Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2018 Jun 1;159(6):2435-2446.
doi: 10.1210/en.2018-00110.

Xenobiotic Nuclear Receptor Signaling Determines Molecular Pathogenesis of Progressive Familial Intrahepatic Cholestasis

Kang Ho Kim  1 , Jong Min Choi  1 , Feng Li  1   2 , Armando Arizpe  3 , Clavia Ruth Wooton-Kee  1 , Sayeepriyadarshini Anakk  4 , Sung Yun Jung  1   5 , Milton J Finegold  6 , David D Moore  1
Affiliations

Xenobiotic Nuclear Receptor Signaling Determines Molecular Pathogenesis of Progressive Familial Intrahepatic Cholestasis

Kang Ho Kim et al. Endocrinology. .

Abstract

Progressive familial intrahepatic cholestasis (PFIC) is a genetically heterogeneous disorder of bile flow disruption due to abnormal canalicular transport or impaired bile acid (BA) metabolism, causing excess BA accumulation and liver failure. We previously reported an intrahepatic cholestasis mouse model based on loss of function of both farnesoid X receptor (FXR; NR1H4) and a small heterodimer partner (SHP; NR0B2) [double knockout (DKO)], which has strong similarities to human PFIC5. We compared the pathogenesis of DKO livers with that of another intrahepatic cholestasis model, Bsep-/-, which represents human PFIC2. Both models exhibit severe hepatomegaly and hepatic BA accumulation, but DKO showed greater circulating BA and liver injury, and Bsep-/- had milder phenotypes. Molecular profiling of BAs uncovered specific enrichment of cholic acid (CA)-derived BAs in DKO livers but chenodeoxycholate-derived BAs in Bsep-/- livers. Transcriptomic and proteomic analysis revealed specific activation of CA synthesis and alternative basolateral BA transport in DKO but increased chenodeoxycholic acid synthesis and canalicular transport in Bsep-/-. The constitutive androstane receptor (CAR)/pregnane X receptor (PXR)-CYP2B/CYP2C axis is activated in DKO livers but not in other cholestasis models. Loss of this axis in Fxr:Shp:Car:Pxr quadruple knockouts blocked Cyp2b/Cyp2c gene induction, impaired bilirubin conjugation/elimination, and increased liver injury. Differential CYP2B expression in DKO and Bsep-/- was recapitulated in human PFIC5 and PFIC2 livers. In conclusion, loss of FXR/SHP results in distinct molecular pathogenesis and CAR/PXR activation, which promotes Cyp2b/Cyp2c gene transcription and bilirubin clearance. CAR/PXR activation was not observed in Bsep-/- mice or PFIC2 patients. These findings provide a deeper understanding of the heterogeneity of intrahepatic cholestasis.

PubMed Disclaimer

Figures

Figure 1.
Figure 1.
DKO andBsep−/− mice showed hepatomegaly. (A) Representative view of whole body and liver at 3 months of age. Scale bar, 10 mm. (B) Body weight (BW), liver weight (LW), and LW/BW ratio at the time of death (n = 3 for each group). (C) Hematoxylin and eosin staining of liver (magnification, ×200). Arrowheads indicate hepatic steatosis; arrows indicate bile duct proliferation. Statistics: Studentt test: *P < 0.05 and ***P < 0.005 compared with each control. C, central vein; P, portal vein.
Figure 2.
Figure 2.
Differential BA accumulation and liver injury in DKO andBsep−/−. (A) Hepatic and serum total BA levels in DKO andBsep−/− serum (n = 3 for each group). (B) Serum ALT, AST, and bilirubin (direct, indirect, and total) levels. Ratio (%) indicated the proportions of direct (white) and indirect (gray) bilirubin. Statistics: Studentt test: *P < 0.05, **P < 0.01, ***P < 0.005 compared with each control. In some cases, the actualP value is presented. TBA, total bile acid.
Figure 3.
Figure 3.
The DKO andBsep−/− mice showed differential BA profiling. BA composition of (A) liver and (B) serum in WT, DKO, andBsep−/− mice. BAs were quantified as the percentage of total BA analyzed (100%). CA-derived primary BAs shown in blue and dark blue [CA, tauro-CA (TCA), glyco-CA (GCA), deoxycholic acid (DCA), and tauro-DCA (TDCA)]. CDCA-derived primary BAs shown in orange or dark red [CDCA, tauro-CDCA (TCDCA),αβMCA, and tauro-αβMCA (TαβMCA)]. TUDCA, tauro-ursodeoxycholic acid.
Figure 4.
Figure 4.
Expression profile of representative genes involved in BA homeostasis. The (A) mRNA and (B) protein profile of representative BA synthesis, metabolism, transport, and uptake genes in 3-month-old DKO (upper) andBsep−/− (lower) mice was analyzed using real-time quantitative PCR and proteomics. Relative protein levels are indicated as intensity-based fraction of the total (iFOT). In addition to the official gene symbol, the common gene name is given in parentheses, if necessary. Statistics: Studentt test: *P < 0.05, **P < 0.01, ***P < 0.005 compared with each control. In some cases, the actualP value is presented. ND, not detected.
Figure 5.
Figure 5.
Specific activation of CYP2B and CYP2C in DKO. (A) Upregulated genes of CYP and GST (GS transferase) gene family (P < 0.005 andP < 0.001 in DKO, respectively).P values in DKO separately indicated by asterisks (*, **, ***). Representative CYP and GST gene expression in (B) DKO and (C)Bsep−/− were confirmed by real-time quantitative PCR. Statistics: Studentt test: *P < 0.05, **P < 0.01, ***P < 0.005 compared with each control. In some cases, the actualP value is presented.
Figure 6.
Figure 6.
Deletion of CAR/PXR exacerbates BA-induced hepatotoxicity. The mRNA expression of representative (A) CYP and (B) GSTs was evaluated by real-time quantitative PCR (n = 3 for each group). (C) Serum bilirubin levels in DKO and QKO. Ratio (%) indicated the proportions of direct (white) or indirect (gray) bilirubin compared with total bilirubin. (D) Hematoxylin and eosin images of WT, DKO, and QKO liver. Black arrowheads indicate hepatic steatosis; white arrowheads indicate cytoplasmic clearing and severe hepatocyte swelling. Statistics: (A and B)P values of one-way ANOVA test indicated.Post hoc t test with Bonferroni correction:†,#P < 0.0167,††,##P < 0.00333,†††,###P < 0.00167;†compared with WT;#compared with DKO. (D) Studentt test: *P < 0.05, ***P < 0.005 compared with DKO.
Figure 7.
Figure 7.
Upregulation of CYP2B expression in DKO and PFIC5 livers. (A) Mouse liver sections from WT, DKO, QKO,Bsep+/−, andBsep−/− mice were stained with anti-BSEP (upper panel) and anti-CYP2B6 (lower panel) antibodies. Next, nuclei were counterstained with hematoxylin. Arrowheads indicate canalicular BSEP expression. (B) Cirrhotic liver sections from end-stage disease of patients with PFIC5 and PFIC2 were stained with anti-CYP2B6 antibody. (Inset) Parenchymal CYP2B6 staining. Arrowhead indicates positive CYP2B6 staining in a biliary tract.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Beuers U,Trauner M,Jansen P,Poupon R. New paradigms in the treatment of hepatic cholestasis: from UDCA to FXR, PXR and beyond.J Hepatol.2015;62(1,Suppl):S25–S37. - PubMed
    1. Mazokopakis EE,Papadakis JA,Kofteridis DP. Unusual causes of intrahepatic cholestatic liver disease.World J Gastroenterol.2007;13(12):1879–1882. - PMC - PubMed
    1. Schady DA,Finegold MJ. Contemporary evaluation of the pediatric liver biopsy.Gastroenterol Clin North Am.2017;46(2):233–252. - PubMed
    1. Amer S,Hajira A. A comprehensive review of progressive familial intrahepatic cholestasis (PFIC): genetic disorders of hepatocanalicular transporters.Gastroenterol Res.2014;7(2):39–43. - PMC - PubMed
    1. Gomez-Ospina N,Potter CJ,Xiao R,Manickam K,Kim MS,Kim KH,Shneider BL,Picarsic JL,Jacobson TA,Zhang J,He W,Liu P,Knisely AS,Finegold MJ,Muzny DM,Boerwinkle E,Lupski JR,Plon SE,Gibbs RA,Eng CM,Yang Y,Washington GC,Porteus MH,Berquist WE,Kambham N,Singh RJ,Xia F,Enns GM,Moore DD. Mutations in the nuclear bile acid receptor FXR cause progressive familial intrahepatic cholestasis.Nat Commun.2016;7:10713. - PMC - PubMed

Publication types

MeSH terms

Substances

Supplementary concepts

LinkOut - more resources