Differentiation and enrichment of hepatocyte-like cells from human embryonic stem cells in vitro and in vivo
Differentiation and enrichment of hepatocyte-like cells from human embryonic stem cells in vitro and in vivo
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DOI：
10.1634/stemcells.2007-0291
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发表时间：
2007-01-01
期刊：
影响因子：
5.2
通讯作者：
Zerna, Mark A.
中科院分区：
文献类型：
--
作者：
Duan, Yuyou;Catana, Andreea;Zerna, Mark A.
人胚胎干细胞（hESC）可能为功能性肝细胞提供细胞来源。本研究的目的是从 hESC 中建立一种可行的人肝细胞样细胞系，可用于基于细胞的治疗。通过转导含有由α1-抗胰蛋白酶启动子驱动的绿色荧光蛋白（GFP）基因的慢病毒载体来富集分化的hESC； GFP基因在定型肝细胞祖细胞和肝细胞中表达。通过激光显微切割和压力弹射纯化GFP+hESC。此外，将用慢病毒三重融合载体转导的分化hESC移植到NOD-SCID小鼠中，并通过电荷耦合装置相机评估肝脏中荧光素酶诱导的生物发光。 GFP+hESC表达了大量肝脏特异性基因，并且通过纯化GFP+hESC显着提高了这些基因的表达水平；我们的结果表明，纯化的分化hESC表达接近生理水平的肝脏特异性基因，并且具有与原代人肝细胞相当的肝脏特异性功能。分化的 hESC 存活并移植到小鼠肝脏中，移植后 3 周在移植的小鼠肝脏和血清中检测到人类肝脏特异性 mRNA 和蛋白质种类。这是首次在动物模型血清中检测到由hESC衍生的肝细胞产生的人白蛋白。这也标志着分化型hESC在动物肝脏中的首次成功移植，以及hESC在肝脏中的首次生物发光成像。这项研究是从 hESC 建立可行的肝细胞样细胞系的第一步。
Human embryonic stem cells (hESC) may provide a cell source for functional hepatocytes. The aim of this study is to establish a viable human hepatocyte-like cell line from hESC that can be used for cell-based therapies. The differentiated hESC were enriched by transducing with a lentivirus vector containing the green fluorescent protein (GFP) gene driven by the alpha 1-antitrypsin promoter; the GFP gene is expressed in committed hepatocyte progenitors and hepatocytes. GFP+hESC were purified by laser microdissection and pressure catapulting. In addition, differentiated hESC that were transduced with a lentivirus triple-fusion vector were transplanted into NOD-SCID mice, and the luciferase-induced bioluminescence in the livers was evaluated by a charge-coupled device camera. GFP+hESC expressed a large series of liver-specific genes, and expression levels of these genes were significantly improved by purifying GFP+hESC; our results demonstrated that purified differentiated hESC express nearly physiological levels of liver-specific genes and have liver-specific functions that are comparable to those of primary human hepatocytes. The differentiated hESC survived and engrafted in mouse livers, and human liver-specific mRNA and protein species were detected in the transplanted mouse liver and serum at 3 weeks after transplantation. This is the first time that human albumin generated by hESC-derived hepatocytes was detected in the serum of an animal model. This also represents the first successful transplantation of differentiated hESC in an animal liver and the first bioluminescence imaging of hESC in the liver. This study is an initial step in establishing a viable hepatocyte-like cell line from hESC.