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. 2000 May 15;348(Pt 1):183–188.

Electron-microscopic demonstration of multidrug resistance protein 2 (Mrp2) retrieval from the canalicular membrane in response to hyperosmolarity and lipopolysaccharide.

F Dombrowski 1, R Kubitz 1, A Chittattu 1, M Wettstein 1, N Saha 1, D Häussinger 1
PMCID: PMC1221052  PMID: 10794730

Abstract

Immunohistochemical studies suggest that canalicular secretion via multidrug resistance protein 2 (Mrp2), a conjugate export pump encoded by the Mrp2 gene, is regulated by rapid transporter retrieval from/insertion into the canalicular membrane. The present study was undertaken in order to investigate this suggestion by means of immunogold electron microscopy. Therefore the effects of lipopolysaccharide (LPS) and osmolarity on Mrp2 localization were studied following immunogold labelling in the perfused rat liver by quantitative electron microscopy and morphometric analyses, and by confocal laser scanning microscopy. Mrp2 activity was assessed in the isolated perfused rat liver by measuring the excretion of dinitrophenyl-S-glutathione as a substrate of Mrp2. Both LPS and hyperosmolarity resulted in a statistically significant decrease in immunogold-labelled Mrp2 in the canalicular membrane and canalicular villi, and an increase in labelling in the pericanalicular cytoplasm. Canalicular morphometric parameters were unchanged under these conditions compared with controls. Under hyperosmolar perfusion Mrp2, but not the canalicular protein dipeptidylpeptidase IV, was found inside the cells, as shown by double immunofluorescence and confocal laser scanning microscopy. The findings suggest a selective retrieval of Mrp2 from the canalicular membrane under the influence of hyperosmolarity and LPS, whereas canalicular morphology remains unchanged.

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Selected References

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  1. Akerboom T. P., Sies H. Transport of glutathione, glutathione disulfide, and glutathione conjugates across the hepatocyte plasma membrane. Methods Enzymol. 1989;173:523–534. doi: 10.1016/s0076-6879(89)73036-6. [DOI] [PubMed] [Google Scholar]
  2. Becker A., Neumeier R., Heidrich C., Loch N., Hartel S., Reutter W. Cell surface glycoproteins of hepatocytes and hepatoma cells identified by monoclonal antibodies. Biol Chem Hoppe Seyler. 1986 Aug;367(8):681–688. doi: 10.1515/bchm3.1986.367.2.681. [DOI] [PubMed] [Google Scholar]
  3. Büchler M., König J., Brom M., Kartenbeck J., Spring H., Horie T., Keppler D. cDNA cloning of the hepatocyte canalicular isoform of the multidrug resistance protein, cMrp, reveals a novel conjugate export pump deficient in hyperbilirubinemic mutant rats. J Biol Chem. 1996 Jun 21;271(25):15091–15098. doi: 10.1074/jbc.271.25.15091. [DOI] [PubMed] [Google Scholar]
  4. Elferink R. P., Ottenhoff R., Liefting W., de Haan J., Jansen P. L. Hepatobiliary transport of glutathione and glutathione conjugate in rats with hereditary hyperbilirubinemia. J Clin Invest. 1989 Aug;84(2):476–483. doi: 10.1172/JCI114189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Häussinger D., Hallbrucker C., Saha N., Lang F., Gerok W. Cell volume and bile acid excretion. Biochem J. 1992 Dec 1;288(Pt 2):681–689. doi: 10.1042/bj2880681. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Häussinger D., Saha N., Hallbrucker C., Lang F., Gerok W. Involvement of microtubules in the swelling-induced stimulation of transcellular taurocholate transport in perfused rat liver. Biochem J. 1993 Apr 15;291(Pt 2):355–360. doi: 10.1042/bj2910355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Jansen P. L., Groothuis G. M., Peters W. H., Meijer D. F. Selective hepatobiliary transport defect for organic anions and neutral steroids in mutant rats with hereditary-conjugated hyperbilirubinemia. Hepatology. 1987 Jan-Feb;7(1):71–76. doi: 10.1002/hep.1840070116. [DOI] [PubMed] [Google Scholar]
  8. Kartenbeck J., Leuschner U., Mayer R., Keppler D. Absence of the canalicular isoform of the MRP gene-encoded conjugate export pump from the hepatocytes in Dubin-Johnson syndrome. Hepatology. 1996 May;23(5):1061–1066. doi: 10.1053/jhep.1996.v23.pm0008621134. [DOI] [PubMed] [Google Scholar]
  9. Keppler D., Konig J. Hepatic canalicular membrane 5: Expression and localization of the conjugate export pump encoded by the MRP2 (cMRP/cMOAT) gene in liver. FASEB J. 1997 Jun;11(7):509–516. doi: 10.1096/fasebj.11.7.9212074. [DOI] [PubMed] [Google Scholar]
  10. Kubitz R., D'urso D., Keppler D., Häussinger D. Osmodependent dynamic localization of the multidrug resistance protein 2 in the rat hepatocyte canalicular membrane. Gastroenterology. 1997 Nov;113(5):1438–1442. doi: 10.1053/gast.1997.v113.pm9352844. [DOI] [PubMed] [Google Scholar]
  11. Kubitz R., Warskulat U., Schmitt M., Häussinger D. Dexamethasone- and osmolarity-dependent expression of the multidrug-resistance protein 2 in cultured rat hepatocytes. Biochem J. 1999 Jun 15;340(Pt 3):585–591. [PMC free article] [PubMed] [Google Scholar]
  12. Kubitz R., Wettstein M., Warskulat U., Häussinger D. Regulation of the multidrug resistance protein 2 in the rat liver by lipopolysaccharide and dexamethasone. Gastroenterology. 1999 Feb;116(2):401–410. doi: 10.1016/s0016-5085(99)70138-1. [DOI] [PubMed] [Google Scholar]
  13. 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]
  14. Müller M., Jansen P. L. Molecular aspects of hepatobiliary transport. Am J Physiol. 1997 Jun;272(6 Pt 1):G1285–G1303. doi: 10.1152/ajpgi.1997.272.6.G1285. [DOI] [PubMed] [Google Scholar]
  15. Müller M., Jansen P. L. The secretory function of the liver: new aspects of hepatobiliary transport. J Hepatol. 1998 Feb;28(2):344–354. doi: 10.1016/0168-8278(88)80024-2. [DOI] [PubMed] [Google Scholar]
  16. Noé B., Schliess F., Wettstein M., Heinrich S., Häussinger D. Regulation of taurocholate excretion by a hypo-osmolarity-activated signal transduction pathway in rat liver. Gastroenterology. 1996 Mar;110(3):858–865. doi: 10.1053/gast.1996.v110.pm8608896. [DOI] [PubMed] [Google Scholar]
  17. Paulusma C. C., Bosma P. J., Zaman G. J., Bakker C. T., Otter M., Scheffer G. L., Scheper R. J., Borst P., Oude Elferink R. P. Congenital jaundice in rats with a mutation in a multidrug resistance-associated protein gene. Science. 1996 Feb 23;271(5252):1126–1128. doi: 10.1126/science.271.5252.1126. [DOI] [PubMed] [Google Scholar]
  18. Rost D., Kartenbeck J., Keppler D. Changes in the localization of the rat canalicular conjugate export pump Mrp2 in phalloidin-induced cholestasis. Hepatology. 1999 Mar;29(3):814–821. doi: 10.1002/hep.510290319. [DOI] [PubMed] [Google Scholar]
  19. Stevenson B. R., Siliciano J. D., Mooseker M. S., Goodenough D. A. Identification of ZO-1: a high molecular weight polypeptide associated with the tight junction (zonula occludens) in a variety of epithelia. J Cell Biol. 1986 Sep;103(3):755–766. doi: 10.1083/jcb.103.3.755. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Taniguchi K., Wada M., Kohno K., Nakamura T., Kawabe T., Kawakami M., Kagotani K., Okumura K., Akiyama S., Kuwano M. A human canalicular multispecific organic anion transporter (cMOAT) gene is overexpressed in cisplatin-resistant human cancer cell lines with decreased drug accumulation. Cancer Res. 1996 Sep 15;56(18):4124–4129. [PubMed] [Google Scholar]
  21. Trauner M., Arrese M., Soroka C. J., Ananthanarayanan M., Koeppel T. A., Schlosser S. F., Suchy F. J., Keppler D., Boyer J. L. The rat canalicular conjugate export pump (Mrp2) is down-regulated in intrahepatic and obstructive cholestasis. Gastroenterology. 1997 Jul;113(1):255–264. doi: 10.1016/s0016-5085(97)70103-3. [DOI] [PubMed] [Google Scholar]
  22. Trauner M., Meier P. J., Boyer J. L. Molecular pathogenesis of cholestasis. N Engl J Med. 1998 Oct 22;339(17):1217–1227. doi: 10.1056/NEJM199810223391707. [DOI] [PubMed] [Google Scholar]
  23. Wahlländer A., Sies H. Glutathione S-conjugate formation from 1-chloro-2,4-dinitrobenzene and biliary S-conjugate excretion in the perfused rat liver. Eur J Biochem. 1979 Jun 1;96(3):441–446. doi: 10.1111/j.1432-1033.1979.tb13056.x. [DOI] [PubMed] [Google Scholar]
  24. Weibel E. R. Stereological principles for morphometry in electron microscopic cytology. Int Rev Cytol. 1969;26:235–302. doi: 10.1016/s0074-7696(08)61637-x. [DOI] [PubMed] [Google Scholar]

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