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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

  • Molecular Toxicology
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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.

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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

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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).

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Authors and Affiliations

  1. Facultad de Ciencias Bioquímicas y Farmacéuticas, Instituto de Fisiología Experimental (IFISE) (CONICET-U.N.R.), Suipacha 570, S2002LRL, Rosario, Argentina

    Romina Belén Andermatten, Nadia Ciriaci, Virginia Soledad Schuck, María Valeria Razori, Gisel Sabrina Miszczuk, Anabela Carolina Medeot, Fernando Ariel Crocenzi, Marcelo Gabriel Roma, Ismael Ricardo Barosso & Enrique Juan Sánchez Pozzi

  2. Facultad de Farmacia y Bioquímica, Instituto de Investigaciones Farmacológicas (ININFA), Universidad de Buenos Aires, CONICET, Buenos Aires, Argentina

    Nicolás Di Siervi & Carlos Alberto Davio

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  1. Romina Belén Andermatten
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  2. Nadia Ciriaci
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Andermatten, R.B., Ciriaci, N., Schuck, V.S. et al. Sphingosine 1-phosphate receptor 2/adenylyl cyclase/protein kinase A pathway is involved in taurolithocholate-induced internalization of Abcc2 in rats. Arch Toxicol 93, 2279–2294 (2019). https://doi.org/10.1007/s00204-019-02514-6

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