Reduction of N-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions.
Reduction of N-glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions.
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DOI：
10.1371/journal.pone.0014099
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发表时间：
2010-11-23
期刊：
影响因子：
3.7
通讯作者：
Asashima M
中科院分区：
文献类型：
--
作者：
Hayashi Y;Chan T;Warashina M;Fukuda M;Ariizumi T;Okabayashi K;Takayama N;Otsu M;Eto K;Furue MK;Michiue T;Ohnuma K;Nakauchi H;Asashima M
人类诱导多能干细胞（hiPSCs）的成功建立增加了干细胞研究在生物学和医学上的应用可能性。特别是，hipsc是再生医学和药理学中很有前途的细胞来源。然而，hipsc应用的主要障碍之一是，在使用血清替代和小鼠胚胎成纤维细胞作为饲养细胞的常规培养条件下，存在被未定义病原体污染的风险。在这里，我们报告了一种简单的方法，在我们之前为人类胚胎干细胞开发的无饲料和无血清的定义培养条件下生成或培养hipsc。所述培养条件包括具有最少数量的所述组分的基础培养基，所述组分包括五种高度纯化的蛋白质和作为底物的纤维连接蛋白。首先，利用Yamanaka的四种因子和传统的未定义培养条件生成的hipsc适应已定义的培养条件。通过形态学评估，这些适应的细胞保留了自我更新的特性，自我更新标记蛋白的表达，标准生长速率，以及通过在体外和体内分化为所有三个主要生殖层的衍生物（免疫缺陷小鼠的畸胎瘤形成）来评估的多能性。此外，在规定的条件下培养的hiPSCs中，非人n -糖基神经氨酸（Neu5Gc）水平显著降低，Neu5Gc是人类iPS细胞培养中病原体污染的异种抗原指标。其次，我们通过重编程步骤在规定的培养条件下使用成人真皮成纤维细胞成功地生成了hipsc。经长时间热培养，生成的细胞具有自我更新和多能性，核型正常，Neu5Gc阴性。本研究表明，在确定的培养条件下进行世代或适应性培养可以消除未定义病原体带来的风险。利用成纤维细胞生成hipsc的成功将有利于临床应用。
The successful establishment of human induced pluripotent stem cells (hiPSCs) has increased the possible applications of stem cell research in biology and medicine. In particular, hiPSCs are a promising source of cells for regenerative medicine and pharmacology. However, one of the major obstacles to such uses for hiPSCs is the risk of contamination from undefined pathogens in conventional culture conditions that use serum replacement and mouse embryonic fibroblasts as feeder cells. Here we report a simple method for generating or culturing hiPSCs under feeder- and serum-free defined culture conditions that we developed previously for human embryonic stem cells. The defined culture condition comprises a basal medium with a minimal number of defined components including five highly purified proteins and fibronectin as a substrate. First, hiPSCs, which were generated using Yamanaka's four factors and conventional undefined culture conditions, adapted to the defined culture conditions. These adapted cells retained the property of self renewal as evaluated morphologically, the expression of self-renewal marker proteins, standard growth rates, and pluripotency as evaluated by differentiation into derivatives of all three primary germ layers in vitro and in vivo (teratoma formation in immunodeficient mice). Moreover, levels of nonhuman N-glycolylneuraminic acid (Neu5Gc), which is a xenoantigenic indicator of pathogen contamination in human iPS cell cultures, were markedly decreased in hiPSCs cultured under the defined conditions. Second, we successfully generated hiPSCs using adult dermal fibroblast under the defined culture conditions from the reprogramming step. For a long therm culture, the generated cells also had the property of self renewal and pluripotency, they carried a normal karyotype, and they were Neu5Gc negative. This study suggested that generation or adaption culturing under defined culture conditions can eliminate the risk posed by undefined pathogens. This success in generating hiPSCs using adult fibroblast would be beneficial for clinical application.
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DOI：
10.1073/pnas.2131556100
发表时间：
2003-10-14
影响因子：
11.1
作者：
Tangvoranuntakul, P;Gagneux, P;Muchmore, E
通讯作者：
Muchmore, E
影响因子：
5.2
作者：
Hayashi, Yohei;Furue, Miho Kusuda;Asashima, Makoto
通讯作者：
Asashima, Makoto
影响因子：
2.1
作者：
Hayashi, Yohei;Furue, Miho Kusuda;Tanaka, Satoshi;Hirose, Michiko;Wakisaka, Noriko;Danno, Hiroki;Ohnuma, Kiyoshi;Oeda, Shiho;Aihara, Yuko;Shiota, Kunio;Ogura, Atsuo;Ishiura, Shoichi;Asashima, Makoto
通讯作者：
Asashima, Makoto
影响因子：
56.9
作者：
Yu, Junying;Vodyanik, Maxim A.;Thomson, James A.
通讯作者：
Thomson, James A.
影响因子：
4
作者：
Ross, Pablo Juan;Suhr, Steven Thomas;Bernardo Cibelli, Jose
通讯作者：
Bernardo Cibelli, Jose