Establishment and characterization of Mardin-Darby canine kidney cells stably expressing human organic anion transporters.

Organic anion transporters (OATs) play essential roles in the renal elimination of clinically important anionic drugs. The purpose of this study was to establish an efficient in vitro assay system to screen the transport characteristics of drugs and to examine drug interaction potentials of drugs with OATs. First, we established Mardin-Darby canine kidney (MDCK) cells stably expressing OAT1, OAT3, and OAT4 (MDCK-OAT1, -OAT3, and -OAT4, respectively). To characterize these cells, [(14)C]para-aminohippuric acid and [(3)H]estrone-3-sulfate transport properties were measured, and these corresponded to the results of the mRNA expression and localization of respective transporters using RT-PCR and immunofluorescence staining assay. Additionally, we screened three herbal medicines, Woohwangcheongsimwon, Hawangyeonhaedoktang, and Aristolochiae fructus extracts, which have been widely used in Korea or reported for nephrotoxicity, in our MDCK-OAT1, -OAT3, and -OAT4 cells. Aristolochiae fructus extracts strongly inhibited organic anion uptake by OAT1, OAT3, and OAT4, whereas Woohwangcheongsimwon only interacted with OAT1, and Hawangyeonhaedoktang with OAT1 and OAT3, suggesting drug interaction potential with OATs-mediated renal eliminating drugs. In conclusion, we established and characterized MDCK cells stably expressing OAT1, OAT3, and OAT4 for the elucidation of substrates or inhibitors of respective transporters as high-throughput screening tools.