Abstract
Sandwich-cultured (SC) rat hepatocytes are employed in drug discovery for pharmacological and toxicological assessment of drug candidates, yet their utility as a functional model for drug transporters has not been fully characterized. To evaluate the system as an in vitro model for drug transport, expression changes of hepatic transporters relative to whole liver and freshly-isolated hepatocytes (day-0) were examined by RT-qPCR for 4 consecutive days of culture. No significant differences in transporters expression levels were observed between freshly-isolated hepatocytes and whole liver. Two distinct mRNA profiles were detected over time showing: (1) more than 5-fold decline in levels of uptake transporters such as Ntcp, Oat2, Oatp1a1, 1a4, and 1b2; and (2) greater than 5-fold increase of efflux transporters Pgp, Bcrp, Mrp1, 2, and 3 and 4. Additionally, protein levels and functional activities for selected transporters were also determined. Protein levels for Mrp2, Bcrp, Pgp, Ntcp, and Oatp1a4 corresponded to changes in mRNA. Functional activities of Oatps and Oct1 exhibited 3- and 4-fold decrease on day-2 and day-4, respectively, relative to day-0, while more than 10-fold reduction in Oat2 activity was observed. These results indicate that cell culture conditions used herein did not provide an optimal environment for expression of all hepatic transporters. Significant time-dependent alterations in basal gene expression patterns of transporters were detected as compared to liver or freshly-isolated hepatocytes. Further work and new strategies are required to improve the validity of this model as an in vitro tool for in vivo drug transport or biliary clearance prediction.
- ABC transporters
- expression profiling
- hepatic elimination
- hepatic transport
- hepatobiliary transport
- hepatocytes
- MRP
- organic anion transport
- organic cation transport
- p-glycoprotein
- Received March 18, 2011.
- Accepted August 23, 2011.
- The American Society for Pharmacology and Experimental Therapeutics