Lithocholic Acid Decreases Expression of Bile Salt Export Pump through Farnesoid X Receptor Antagonist Activity

Jinghua Yu,Jane-L Lo,Li Huang,Annie Zhao,Edward Metzger,Alan Adams,Peter T Meinke,Samuel D Wright,Jisong Cui

doi:10.1074/jbc.m200474200

Abstract

Bile salt export pump (BSEP) is a major bile acid transporter in the liver. Mutations in BSEP result in progressive intrahepatic cholestasis, a severe liver disease that impairs bile flow and causes irreversible liver damage. BSEP is a target for inhibition and down-regulation by drugs and abnormal bile salt metabolites, and such inhibition and down-regulation may result in bile acid retention and intrahepatic cholestasis. In this study, we quantitatively analyzed the regulation of BSEP expression by FXR ligands in primary human hepatocytes and HepG2 cells. We demonstrate that BSEP expression is dramatically regulated by ligands of the nuclear receptor farnesoid X receptor (FXR). Both the endogenous FXR agonist chenodeoxycholate (CDCA) and synthetic FXR ligand GW4064 effectively increased BSEP mRNA in both cell types. This up-regulation was readily detectable at as early as 3 h, and the ligand potency for BSEP regulation correlates with the intrinsic activity on FXR. These results suggest BSEP as a direct target of FXR and support the recent report that the BSEP promoter is transactivated by FXR. In contrast to CDCA and GW4064, lithocholate (LCA), a hydrophobic bile acid and a potent inducer of cholestasis, strongly decreased BSEP expression. Previous studies did not identify LCA as an FXR antagonist ligand in cells, but we show here that LCA is an FXR antagonist with partial agonist activity in cells. In an in vitro co-activator association assay, LCA decreased CDCA- and GW4064-induced FXR activation with an IC(50) of 1 microm. In HepG2 cells, LCA also effectively antagonized GW4064-enhanced FXR transactivation. These data suggest that the toxic and cholestatic effect of LCA in animals may result from its down-regulation of BSEP through FXR. Taken together, these observations indicate that FXR plays an important role in BSEP gene expression and that FXR ligands may be potential therapeutic drugs for intrahepatic cholestasis.

Highlights

Bile salt export pump (BSEP)1 mediates the rate-limiting step in overall hepatocellular bile salt excretion, the transport of bile acids across the canalicular membrane [1,2,3]
Mutations in BSEP result in progressive familial intrahepatic cholestasis type 2, a severe liver disease characterized by impaired bile flow and irreversible liver damage [4]
We demonstrate that CDCA and a synthetic farnesoid X receptor (FXR) agonist, GW4064 [22], dramatically up-regulate expression of BSEP mRNA in primary human hepatocytes and HepG2

Summary

Introduction

Bile salt export pump (BSEP) mediates the rate-limiting step in overall hepatocellular bile salt excretion, the transport of bile acids across the canalicular membrane [1,2,3]. We show that LCA down-regulates BSEP expression through its antagonist activity on the nuclear hormone receptor FXR. This down-regulation may explain the cholestatic effect of LCA. It has been shown that FXR inhibits expression of cholesterol 7␣-hydroxylase (Cyp 7a) (14 –17), the enzyme catalyzing the first and rate-limiting step of bile acid synthesis [18], and activates expression of intestinal bile acidbinding protein [11], phospholipid transfer protein [19], BSEP [20], and dehydroepiandrosterone sulfotransferase [21]. The FXR antagonist LCA effectively down-regulates BSEP gene expression. These data support the notion of BSEP as a direct target of FXR

Methods

Results

Conclusion