Oxidative stress plays a major role in... : Hepatology (original) (raw)

Autoimmune, Cholestatic and Biliary Disease

Oxidative stress plays a major role in chlorpromazine-induced cholestasis in human HepaRG cells

Anthérieu, Sébastien1,2; Azzi, Pamela Bachour-El1,2,3; Dumont, Julie1,2; Abdel-Razzak, Ziad3; Guguen-Guillouzo, Christiane1,2,4; Fromenty, Bernard1,2; Robin, Marie-Anne1,2; Guillouzo, André1,2,*

1_Inserm UMR991, Liver Metabolisms and Cancer, Rennes, France_

2_Université de Rennes 1, Rennes, France_

3_Université Libanaise, EDST-PRASE and EDST-AZM-center-LBA3B, Lebanon_

4_Biopredic International, St Grégoire, France_

curr1_Inserm UMR 744, Université Lille Nord de France, Institut Pasteur, Lille, France_

*Inserm UMR 991, Faculté des Sciences Pharmaceutiques et Biologiques, 35043 Rennes Cedex, France

Email:[email protected]

Received 24 January 2012; Accepted 24 October 2012

View this article online atwileyonlinelibrary.com.

Grant sponsor: European Community Contracts; Grant Numbers: LIINTOP-STREP-037499 Predict-IV-202222; Grant sponsor: Pamela Bachour-El Azzi was provided by Lebanese University, Lebanese CNRS, Philippe Jabre association and the Ecole Doctorale VAS Rennes.

Potential conflict of interest: Nothing to report.

These authors contributed equally to this work.

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Abstract

Drugs induce cholestasis by diverse and still poorly understood mechanisms in humans. Early hepatic effects of chlorpromazine (CPZ), a neuroleptic drug known for years to induce intrahepatic cholestasis, were investigated using the differentiated human hepatoma HepaRG cells. Generation of reactive oxygen species (ROS) was detected as early as 15 minutes after CPZ treatment and was associated with an altered mitochondrial membrane potential and disruption of the pericanalicular distribution of F-actin. Inhibition of [3H]-taurocholic acid efflux was observed after 30 minutes and was mostly prevented by N-acetyl cysteine (NAC) cotreatment, indicating a major role of oxidative stress in CPZ-induced bile acid (BA) accumulation. Moreover, 24-hour treatment with CPZ decreased messenger RNA (mRNA) expression of the two main canalicular bile transporters, bile salt export pump (BSEP) and multidrug resistance protein 3 (MDR3). Additional CPZ effects included inhibition of Na+-dependent taurocholic cotransporting polypeptide (NTCP) expression and activity, multidrug resistance-associated protein 4 (MRP4) overexpression and CYP8B1 inhibition that are involved in BA uptake, basolateral transport, and BA synthesis, respectively. These latter events likely represent hepatoprotective responses which aim to reduce intrahepatic accumulation of toxic BA. Compared to CPZ effects, overloading of HepaRG cells with high concentrations of cholic and chenodeoxycholic acids induced a delayed oxidative stress and, similarly, after 24 hours it down-regulated BSEP and MDR3 in parallel to a decrease of NTCP and CYP8B1 and an increase of MRP4. By contrast, low BA concentrations up-regulated BSEP and MDR3 in the absence of oxidative stress. Conclusion : These data provide evidence that, among other mechanisms, oxidative stress plays a major role as both a primary causal and an aggravating factor in the early CPZ-induced intrahepatic cholestasis in human hepatocytes.