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This Article Full Text Full Text (PDF) Supplemental Tables All Versions of this Article: 296/5/G1119    most recent 90371.2008v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Martínez-Fernández, P. Articles by Alvarez, L. Search for Related Content PubMed PubMed Citation Articles by Martínez-Fernández, P. Articles by Alvarez, L.

LIVER AND BILIARY TRACT

Knockdown of ATP8B1 expression leads to specific downregulation of the bile acid sensor FXR in HepG2 cells: effect of the FXR agonist GW4064

¹Research Unit, La Paz University Hospital-Fundación para la Investigación Biomédica del Hospital Universitario La Paz (FIBHULP) and ²Pediatric Liver Service, La Paz Children's University Hospital, Madrid, Spain

Submitted 10 June 2008 ; accepted in final form 16 February 2009

Farnesoid X receptor (FXR) is a bile acid-sensing nuclear receptor that controls bile acid homeostasis. It has been suggested that downregulation of FXR contributes to the pathogenesis of an inherited disorder of bile secretion caused by mutations in ATP8B1. We have investigated the relationship between ATP8B1 knockdown and FXR downregulation in the human hepatoblastoma cell line HepG2. Transfection of HepG2 cells with ATP8B1 small interfering RNA (siRNA) duplexes led to a 60% reduction in the endogenous levels of ATP8B1 mRNA and protein and a concomitant decrease in FXR mRNA and protein content, as well as in FXR phosphorylation. This decrease was accompanied by a marked reduction in mRNA levels of a subset of FXR targets, such as bile salt export pump (ABCB11), small heterodimer partner, and uridine 5'-diphosphate-glucuronosyltransferase. ATP8B1 inhibition specifically targeted FXR since mRNA expression of other prominent nuclear receptors, such as pregnane X receptor and constitutive androstane receptor, or liver-enriched transcription factors, such as hepatocyte nuclear factor 1 (HNF-1) and HNF-4, was not altered. The expression of other key genes involved in bile acid synthesis, detoxification, and transport also remained unchanged upon ATP8B1 knockdown. Supporting the specificity of the effect, siRNA-mediated silencing of ABCB11, whose defect is associated with another inherited disorder of bile secretion, did not affect FXR expression. Treatment with the synthetic FXR agonist GW4064 was able to partially neutralize ATP8B1 siRNA-mediated FXR downregulation and fully counteract inhibition of FXR target genes. Collectively these findings indicate that ATP8B1 knockdown specifically downregulates FXR, and this action can be circumvented by treatment with FXR agonists.

farnesoid X receptor; ABCB11; nuclear receptors; progressive familial intrahepatic cholestasis