Differentiation and Characterization of Metabolically Functioning Hepatocytes from Human Embryonic Stem Cells
Differentiation and Characterization of Metabolically Functioning Hepatocytes from Human Embryonic Stem Cells
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DOI：
10.1002/stem.315
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发表时间：
2010-04-01
期刊：
影响因子：
5.2
通讯作者：
Zern, Mark A.
中科院分区：
文献类型：
--
作者：
Duan, Yuyou;Ma, Xiaochui;Zern, Mark A.
人类胚胎干细胞（hESC）可以为临床应用和药物开发提供功能性肝细胞的细胞来源。最初，通过CXCR 4、SOX 17和FOXA 2的表达评估，hESC群体富集为超过85%的定形内胚层（DE）。然后，我们成功地将DE转化为肝祖细胞，93%的细胞在9天内对甲胎蛋白呈阳性。白蛋白阳性细胞比例在分化后20-22天逐渐增加至90%。此外，我们的hESC衍生的肝细胞（hEH）开发了一个完整的生物转化系统，包括I相和II相代谢酶和III相转运蛋白。在调节代谢酶表达中起关键作用的核受体也在我们的hEH中表达。利用超高效液相色谱-串联质谱技术，我们在hEH中鉴定了药物丁呋洛尔的7条代谢途径，其中包括4条新报道的代谢途径，这些代谢途径与新鲜分离的人原代肝细胞（hPH）中的代谢途径相同。此外，四种药物的代谢结果表明，我们的hEH有能力以与hPH相当的水平代谢这些药物。总之，我们已经从hESC产生了相对同质的肝细胞群体，其似乎具有与原代肝细胞相当的完整代谢功能。这些结果代表了成熟肝细胞有效分化用于基于细胞的治疗以及药理学和毒理学研究的重要一步。干细胞2010;28：674-686
Human embryonic stem cells (hESCs) may provide a cell source for functional hepatocytes for clinical applications and drug development. Initially, the hESC population was enriched to be more than 85% definitive endoderm (DE) as assessed by the expression of CXCR4, SOX17, and FOXA2. We then successfully converted DE into hepatic progenitors with 93% of the cells being positive for alpha-feto protein within 9 days. The percentage of albumin positive cells gradually increased to 90% at days 20-22 after differentiation. Moreover, our hESC-derived hepatocytes (hEH) developed a complete biotransformation system including phase I and II metabolizing enyzmes and phase III transporters. Nuclear receptors, which are critical in regulating the expression of metabolizing enzymes, were also expressed by our hEH. Using ultraperformance liquid chromatography-tandem mass spectrometry technology, we identified seven metabolic pathways of the drug bufuralol including four newly-reported ones in our hEH, which are the same as those in freshly isolated human primary hepatocytes (hPH). In addition, the results of the metabolism of four drugs indicate that our hEH have the capacity to metabolize these drugs at levels that are comparable to hPH. In conclusion, we have generated a relatively homogenous population of hepatocytes from hESCs, which appear to have complete metabolic function that is comparable to primary liver cells. These results represent a significant step towards the efficient differentiation of mature hepatocytes for cell-based therapeutics as well as for pharmacology and toxicology studies. STEM CELLS 2010;28:674-686