An in vitro expansion system for generation of human iPS cell-derived hepatic progenitor-like cells exhibiting a bipotent differentiation potential.
An in vitro expansion system for generation of human iPS cell-derived hepatic progenitor-like cells exhibiting a bipotent differentiation potential.
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DOI：
10.1371/journal.pone.0067541
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Kamiya A
中科院分区：
文献类型：
--
作者：
Yanagida A;Ito K;Chikada H;Nakauchi H;Kamiya A
肝母细胞是肝脏发育过程中的肝干/祖细胞，具有较高的增殖能力和分化为肝细胞和胆管细胞的能力。在再生医学和治疗严重肝病的药物筛选中，人诱导多能干细胞来源的成熟功能性肝细胞被认为是一种潜在的良好细胞来源。然而，在体外诱导这些细胞增殖是困难的。为了避免这个问题，我们使用一系列细胞因子在体外从人的iPS细胞中产生了肝祖细胞样细胞。利用CD13和CD133的抗体通过荧光激活细胞分选纯化高增殖的肝祖细胞，CD13和CD133是胚胎和成年小鼠肝脏中肝干细胞/祖细胞的已知细胞表面标志。纯化的CD13HighCD133+细胞与饲养层细胞一起低密度培养时，加入合适的生长因子和信号转导抑制剂(碱性磷酸酶抑制剂A-83-01和ROCK抑制剂Y-27632)，单个细胞可形成较大的集落。这些克隆由表达肝细胞标志基因(肝细胞核因子4α和α胎蛋白)和胆管标志基因(细胞角蛋白7)的两种类型的细胞组成，并在较长时间内继续增殖。在球体形成试验中，这些细胞被发现表达成熟肝功能所需的基因，如细胞色素P450酶和分泌白蛋白。当这些细胞在合适的细胞外基质凝胶中培养时，它们最终形成了具有上皮极性的胆管囊样结构，表明人iPS细胞来源的肝祖细胞具有双向分化能力。总而言之，这些数据表明，这种使用体外扩增系统的新程序不仅对肝脏再生有用，而且对于确定调节肝脏发育的分子机制也是有用的。
Hepatoblasts, hepatic stem/progenitor cells in liver development, have a high proliferative potential and the ability to differentiate into both hepatocytes and cholangiocytes. In regenerative medicine and drug screening for the treatment of severe liver diseases, human induced pluripotent stem (iPS) cell-derived mature functional hepatocytes are considered to be a potentially good cell source. However, induction of proliferation of these cells is difficult ex vivo. To circumvent this problem, we generated hepatic progenitor-like cells from human iPS cells using serial cytokine treatments in vitro. Highly proliferative hepatic progenitor-like cells were purified by fluorescence-activated cell sorting using antibodies against CD13 and CD133 that are known cell surface markers of hepatic stem/progenitor cells in fetal and adult mouse livers. When the purified CD13highCD133+ cells were cultured at a low density with feeder cells in the presence of suitable growth factors and signaling inhibitors (ALK inhibitor A-83-01 and ROCK inhibitor Y-27632), individual cells gave rise to relatively large colonies. These colonies consisted of two types of cells expressing hepatocytic marker genes (hepatocyte nuclear factor 4α and α-fetoprotein) and a cholangiocytic marker gene (cytokeratin 7), and continued to proliferate over long periods of time. In a spheroid formation assay, these cells were found to express genes required for mature liver function, such as cytochrome P450 enzymes, and secrete albumin. When these cells were cultured in a suitable extracellular matrix gel, they eventually formed a cholangiocytic cyst-like structure with epithelial polarity, suggesting that human iPS cell-derived hepatic progenitor-like cells have a bipotent differentiation ability. Collectively these data indicate that this novel procedure using an in vitro expansion system is useful for not only liver regeneration but also for the determination of molecular mechanisms that regulate liver development.
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影响因子：
6
作者：
Asahina, K;Sato, H;Yoshizato, K
通讯作者：
Yoshizato, K
影响因子：
56.9
作者：
Matsumoto, K;Yoshitomi, H;Zaret, KS
通讯作者：
Zaret, KS
影响因子：
29.4
作者：
Kamiya, Akihide;Kakinuma, Sei;Nakauchi, Hiromitsu
通讯作者：
Nakauchi, Hiromitsu
DOI：
10.1016/0006-291x(89)91609-4
发表时间：
1989-05-30
影响因子：
3.1
作者：
KOIDE, N;SHINJI, T;TSUJI, T
通讯作者：
TSUJI, T
DOI：
10.1073/pnas.0806522105
发表时间：
2008-08-26
影响因子：
11.1
作者：
Hay, David C.;Fletcher, Judy;Iredale, John P.
通讯作者：
Iredale, John P.