Obeticholic acid, a selective farnesoid X receptor agonist, regulates bile acid homeostasis in sandwich‐cultured human hepatocytes

Abstract

Farnesoid X receptor (FXR) is a master regulator of bile acid homeostasis through transcriptional regulation of genes involved in bile acid synthesis and cellular membrane transport. Impairment of bile acid efflux due to cholangiopathies results in chronic cholestasis leading to abnormal elevation of intrahepatic and systemic bile acid levels. Obeticholic acid (OCA) is a potent and selective FXR agonist that is 100‐fold more potent than the endogenous ligand chenodeoxycholic acid (CDCA). The effects of OCA on genes involved in bile acid homeostasis were investigated using sandwich‐cultured human hepatocytes. Gene expression was determined by measuring mRNA levels. OCA dose‐dependently increased fibroblast growth factor‐19 (FGF‐19) and small heterodimer partner (SHP) which, in turn, suppress mRNA levels of cholesterol 7‐alpha‐hydroxylase (CYP7A1), the rate‐limiting enzyme for de novo synthesis of bile acids. Consistent with CYP7A1 suppression, total bile acid content was decreased by OCA (1 μmol/L) to 42.7 ± 20.5% relative to control. In addition to suppressing de novo bile acids synthesis, OCA significantly increased the mRNA levels of transporters involved in bile acid homeostasis. The bile salt excretory pump (BSEP), a canalicular efflux transporter, increased by 6.4 ± 0.8‐fold, and the basolateral efflux heterodimer transporters, organic solute transporter α (OST α) and OST β increased by 6.4 ± 0.2‐fold and 42.9 ± 7.9‐fold, respectively. The upregulation of BSEP and OST α and OST β, by OCA reduced the intracellular concentrations of d8‐TCA, a model bile acid, to 39.6 ± 8.9% relative to control. These data demonstrate that OCA does suppress bile acid synthesis and reduce hepatocellular bile acid levels, supporting the use of OCA to treat bile acid‐induced toxicity observed in cholestatic diseases.

Comparative potency of obeticholic acid and natural bile acids on FXR in hepatic and intestinal in vitro cell models

“… FXR activation by OCA has an EC 50 value of approximately 100 nmol/L . Previous studies by the authors utilizing a physiologically relevant cellular model of sandwich‐cultured human hepatocytes (SCHH) demonstrated a significant reduction in endogenous bile acids following 72‐hour incubation of OCA . In the same study, a head‐to‐head comparison confirmed that OCA was 100‐fold more potent than CDCA on FXR.…”

“…Previous studies in SCHH have demonstrated that OCA and CDCA up to 100 μmol/L showed no overt cytotoxicity . To optimize treatment concentration of UDCA, cytotoxicity of increasing concentrations of UDCA (1.0‐1000 μmol/L) was evaluated by morphological and ATP assessments (Figure ).…”

“…7 Obeticholic acid (OCA) is an analog of CDCA that differs in structure by a single ethyl group at C 6. 14 FXR activation by OCA has an EC 50 value of approximately 100 nmol/L. 14 15 In the same study, a head-to-head comparison confirmed that OCA was 100-fold more potent than CDCA on FXR. In a Phase 3 clinical trial 16 , patients with primary biliary cholangitis (PBC), treated with OCA once daily, consistently had significant reductions in plasma bile acid levels compared to placebo controls.…”

“…Obeticholic acid does activate hepatic and intestinal FXR‐FGF‐19/SHP cascades, thereby substantially reducing bile acid synthesis. We have previously reported and discussed the hepatic mechanisms of action of OCA . Herein, we further characterized glyco‐OCA in Caco‐2 cells to understand its intestinal mechanisms of action.…”

“…EBAP in SCHH was analyzed using B‐CLEAR ® technology and calculated as the sum of endogenous primary bile acids (CA and CDCA) and their glycine and taurine conjugates (glyco‐CA, tauro‐CA, glyco‐CDCA, and tauro‐CDCA) in cell culture medium, and hepatocyte lysate (cell + bile). Endogenous bile acids were extracted from cell culture medium samples and hepatocyte lysate using the same procedure described in the previous paper . Prepared samples were filtered and analyzed by LC‐MS/MS using a Shimadzu binary HPLC system (Columbia, MD) and tandem mass spectrometry using Thermo Electron TSQ ® Quantum Discovery MAX™ (Waltham, MA) with an Ion Max ESI source operating in negative ion electrospray ionization mode using multiple reaction monitoring.…”