Progressive familial intrahepatic cholestasis 1 (PFIC1) results from mutations in ATP8B1 (also known as FIC1), a putative aminophospholipid flippase. However, conflicting hypotheses have been proposed for the pathogenesis of PFIC1. The aim of this study was to determine whether ATP8B1 deficiency produces cholestasis by altering the activity of the nuclear receptor farnesoid X receptor (FXR) or by impairing the structure of the canalicular membrane.

ATP8B1/Atp8b1 was knocked down in human and rat hepatocytes and Caco2 cells using adenoviral and oligonucleotide small interfering RNAs.

ATP8B1 messenger RNA and protein expression was greatly reduced in human and rat hepatocytes and Caco2 cells. In contrast, FXR expression and several FXR-dependent membrane transporters (bile salt export pump [BSEP], multidrug resistance-associated protein [MRP] 2) were unchanged at messenger RNA or protein levels in ATP8B1-deficient cells, whereas Mrp3 and Mrp4 were up-regulated in rat hepatocytes. FXR activity remained intact in these cells, as evidenced by 6α-ethyl chenodeoxycholic acid–mediated induction of small heterodimer partner (SHP), BSEP, and multidrug-resistant protein (MDR) 3/Mdr2. Fluorescent substrate excretion assays indicate that Bsep function was significantly reduced in Atp8b1-deficient rat hepatocytes, although Bsep remained localized to the canalicular membrane. Exposure to the hydrophobic bile acid CDCA resulted in focal areas of canalicular membrane disruption by electron microscopy and luminal accumulation of NBD-phosphatidylserine, consistent with the function of Atp8b1 as an aminophospholipid flippase.

ATP8B1 deficiency predisposes to cholestasis by favoring bile acid–induced injury in the canalicular membrane but does not directly affect FXR expression, which may occur in PFIC1 as a secondary phenomenon associated with bile acid accumulation.