Rapid generation of mature hepatocyte-like cells from human induced pluripotent stem cells by an efficient three-step protocol.
Rapid generation of mature hepatocyte-like cells from human induced pluripotent stem cells by an efficient three-step protocol.
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DOI：
10.1002/hep.24790
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发表时间：
2012-04
期刊：
影响因子：
13.5
通讯作者：
Lee, Oscar K.
中科院分区：
文献类型：
--
作者：
Chen, Yu-Fan;Tseng, Chien-Yu;Wang, Hsei-Wei;Kuo, Hung-Chih;Yang, Vincent W.;Lee, Oscar K.
肝移植是终末期肝硬化和暴发性肝衰竭的唯一确定性治疗方法，但缺乏可用的供体肝脏是肝移植的主要障碍。最近，源自体成纤维细胞重编程的诱导多能干细胞 (iPSC) 已被证明与胚胎干 (ES) 细胞相似，因为它们具有多能特性，并且有可能在体外分化成所有细胞谱系，包括肝细胞。因此，iPSC 可作为广泛应用的有利细胞来源，包括药物毒性测试、细胞移植和患者特异性疾病建模。在这里，我们描述了一种高效、快速的三步方案，能够从人类 iPSC 中快速生成肝细胞样细胞。发生这种情况是因为内胚层诱导步骤允许更有效和确定的内胚层细胞形成。我们发现，在内胚层诱导步骤中，与不存在 HGF 时 (14.2%) 相比，肝细胞生长因子 (HGF) 与激活素 A 和 Wnt3a 协同作用，可将内胚层标志物 Foxa2（叉头盒 a2）的表达提高 39.3%。此外，iPSC 衍生的肝细胞与成熟肝细胞具有相似的基因表达谱。重要的是，肝细胞样细胞表现出细胞色素P450 3A4 (CYP3A4)酶活性、分泌尿素、摄取低密度脂蛋白(LDL)，并具有储存糖原的能力。此外，肝细胞样细胞在非肥胖糖尿病严重联合免疫缺陷小鼠模型中挽救了致命的暴发性肝衰竭。结论：我们建立了一种快速有效的分化方案，能够从人 iPSC 中产生功能性肝细胞样细胞。这可能为治疗肝脏疾病提供另一种选择。
Liver transplantation is the only definitive treatment for end-stage cirrhosis and fulminant liver failure, but the lack of available donor livers is a major obstacle to liver transplantation. Recently, induced pluripotent stem cells (iPSCs) derived from the reprogramming of somatic fibroblasts, have been shown to resemble embryonic stem (ES) cells in that they have pluripotent properties and the potential to differentiate into all cell lineages in vitro, including hepatocytes. Thus, iPSCs could serve as a favorable cell source for a wide range of applications, including drug toxicity testing, cell transplantation, and patient-specific disease modeling. Here, we describe an efficient and rapid three-step protocol that is able to rapidly generate hepatocyte-like cells from human iPSCs. This occurs because the endodermal induction step allows for more efficient and definitive endoderm cell formation. We show that hepatocyte growth factor (HGF), which synergizes with activin A and Wnt3a, elevates the expression of the endodermal marker Foxa2 (forkhead box a2) by 39.3% compared to when HGF is absent (14.2%) during the endodermal induction step. In addition, iPSC-derived hepatocytes had a similar gene expression profile to mature hepatocytes. Importantly, the hepatocyte-like cells exhibited cytochrome P450 3A4 (CYP3A4) enzyme activity, secreted urea, uptake of low-density lipoprotein (LDL), and possessed the ability to store glycogen. Moreover, the hepatocyte-like cells rescued lethal fulminant hepatic failure in a nonobese diabetic severe combined immunodeficient mouse model. Conclusion: We have established a rapid and efficient differentiation protocol that is able to generate functional hepatocyte-like cells from human iPSCs. This may offer an alternative option for treatment of liver diseases.
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