Identification of drug transporters involved in the uptake and efflux of rhein in hepatocytes

Aim: Rhein is a herbal medicine with various bioactivities and is derived from an anthraquinone compound. In this study, we aimed to identify drug transporters involved in the uptake and efflux of rhein in hepatocytes. Methods: Rhein uptake and efflux were investigated in rat liver slices, primary rat hepatocytes, human transporter-transfected cells, membrane vesicles, and rats. Rhein was quantified using liquid chromatography-tandem mass spectrometry. Results: Rhein uptake was fitted to a Michaelis-Menten model with a K-m = 229.4 +/- 58.5 mu M and V-max = 428.7 +/- 49.2 pmol mg(-1) protein in liver slices and a K-m = 129.6 +/- 22.3 mu M and V-max = 9.1 +/- 0.6 nmol mg(-1) protein in primary rat hepatocytes. Rhein uptake by primary rat hepatocytes was inhibited by organic anion-transporting polypeptide (OATP) inhibitors (rifampicin and fluvastatin) and an organic anion transporter (OAT) inhibitor (probenecid). Rhein was taken up by HEK293 cells expressing OATP1B3, OATP2B1, and OAT2 and pumped out of MDCKII cells expressing breast cancer resistance protein (BCRP), but not from those expressing P-glycoprotein (P-gp) or from membrane vesicles expressing multidrug resistance-associated proteins (MRPs) 2 and 3. In rats, hepatic exposure to rhein was reduced by rifampicin and probenecid (AUC(0-2) h decreased 0.61- and 0.12-fold, respectively), but increased by pantoprazole (AUC(0-2) h increased 0.64-fold), a BCRP inhibitor. Conclusion: These results show that OATP1B3, OATP2B1, and OAT2 contribute to hepatic rhein uptake, while BCRP contributes to its hepatic efflux. The results provide new insights into the mechanisms of rhein uptake and efflux in hepatocytes.