Derivation, characterization and differentiation of human embryonic stem cells: comparing serum-containing versus serum-free media and evidence of germ cell differentiation
Derivation, characterization and differentiation of human embryonic stem cells: comparing serum-containing versus serum-free media and evidence of germ cell differentiation
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DOI：
10.1093/humrep/del412
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发表时间：
2007-02-01
影响因子：
6.1
通讯作者：
Ho, H-N.
中科院分区：
文献类型：
--
作者：
Chen, H. -F.;Kuo, H. -C.;Ho, H-N.
背景：本研究旨在建立人胚胎干细胞（hESC）系，以确定在含血清培养基和无血清培养基中维持时的差异，并测试其体外分化的潜力。方法：对 11 个捐赠的人类囊胚进行包括免疫手术在内的程序，以建立 hESC 系。使用无血清或含血清培养基对细胞系进行表征和维持，以比较其形态、Oct-4 表达、细胞凋亡和生长速度。通过属于三个胚胎胚层和生殖细胞谱系的基因的形态和表达来评估这些系的分化。结果：建立了三个 hESC 系，它们在两种培养基（含血清或无血清）中以相似的速度生长，但在含血清培养基中培养的 hESC 产生形态良好的集落和表达 Oct-4 的细胞的百分比显着更高。这些细胞系在直接贴壁培养中在体外自发分化为表达属于所有三个胚胎胚层的标记物和生殖细胞标记物的细胞，包括 c-Kit、STELLA、VASA 和生长分化因子 9 (GDF9)。结论：已经建立了三个具有台湾血统的hESC系，并且它们在有或没有胚状体（EB）形成的情况下均保留了体外分化潜力。数据支持 hESC 可能能够分化为生殖细胞，但还需要进一步证实。还建议在对先前在含血清培养基中衍生的 hESC 系实施无血清培养条件之前，需要采取逐步适应等策略。
BACKGROUND: This study was designed to establish human embryonic stem cell (hESC) lines, to identify the differences when maintained in serum-containing versus serum-free medium and to test their potential of in vitro differentiation. METHODS: Procedures including immunosurgery were performed on 11 donated human blastocysts to establish hESC lines. The cell lines were characterized and maintained using either serum-free or serum-containing media to compare their morphology, Oct-4 expression, apoptosis and growth speed. Differentiation of these lines was evaluated by the morphology and the expression of genes belonging to the three embryonic germ layers and the germ cell lineage. RESULTS: Three hESC lines were established, and they grew at similar speed in both media (serum-containing or serum-free), but hESC cultured in serum-containing medium yielded significantly higher percentages of morphologically good colonies and cells expressing Oct-4. These cell lines differentiated spontaneously in vitro into cells expressing markers belonging to all three embryonic germ layers and germ cell markers, including c-Kit, STELLA, VASA and growth differentiation factor 9 (GDF9), in directly adherent culture. CONCLUSIONS: Three hESC lines with Taiwanese ancestry have been established, and they retain the in vitro differentiation potential with or without embryoid body (EB) formation. The data support that hESC may be capable of differentiation into germ cells although further confirmation is needed. It is also suggested that strategies such as stepwise adaptation will be needed before implementing a serum-free culture condition for hESC lines that have previously been derived in a medium containing serum.