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胚胎干细胞是取自胚泡期胚胎内细胞团的多能干细胞。它们能够维持未分化状态，并且能够分化成内胚层，中胚层和外胚层所有三个胚层的细胞。尽管在胚胎干细胞的增殖和分化方面，人们已经作了很多研究，但是 光受体细胞的诱导依旧需要和发育中视网膜组织细胞共培养或者移植到发育的视网膜组织中。我们在这里介绍了一种通过确定的因子处理诱导干细胞分化为视网膜细胞的方法。这种方法可以使人或小鼠的胚胎干细胞倾向的分化为光受体细胞和视网膜色素上皮细胞。在我们的方法中，小鼠胚胎干细胞向光受体细胞和色素上皮细胞分化需要 28 天，而人的分化则分别需要 150 天和 120 天。这种分化系统和干细胞来源的视网膜细胞将促进视网膜疾病的移植治疗，药物检测和体外的疾病模型的发展。同样也将提高我们对中枢神经系统发育, 特别是眼发育的理解。
Stepwise differentiation of pluripotent stem cells into retinal cells
Fumitaka Osakada1,2, Hanako Ikeda1–3, Yoshiki Sasai2 & Masayo Takahashi1
1Laboratory for Retinal Regeneration, Center for Developmental Biology, RIKEN, Kobe, Japan. 2Organogenesis and Neurogenesis Group, Center for Developmental
Biology, RIKEN, Kobe, Japan. 3Shiga Medical Center for Adults, Shiga, Japan. Correspondence should be addressed to F.O. (osakada@cdb.riken.jp).
Published online 7 May 2009; doi:10.1038/nprot.2009.51
Embryonic stem (ES) cells are pluripotent cells derived from the inner cell mass of blastocyst-stage embryos. They can maintain an undifferentiated state indefinitely and can differentiate into derivatives of all three germ layers, namely ectoderm, endoderm and mesoderm. Although much progress has been made in the propagation and differentiation of ES cells, induction of photoreceptors has generally required coculture with or transplantation into developing retinal tissue. Here, we describe a protocol for generating retinal cells from ES cells by stepwise treatment with defined factors. This method preferentially induces photoreceptor and retinal pigment epithelium (RPE) cells from mouse and human ES cells. In our protocol, differentiation of RPE and photoreceptors from mouse ES cells requires 28 d and the differentiation of human ES cells into mature RPE and photoreceptors requires 120 and 150 d, respectively. This differentiation system and the resulting pluripotent stem cell-derived retinal cells will facilitate the development of transplantation therapies for retinal diseases, drug testing and in vitro disease modeling. It will also improve our understanding of the development of the central nervous system, especially the eye.
