中性化合物和单阴离子胆汁酸抑制。这表明一种多特异性有机阴离子转运体负责这些多价阴离子的转运，这与所有这些化合物在突变的TR4大鼠中的胆汁转运存在缺陷这一事实密切相符。
