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Biochemical Journal logoLink to Biochemical Journal
. 1996 Jun 15;316(Pt 3):901–904. doi: 10.1042/bj3160901

Effect of antisense oligonucleotides on the expression of hepatocellular bile acid and organic anion uptake systems in Xenopus laevis oocytes.

B Hagenbuch 1, B F Scharschmidt 1, P J Meier 1
PMCID: PMC1217435  PMID: 8670169

Abstract

A Na(+)-dependent bile acid (Na+/taurocholate co-transporting polypeptide; Ntcp) and a Na(+)-independent bromosulphophthalein (BSP)/bile acid uptake system (organic-anion-transporting polypeptide; oatp) have been cloned from rat liver by using functional expression cloning in Xenopus laevis oocytes. To evaluate the extent to which these cloned transporters could account for overall hepatic bile acid and BSP uptake, we used antisense oligonucleotides to inhibit the expression of Ntcp and oatp in Xenopus laevis oocytes injected with total rat liver mRNA. An Ntcp-specific antisense oligonucleotide co-injected with total rat liver mRNA blocked the expression of Na(+)-dependent taurocholate uptake by approx. 95%. In contrast, an oatp-specific antisense oligonucleotide when co-injected with total rat liver mRNA had no effect on the expression of Na(+)-dependent taurocholate uptake, but it blocked Na(+)-independent uptake of taurocholate by approx. 80% and of BSP by 50%. Assuming similar expression of hepatocellular bile acid and organic anion transporters in Xenopus laevis oocytes, these results indicate that Ntcp and oatp respectively represent the major, if not the only, Na(+)-dependent and Na(+)-independent taurocholate uptake systems in rat liver. By contrast, the cloned oatp accounts for only half of BSP transport, suggesting that there must be additional, non-bile acid transporting organic anion uptake systems in rat liver.

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Selected References

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  1. Akagi H., Patton D. E., Miledi R. Discrimination of heterogenous mRNAs encoding strychnine-sensitive glycine receptors in Xenopus oocytes by antisense oligonucleotides. Proc Natl Acad Sci U S A. 1989 Oct;86(20):8103–8107. doi: 10.1073/pnas.86.20.8103. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Ananthanarayanan M., Bucuvalas J. C., Shneider B. L., Sippel C. J., Suchy F. J. An ontogenically regulated 48-kDa protein is a component of the Na(+)-bile acid cotransporter of rat liver. Am J Physiol. 1991 Nov;261(5 Pt 1):G810–G817. doi: 10.1152/ajpgi.1991.261.5.G810. [DOI] [PubMed] [Google Scholar]
  3. Ananthanarayanan M., von Dippe P., Levy D. Identification of the hepatocyte Na+-dependent bile acid transport protein using monoclonal antibodies. J Biol Chem. 1988 Jun 15;263(17):8338–8343. [PubMed] [Google Scholar]
  4. Anwer M. S., Hegner D. Effect of Na on bile acid uptake by isolated rat hepatocytes. Evidence for a heterogeneous system. Hoppe Seylers Z Physiol Chem. 1978 Feb;359(2):181–192. [PubMed] [Google Scholar]
  5. Berk P. D., Potter B. J., Stremmel W. Role of plasma membrane ligand-binding proteins in the hepatocellular uptake of albumin-bound organic anions. Hepatology. 1987 Jan-Feb;7(1):165–176. doi: 10.1002/hep.1840070131. [DOI] [PubMed] [Google Scholar]
  6. Bossuyt X., Müller M., Hagenbuch B., Meier P. J. Polyspecific drug and steroid clearance by an organic anion transporter of mammalian liver. J Pharmacol Exp Ther. 1996 Mar;276(3):891–896. [PubMed] [Google Scholar]
  7. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  8. Frimmer M., Ziegler K. The transport of bile acids in liver cells. Biochim Biophys Acta. 1988 Feb 24;947(1):75–99. doi: 10.1016/0304-4157(88)90020-2. [DOI] [PubMed] [Google Scholar]
  9. García-Ruiz C., Morales A., Colell A., Rodés J., Yi J. R., Kaplowitz N., Fernández-Checa J. C. Evidence that the rat hepatic mitochondrial carrier is distinct from the sinusoidal and canalicular transporters for reduced glutathione. Expression studies in Xenopus laevis oocytes. J Biol Chem. 1995 Jul 7;270(27):15946–15949. doi: 10.1074/jbc.270.27.15946. [DOI] [PubMed] [Google Scholar]
  10. Hagenbuch B., Lübbert H., Stieger B., Meier P. J. Expression of the hepatocyte Na+/bile acid cotransporter in Xenopus laevis oocytes. J Biol Chem. 1990 Apr 5;265(10):5357–5360. [PubMed] [Google Scholar]
  11. Hagenbuch B., Stieger B., Foguet M., Lübbert H., Meier P. J. Functional expression cloning and characterization of the hepatocyte Na+/bile acid cotransport system. Proc Natl Acad Sci U S A. 1991 Dec 1;88(23):10629–10633. doi: 10.1073/pnas.88.23.10629. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Inoue M., Kinne R., Tran T., Arias I. M. Taurocholate transport by rat liver sinusoidal membrane vesicles: evidence of sodium cotransport. Hepatology. 1982 Sep-Oct;2(5):572–579. doi: 10.1002/hep.1840020510. [DOI] [PubMed] [Google Scholar]
  13. Jacquemin E., Hagenbuch B., Stieger B., Wolkoff A. W., Meier P. J. Expression cloning of a rat liver Na(+)-independent organic anion transporter. Proc Natl Acad Sci U S A. 1994 Jan 4;91(1):133–137. doi: 10.1073/pnas.91.1.133. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kramer W., Bickel U., Buscher H. P., Gerok W., Kurz G. Bile-salt-binding polypeptides in plasma membranes of hepatocytes revealed by photoaffinity labelling. Eur J Biochem. 1982 Dec;129(1):13–24. doi: 10.1111/j.1432-1033.1982.tb07015.x. [DOI] [PubMed] [Google Scholar]
  15. Kullak-Ublick G. A., Hagenbuch B., Stieger B., Wolkoff A. W., Meier P. J. Functional characterization of the basolateral rat liver organic anion transporting polypeptide. Hepatology. 1994 Aug;20(2):411–416. [PubMed] [Google Scholar]
  16. Kurisu H., Nilprabhassorn P., Wolkoff A. W. Preparation of [35S]sulfobromophthalein of high specific activity. Anal Biochem. 1989 May 15;179(1):72–74. doi: 10.1016/0003-2697(89)90202-9. [DOI] [PubMed] [Google Scholar]
  17. Liang D., Hagenbuch B., Stieger B., Meier P. J. Parallel decrease of Na(+)-taurocholate cotransport and its encoding mRNA in primary cultures of rat hepatocytes. Hepatology. 1993 Nov;18(5):1162–1166. [PubMed] [Google Scholar]
  18. Lotan I., Goelet P., Gigi A., Dascal N. Specific block of calcium channel expression by a fragment of dihydropyridine receptor cDNA. Science. 1989 Feb 3;243(4891):666–669. doi: 10.1126/science.2464853. [DOI] [PubMed] [Google Scholar]
  19. Lotan I., Volterra A., Dash P., Siegelbaum S. A., Goelet P. Blockade of ion channel expression in Xenopus oocytes with complementary DNA probes to Na+ and K+ channel mRNAs. Neuron. 1988 Dec;1(10):963–971. doi: 10.1016/0896-6273(88)90153-5. [DOI] [PubMed] [Google Scholar]
  20. Magagnin S., Bertran J., Werner A., Markovich D., Biber J., Palacín M., Murer H. Poly(A)+ RNA from rabbit intestinal mucosa induces b0,+ and y+ amino acid transport activities in Xenopus laevis oocytes. J Biol Chem. 1992 Aug 5;267(22):15384–15390. [PubMed] [Google Scholar]
  21. Markovich D., Bissig M., Sorribas V., Hagenbuch B., Meier P. J., Murer H. Expression of rat renal sulfate transport systems in Xenopus laevis oocytes. Functional characterization and molecular identification. J Biol Chem. 1994 Jan 28;269(4):3022–3026. [PubMed] [Google Scholar]
  22. Meier P. J. Molecular mechanisms of hepatic bile salt transport from sinusoidal blood into bile. Am J Physiol. 1995 Dec;269(6 Pt 1):G801–G812. doi: 10.1152/ajpgi.1995.269.6.G801. [DOI] [PubMed] [Google Scholar]
  23. Nathanson M. H., Boyer J. L. Mechanisms and regulation of bile secretion. Hepatology. 1991 Sep;14(3):551–566. [PubMed] [Google Scholar]
  24. Petzinger E. Transport of organic anions in the liver. An update on bile acid, fatty acid, monocarboxylate, anionic amino acid, cholephilic organic anion, and anionic drug transport. Rev Physiol Biochem Pharmacol. 1994;123:47–211. doi: 10.1007/BFb0030903. [DOI] [PubMed] [Google Scholar]
  25. Reichen J., Paumgartner G. Uptake of bile acids by perfused rat liver. Am J Physiol. 1976 Sep;231(3):734–742. doi: 10.1152/ajplegacy.1976.231.3.734. [DOI] [PubMed] [Google Scholar]
  26. Schwarz L. R., Burr R., Schwenk M., Pfaff E., Greim H. Uptake of taurocholic acid into isolated rat-liver cells. Eur J Biochem. 1975 Jul 15;55(3):617–623. doi: 10.1111/j.1432-1033.1975.tb02199.x. [DOI] [PubMed] [Google Scholar]
  27. Stieger B., Hagenbuch B., Landmann L., Höchli M., Schroeder A., Meier P. J. In situ localization of the hepatocytic Na+/Taurocholate cotransporting polypeptide in rat liver. Gastroenterology. 1994 Dec;107(6):1781–1787. doi: 10.1016/0016-5085(94)90821-4. [DOI] [PubMed] [Google Scholar]
  28. Tiribelli C., Lunazzi G. C., Sottocasa G. L. Biochemical and molecular aspects of the hepatic uptake of organic anions. Biochim Biophys Acta. 1990 Oct 8;1031(3):261–275. doi: 10.1016/0304-4157(90)90012-2. [DOI] [PubMed] [Google Scholar]
  29. Van Dyke R. W., Stephens J. E., Scharschmidt B. F. Bile acid transport in cultured rat hepatocytes. Am J Physiol. 1982 Dec;243(6):G484–G492. doi: 10.1152/ajpgi.1982.243.6.G484. [DOI] [PubMed] [Google Scholar]
  30. Von Dippe P., Amoui M., Alves C., Levy D. Na(+)-dependent bile acid transport by hepatocytes is mediated by a protein similar to microsomal epoxide hydrolase. Am J Physiol. 1993 Mar;264(3 Pt 1):G528–G534. doi: 10.1152/ajpgi.1993.264.3.G528. [DOI] [PubMed] [Google Scholar]
  31. Wieland T., Nassal M., Kramer W., Fricker G., Bickel U., Kurz G. Identity of hepatic membrane transport systems for bile salts, phalloidin, and antamanide by photoaffinity labeling. Proc Natl Acad Sci U S A. 1984 Aug;81(16):5232–5236. doi: 10.1073/pnas.81.16.5232. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Wolkoff A. W., Chung C. T. Identification, purification, and partial characterization of an organic anion binding protein from rat liver cell plasma membrane. J Clin Invest. 1980 May;65(5):1152–1161. doi: 10.1172/JCI109770. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Zimmerli B., Valantinas J., Meier P. J. Multispecificity of Na+-dependent taurocholate uptake in basolateral (sinusoidal) rat liver plasma membrane vesicles. J Pharmacol Exp Ther. 1989 Jul;250(1):301–308. [PubMed] [Google Scholar]
  34. von Dippe P., Levy D. Characterization of the bile acid transport system in normal and transformed hepatocytes. Photoaffinity labeling of the taurocholate carrier protein. J Biol Chem. 1983 Jul 25;258(14):8896–8901. [PubMed] [Google Scholar]

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