Fukunaga Naoto, Teramura Takeshi, Onodera Yuta, Takehara Toshiyuki, Fukuda Kanji, Hosoi Yoshihiko
Department of Biology Oriented Science and Technology, Kinki University, Osaka, Japan.
Cell Reprogram. 2010 Aug;12(4):369-76. doi: 10.1089/cell.2009.0097.
Recently, several research groups have shown that germ cells can be produced in vitro from pluripotent embryonic stem cells (ESCs). In the mouse, live births of offspring using germ cells induced from ESCs in vitro have been reported. Furthermore, some efficient methods for inducing the useful number of germ cells from ESCs have also been developed. On the other hand, in primates, despite the appearances of germ cell-like cells including meiotic cells were observed by spontaneous differentiation or introducing transgenes, it has not been determined whether fully functional germ cells can be derived from ESCs. To elucidate the property for the germ cells induced from primate ESCs, specification of the promoting factors for the germ cell development and improving the efficiency of germ cell derivation are essential. Leukemia inhibitory factor (LIF) has been reported as one of the important factors for mouse primordial germ cell (PGC) survival in vitro. However, the effects of LIF on germ cell formation from pluripotent cells of primates have not been examined. The aim of this study is to determine whether LIF addition can improve in vitro germ cell production from cynomolgus monkey ESCs (cyESCs). After 8 days of differentiation, LIF added culture induced dome-shaped germ cell colonies as indicated by the intense expression of alkaline phosphatase activity (ALP). These cells also demonstrate high-level expression of the germ cell-marker VASA, OCT-4, and BLIMP-1, and show SSEA-1 expression that supports their early stage germ cell identity. Finally, we observed that adding LIF to differentiating cultures inhibited meiotic gene expressions and increased the percentage of ALP-positive cells, and demonstrate that the addition of LIF to differentiation media increases differentiation of early germ cells from the cyESCs.
最近，几个研究小组表明，可以从多能胚胎干细胞（ESC）体外产生生殖细胞。在小鼠中，已经报道了使用体外从ESC诱导的生殖细胞生出活的后代。此外，还开发了一些从ESC诱导产生有用数量生殖细胞的有效方法。另一方面，在灵长类动物中，尽管通过自发分化或导入转基因观察到了包括减数分裂细胞在内的生殖细胞样细胞的出现，但尚未确定是否能从ESC中获得功能完全正常的生殖细胞。为了阐明从灵长类ESC诱导产生的生殖细胞的特性，确定促进生殖细胞发育的因子并提高生殖细胞衍生效率至关重要。白血病抑制因子（LIF）已被报道为小鼠原始生殖细胞（PGC）体外存活的重要因子之一。然而，LIF对灵长类多能细胞形成生殖细胞的影响尚未得到研究。本研究的目的是确定添加LIF是否能提高食蟹猴ESC（cyESC）体外生殖细胞的产生。分化8天后，添加LIF的培养物诱导出圆顶形生殖细胞集落，碱性磷酸酶活性（ALP）的强烈表达表明了这一点。这些细胞还显示出生殖细胞标记物VASA、OCT-4和BLIMP-1的高水平表达，并显示出SSEA-1表达，支持它们早期生殖细胞的身份。最后，我们观察到在分化培养物中添加LIF会抑制减数分裂基因表达并增加ALP阳性细胞的百分比，并证明在分化培养基中添加LIF会增加cyESC早期生殖细胞的分化。
