Functional Expression of Organic Anion Transporters in Hepatic Organoids Reconstructed by Rat Small Hepatocytes
Functional Expression of Organic Anion Transporters in Hepatic Organoids Reconstructed by Rat Small Hepatocytes
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DOI：
10.1002/jcb.21601
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发表时间：
2008-05-01
影响因子：
4
通讯作者：
Mitaka, Toshihiro
中科院分区：
文献类型：
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作者：
Oshima, Hideki;Kon, Junko;Mitaka, Toshihiro
小肝细胞（small hepatocytes，SH）是具有肝脏特征的肝祖细胞。它们可以在培养物中增殖形成集落，并将其形态从扁平变为与胆小管（BC）一起升高/堆积，这导致成熟。在这项研究中，我们研究了是否SH可以表达极性的肝转运蛋白，转运蛋白是否可以运输有机阴离子底物进入BC，以及分泌的物质是否可以从BC中回收。采用免疫细胞化学和RT-PCR方法进行检测。[H-3]-标记的雌激素衍生物用于测量从正常和多药耐药相关蛋白（Mrp）2缺陷大鼠分离的SH中转运体的功能。结果表明，有机阴离子转运蛋白（Oatps）1和2，Na+依赖的牛磺胆酸共转运多肽（Ntcp），Mrp 2和胆盐输出泵（Bsep）在上升/堆积细胞中表达良好，并且它们的表达与肝细胞核因子4 α的表达相关。虽然小SI-Is不表达Oatps和Mrp 2，但表达Mrp 3，上升/堆积的SH分别在窦状隙和BC膜中表达Oatp 1和2以及Mrp 2蛋白。另一方面，乳腺癌耐药蛋白（Bcrp）和Mrp 3的表达下降，随着SH成熟。通过Oatps和Mrp 2转运的底物被分泌到BC中，并在BC和囊样结构中积累。分泌的底物可以有效地恢复从BC重建的SH来自正常大鼠，但不是从Mrp 2缺陷大鼠。总之，SHs可以在培养中重建表达功能性有机阴离子转运蛋白的肝类器官。该培养体系可用于药物代谢和排泄机制的研究。J.细胞。104：68-81，2008. (C)2007 Wiley-Liss，Inc.
Small hepatocytes (SHs) are hepatic progenitor cells with hepatic characteristics. They can proliferate to form colonies in culture and change their morphology from flat to rising/piled-up with bile canaliculi (BC), which results in maturation. In this study, we examined whether SHs could express hepatic transporters with polarity, whether the transporters could transport organic anion substrates into BC, and whether the secreted substances could be recovered from BC. Immunocytochemistry and RT-PCR were carried out. [H-3]-labeled estrogen derivatives were used to measure the functions of the transporlers in SHs isolated from normal and multidrug resistance-associated protein (Mrp) 2-deficient rats. The results showed that organic anion-transporting proteins (Oatps) 1 and 2, Na+-dependent taurocholate cotransporting polypeptide (Ntcp), Mrp2, and bile-salt export pump (Bsep) were well expressed in rising/piled-up cells and that their expression was correlated to that of hepatocyte nuclear factor 4 alpha. Although small SI-Is expressed not Oatps and Mrp2 but Mrp3, rising/piled-up SHs expressed Oatp1 and 2 and Mrp2 proteins in the sinusoidal and BC membranes, respectively. On the other hand, breast cancer resistant protein (Bcrp) and Mrp3 expression decreased as SHs matured. The substrate transported via Oatps and Mrp2 was secreted into BC and it accumulated in both BC and cyst-like structures. The secreted substrate could be efficiently recovered from BC reconstructed by SHs derived from a normal rat, but not from an Mrp2-deficient rat. In conclusion, SHs can reconstitute hepatic organoids expressing functional organic anion transporters in culture. This culture system may be useful to analyze the metabolism and excretion mechanisms of drugs. J. Cell. Biochem. 104: 68-81, 2008. (C) 2007 Wiley-Liss, Inc.