Laboratory of Pharmacology and Toxicology, Graduate School of Pharmaceutical Sciences, Chiba University, Chiba-shi, Chiba, Japan.
J Hepatol. 2010 Apr;52(4):486-92. doi: 10.1016/j.jhep.2010.01.011. Epub 2010 Feb 4.
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.
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.
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.
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.
本研究旨在鉴定人肝细胞中主要的利巴韦林摄取转运体，并确定这些先前未被识别的转运体是否参与了肝内利巴韦林摄取。此外，我们旨在探讨造成人肝细胞摄取水平差异的原因。
通过转运试验和逆转录-聚合酶链反应（RT-PCR）分析高加索人肝细胞（HH268、HH283 和 HH291）中利巴韦林摄取和核苷转运体 mRNA 表达谱。通过确定转录起始位点和 RT-PCR 来描述 SLC29A1 基因结构的 5'-末端。
协同核苷转运蛋白 1（ENT1）介导的摄取完全参与 HH268 和 HH283 中的利巴韦林摄取，并且是 HH291 中最大的利巴韦林摄取部分的原因。HH291 中 ENT1 介导的摄取水平高于 HH268 和 HH283。SLC29A1 基因结构的特征显示，人肝中存在几种 ENT1 mRNA 异构体，HH291 中四种 ENT1 mRNA 异构体的水平高于 HH268 或 HH283。在任何肝细胞系中均未观察到 ENT2 介导的摄取。仅在 HH291 中检测到 Na+依赖性摄取；然而，所有肝细胞系中浓缩核苷转运蛋白（CNT）的 mRNA 水平均处于痕量水平。
ENT1 而非 ENT2 或 CNT 是人肝细胞中利巴韦林摄取的主要转运体。不同肝细胞系中 ENT1 介导的利巴韦林摄取水平的差异与特定 ENT1 mRNA 异构体的不同表达水平有关。此外，人肝细胞中可能存在一种未被识别的 Na+依赖性利巴韦林转运系统。
