Enhancement of genotype 1 hepatitis C virus replication by bile acids through FXR

Background/Aims

Hepatitis C virus (HCV) infected patients with high serum levels of bile acids (BAs) usually fail to respond to antiviral therapy. Besides, BAs are essential factors for replication of the porcine enteric calicivirus by inhibiting interferon signaling. The role of BAs on HCV RNA replication was thus assessed.

Methods

BAs and other compounds were tested using an HCV-replication model containing a luciferase reporter gene.

Results

BAs, especially chenodeoxycholate and deoxycholate, up-regulated genotype 1 HCV RNA replication by more than tenfold. Only free but not conjugated BAs were active, suggesting that their effect was mediated by a nuclear receptor. Only farnesoid X receptor (FXR) ligands stimulated HCV replication while FXR silencing and FXR antagonism by guggulsterone blocked the up-regulation induced by BAs. Furthermore, guggulsterone alone inhibited basal level of HCV replication by tenfold. Modulation of HCV replication by FXR ligands occurred in the same proportion in presence or absence of type I interferon, suggesting a mechanism of action independent of this control of viral replication. However, BAs or guggulsterone did not affect replication of genotype 2a-JFH1.

Conclusions

Exposure to routinely measured concentrations of BAs increases HCV replication by a novel mechanism involving activation of the nuclear receptor FXR.

Introduction

Hepatitis C virus (HCV) is a single-stranded positive RNA virus, which belongs to the genus Hepacivirus of the family Flaviviridæ. Around 170 million patients worldwide are chronically infected with HCV, frequently resulting in liver cirrhosis or hepatocellular carcinoma [1]. The outcome of HCV infection varies among individuals and the likelihood of achieving a sustained response to antiviral treatment (SVR) depends on both viral and host characteristics. HCV variants are classified into six major genotypes that have a varied response to antiviral therapy. Indeed, only 45% of genotype 1 infected patients but 80% of those infected with genotype 2 or 3 reach a SVR with the current therapy combining ribavirin and peginterferon [2]. Among host factors associated with therapy outcome, serum bile acids (BAs) have been recently described as prognostic markers predicting a failure to reach SVR [3], [4]. Particularly, concentrations exceeding 15 μM associated with ferritin concentrations over 300 μg/ml are predictive of non-response to therapy. Primary BAs cholate (CA) and chenodeoxycholate (CDCA) are synthesized in the liver from cholesterol, conjugated with glycine or taurine to increase their hydrophilicity and secreted into the bile. In the intestinal tract, primary BAs are 7α-dehydroxylated by commensal bacteria, producing secondary BAs lithocholate (LCA) and deoxycholate (DCA). They are reabsorbed in the ileum and transported back to the liver [5]. BAs are not simply cholesterol’s catabolites, but are truly regulatory molecules with specific receptors. They control gene expression via nuclear receptors of the NR1 family: farnesoid X receptor (FXR; NR1H4) [6], [7], pregnane X receptor (PXR) [8], and vitamin D receptor [9]. Natural ligands of these receptors (BAs, sterols, fatty acids) are physiologically present at high concentrations [10] and activate their receptors with EC50 of 10–15 μM [11]. FXR is mainly expressed in the liver and intestine [12]. Hepatic FXR target genes are classified into several groups [11], [13]; one decreases hepatic BA concentrations by increasing export and decreasing synthesis; a second modulates seric lipoprotein levels and decreases plasma triglyceride concentrations. FXR also controls liver growth and regeneration in response to variations of BA concentrations [14], [15]. BAs also activate MAPK pathways through TGR5, a G-protein-coupled membrane receptor [16], [17]. TGR5 increases intracellular cAMP levels, thus inhibiting the activation of STAT1 by type I interferon (IFN). This IFN interference by BAs is an essential factor for the growth of the porcine enteric calicivirus (PEC), another single-stranded positive RNA virus [18].

In the light of recent observations that lipids play an important role in regulating HCV replication and eventually virus infectivity [19], [20], [21], the aim of the present study was to assess whether BAs had a direct effect on HCV RNA replication in cell lines harbouring HCV replicons.