Alternative Routes to Induce Naive Pluripotency in Human Embryonic Stem Cells
Alternative Routes to Induce Naive Pluripotency in Human Embryonic Stem Cells
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DOI：
10.1002/stem.2071
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发表时间：
2015-09-01
期刊：
影响因子：
5.2
通讯作者：
Heindryckx, Bjorn
中科院分区：
文献类型：
--
作者：
Duggal, Galbha;Warrier, Sharat;Heindryckx, Bjorn
人类胚胎干细胞(HESCs)与小鼠胚胎干细胞(MESCs)非常相似，表现出启动的多能性，不同于小鼠胚胎干细胞(MESCs)，它获得了一种幼稚的多能性状态。已经努力在hESCs中激发幼稚的多能性，以实现随后的无偏见的谱系特异性分化，这是启动的多能性hESCs所面临的常见难题，因为hESC系内和hESC系之间存在基因表达的异质性。这需要NANOG和KLF2等幼稚基因的异位表达，或者包含多种与多能性相关的因子。在此，我们报道了一种新的小分子和生长因子的组合(2I/LIF/碱性成纤维细胞生长因子1抗坏血酸1Forsklin)，有助于在hESCs和mESCs中快速诱导无转基因的幼稚多能性，这是以前没有显示过的。转化后的naive hESCs能在单细胞长期传代中存活，保持正常的核型，上调naive多能性基因，并表现出与naive mESCs相似的信号通路依赖性。此外，它们经历了全球DNA去甲基化，并显示出独特的长非编码RNA图谱。我们认为，在我们的介质中，通过PI3K/AKT/mTORC的成纤维细胞生长因子信号通路诱导了启动的hESCs向幼稚多能性的转化。总而言之，我们展示了一种在hESCs中捕获朴素多能性的替代途径，该途径快速、可重复性、支持朴素的mESC派生，并允许有效的分化。
Human embryonic stem cells (hESCs) closely resemble mouse epiblast stem cells exhibiting primed pluripotency unlike mouse ESCs (mESCs), which acquire a naive pluripotent state. Efforts have been made to trigger naive pluripotency in hESCs for subsequent unbiased lineage-specific differentiation, a common conundrum faced by primed pluripotent hESCs due to heterogeneity in gene expression existing within and between hESC lines. This required either ectopic expression of naive genes such as NANOG and KLF2 or inclusion of multiple pluripotency-associated factors. We report here a novel combination of small molecules and growth factors in culture medium (2i/LIF/basic fibroblast growth factor1Ascorbic Acid1Forskolin) facilitating rapid induction of transgene-free naive pluripotency in hESCs, as well as in mESCs, which has not been shown earlier. The converted naive hESCs survived long-term single-cell passaging, maintained a normal karyotype, upregulated naive pluripotency genes, and exhibited dependence on signaling pathways similar to naive mESCs. Moreover, they undergo global DNA demethylation and show a distinctive long noncoding RNA profile. We propose that in our medium, the FGF signaling pathway via PI3K/AKT/mTORC induced the conversion of primed hESCs toward naive pluripotency. Collectively, we demonstrate an alternate route to capture naive pluripotency in hESCs that is fast, reproducible, supports naive mESC derivation, and allows efficient differentiation.