Introduction: In vitro bile salt export pump (BSEP) inhibition alone does not accurately predict in vivo drug-induced liver injury in humans, suggesting that other mechanisms may be involved. Inhibition of BSEP has been shown to increase the hepatocellular concentrations of bile acids (BAs), and hepatobiliary disposition of BAs is tightly regulated by the farnesoid X receptor (FXR). Activation of FXR leads to decreased synthesis of BAs and increased expression of BA efflux transporters, BSEP, and organic solute and steroid transporter (OSTa/b). The link between BSEP inhibition and activation of the BA compensatory mechanism has not been clearly demonstrated. Materials and Methods: Therefore, we utilized BSEP inhibitors, cyclosporine A (CsA) and troglitazone (Trog), to explore this ''triggering'' event using Transporter CertifiedÔ sandwich-cultured human hepatocytes (SCHH). Results: Biliary excretion of glycine-cholic acid (GCA) as a percentage of total GCA accumulation was reduced in SCHH treated with either CsA or Trog. Within 12 hours, CsA treatment concomitantly increased intracellular concentrations (ICCs) of GCA and FGF19 mRNA content, an FXR-target gene. Separately, a synergistic 76.6fold increase of OSTb mRNA was observed following CsA and chenodeoxycholic acid co-exposure. In contrast, Trog exposure prevented the synergistic increase of OSTb, a component of the basolateral BA efflux transporter, under the same conditions. Finally, BA toxicity potency was increased in SCHH exposed to Trog but not CsA. Discussion and Conclusions: Overall, these results suggested that BSEP inhibition activates the BA compensatory mechanism reducing ICCs of BAs. However, Trog possesses both BSEP inhibition and FXR antagonist properties blocking the activation of the compensatory mechanism resulting in BA-induced hepatotoxicity.