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Sphingosine 1-phosphate receptor 2/adenylyl cyclase/protein kinase A pathway is involved in taurolithocholate-induced internalization of Abcc2 in rats

  • Romina Belén Andermatten
  • Nadia Ciriaci
  • Virginia Soledad Schuck
  • Nicolás Di Siervi
  • María Valeria Razori
  • Gisel Sabrina Miszczuk
  • Anabela Carolina Medeot
  • Carlos Alberto Davio
  • Fernando Ariel Crocenzi
  • Marcelo Gabriel Roma
  • Ismael Ricardo Barosso
  • Enrique Juan Sánchez PozziEmail author
Molecular Toxicology

Abstract

Taurolithocholate (TLC) is a cholestatic bile salt that induces disinsertion of the canalicular transporter Abcc2 (Mrp2, multidrug resistance-associated protein 2). This internalization is mediated by different intracellular signaling proteins such as PI3K, PKCε and MARCK but the initial receptor of TLC remains unknown. A few G protein-coupled receptors interact with bile salts in hepatocytes. Among them, sphingosine-1 phosphate receptor 2 (S1PR2) represents a potential initial receptor for TLC. The aim of this study was to evaluate the role of this receptor and its downstream effectors in the impairment of Abcc2 function induced by TLC. In vitro, S1PR2 inhibition by JTE-013 or its knockdown by small interfering RNA partially prevented the decrease in Abcc2 activity induced by TLC. Moreover, adenylyl cyclase (AC)/PKA and PI3K/Akt inhibition partially prevented TLC effect on canalicular transporter function. TLC produced PKA and Akt activation, which were blocked by JTE-013 and AC inhibitors, connecting S1PR2/AC/PKA and PI3K/Akt in a same pathway. In isolated perfused rat liver, injection of TLC triggered endocytosis of Abcc2 that was accompanied by a sustained decrease in the bile flow and the biliary excretion of the Abcc2 substrate dinitrophenyl-glutathione until the end of the perfusion period. S1PR2 or AC inhibition did not prevent the initial decay, but they accelerated the recovery of these parameters and the reinsertion of Abcc2 into the canalicular membrane. In conclusion, S1PR2 and the subsequent activation of AC, PKA, PI3K and Akt is partially responsible for the cholestatic effects of TLC through sustained internalization of Abcc2.

Keywords

Mrp2 S1PR2 Cholestasis ABC transporters 

Abbreviations

Abcc2

Multidrug resistance-associated protein 2

TLC

Taurolithocholate

S1PR2

Sphingosine 1-phosphate receptor 2

PKA

Protein kinase A

AC

Adenylyl cyclase

CMFDA

5-Chloromethylfluorescein diacetate

GSMF

Glutathione methyl fluorescein

DMSO

Dimethylsulfoxide

IRHC

Isolated rat hepatocyte couplets

cVA

Canalicular vacuolar accumulation

SCRH

Sandwich-cultured rat hepatocytes

IPRL

Isolated perfused rat liver

Notes

Acknowledgements

We thank José Pellegrino and Rodrigo Vena for assistance with confocal microscopy, and Diego Taborda for technical assistance with animal procedures and cell isolation.

Funding

This work was supported by grants from Agencia Nacional de Promoción Científica y Tecnológica (PICTs 2013 No. 1222 and 2016 No. 2166) and National University of Rosario-UNR (BIO436/2015).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

204_2019_2514_MOESM1_ESM.docx (296 kb)
Supplementary material 1 (DOCX 295 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Romina Belén Andermatten
    • 1
  • Nadia Ciriaci
    • 1
  • Virginia Soledad Schuck
    • 1
  • Nicolás Di Siervi
    • 2
  • María Valeria Razori
    • 1
  • Gisel Sabrina Miszczuk
    • 1
  • Anabela Carolina Medeot
    • 1
  • Carlos Alberto Davio
    • 2
  • Fernando Ariel Crocenzi
    • 1
  • Marcelo Gabriel Roma
    • 1
  • Ismael Ricardo Barosso
    • 1
  • Enrique Juan Sánchez Pozzi
    • 1
    Email author
  1. 1.Facultad de Ciencias Bioquímicas y FarmacéuticasInstituto de Fisiología Experimental (IFISE) (CONICET-U.N.R.)RosarioArgentina
  2. 2.Facultad de Farmacia y Bioquímica, Instituto de Investigaciones Farmacológicas (ININFA)Universidad de Buenos Aires, CONICETBuenos AiresArgentina

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