Background & aims: The purpose of this study was to identify the major ribavirin uptake transporter(s) in human hepatocytes and to determine if these previously unidentified transporters are involved in hepatic ribavirin uptake. Furthermore, we aimed to address what causes the difference in uptake levels among human hepatocytes.
Methods: Profiles of ribavirin uptake and nucleoside transporter mRNA expression in Caucasian hepatocytes (HH268, HH283 and HH291) were characterized by transport assay and reverse transcription-polymerase chain reaction (RT-PCR). The 5'-side of the SLC29A1 gene structure was characterized by determination of transcription start sites and by RT-PCR.
Results: Equilibrative nucleoside transporter 1 (ENT1)-mediated uptake was exclusively involved in ribavirin uptake in HH268 and HH283 and was responsible for the largest ribavirin uptake fraction in HH291. The level of ENT1-mediated uptake in HH291 was higher than that in HH268 and HH283. Characterization of the SLC29A1 gene structure revealed the existence of several ENT1 mRNA isoforms in the human liver, and the levels of four ENT1 mRNA isoforms in HH291 were higher than those in HH268 or HH283. No ENT2-mediated uptake was observed in any hepatocyte lines. Na(+)-dependent uptake was detected only in HH291; however, mRNA levels of concentrative nucleoside transporters (CNTs) were at trace levels in all hepatocyte lines.
Conclusions: ENT1, but not ENT2 or CNTs, is a major ribavirin uptake transporter in human hepatocytes. The different ENT1-mediated ribavirin uptake levels in different hepatocyte lines are associated with different expression levels of specific isoforms of ENT1 mRNAs. Furthermore, an unidentified Na(+)-dependent ribavirin transport system might exist in human hepatocytes.