小鼠胚胎干细胞中2C-like细胞研究进展

Review on 2C-like state of mouse embryonic stem cells

小鼠胚胎干细胞(embryonic stem cell, ESC)由小鼠囊胚的内细胞团体外培养获得.小鼠胚胎干细胞可贡献到胚胎的3个胚层,但缺乏形成胚外组织的能力,因此,小鼠胚胎干细胞被认为具有多能性(pluripotent),而不是全能性(totipotent).值得注意的是,小鼠胚胎干细胞中含有少量的亚细胞群(小于1%),其基因表达模式类似2-细胞期胚胎,这一类细胞称为2-细胞期胚胎样细胞(2-cell embryo like cell, 2C-like cell).在适当的体外培养条件下,小鼠胚胎干细胞和诱导多能性干细胞(induced pluripotent stem cell, iPSC)中均会自发产生少量的2C-like细胞. 2C-like细胞具有全能性和独特的发育特性,嵌合体实验证明其可同时贡献到胚胎和胚外组织中,因此2C-like细胞可以作为体外研究全能性干细胞的理想模型.本文就2C-like细胞的发现、分子调控机制、染色质特征、激活策略以及与体细胞重编程之间的关系等进行综述.

Mouse embryonic stem cells(ESCs) are derived from the inner cell mass of mouse blastocysts. Mouse ESCs are pluripotent and contribute widely to the formation of the three embryonic germ layers in chimeric mice but not to that of the extraembryonic trophectoderm. A small population of transient cells(less than 1%) has been identified occasionally in the culture of mouse ESCs, which has been shown to exhibit a gene expression profile similar to that of cells in 2-cell stage embryos(2C-like cells). 2C-like cells were found to be generated spontaneously from mouse ESCs and induced pluripotent stem cells(iPSCs) when they were cultured in the serum-supplemented with leukemia inhibitory factor(LIF) on feeder cells in vitro. 2C-like cells have been considered to be totipotent. There is evidence suggesting that 2C-like cells can effectively generate both embryonic and extra-embryonic tissues, and therefore, 2C-like cells can be potentially used as a model for in vitro totipotent stem cell research. However, only a small population of ESCs could transit to 2C-like state in the serum-containing culture medium. There are several 2C specific genes that express when ESCs enter 2C-like state, such as Zscan4, Mervl, Dux, Nelfa, Tdpoz1/2/4 and Tcstv1/3. Notably, 2C-like cells exhibit highly relaxed chromatin architecture, which is similar to that observed in 2-cell stage embryos. Genome-wide demethylation occurs upon transition of mouse ESCs into 2C-like state, including gene bodies, promoters, and repeat elements. The dynamics of the reprogramming process from ESCs to 2C-like cells are still unclear, which have certainly uncovered an intricate regulatory mechanism mediated by specific genes, proteins, as well as some metabolites and non-coding RNAs.Additionally, one of the important features of totipotency is that a single cell can form an entire organism. Therefore,totipotent stem cells form a valuable tool to study the early developmental processes in vitro and prove to be beneficial for regenerative medicine applications. For instance, the embryonic stem cells have been used to generate embryo-like structures. The artificial embryo-like structures not only facilitate the studies on the key events in early embryonic development, but also serve as a model for investigating the early developmental defects occurring in humans. Several previous studies revealed that ESCs at different totipotent states can arise in in vitro cultures, and they focused on identifying culture conditions that induce 2C-like states. For example, knockout serum replacement(KOSR) media promotes the induction of 2C-like cells from ESCs in in vitro culture, as indicated by the increased levels of ZSCAN4-positive cells. Additionally, the suppression of glycolysis mediated by 2-deoxy-d-glucose(2-DG) in serum plus LIF condition induces the 2C transcriptional programming in ESCs. Interestingly, the most recent research indicates that the spliceosomal repression in mouse ESCs drives the pluripotent-to-totipotent state transition. Using the splicing inhibitor pladienolide B, they achieved stable in vitro culture of totipotent ESCs comparable at the molecular levels with 2-and 4-cell blastomeres. In our recent study, we also demonstrated that retinoic acid induces NELFA-mediated 2C-like state of mouse embryonic stem cells, which is associated with epigenetic modifications and metabolic processes in chemically defined media. Here, we review the discovery, molecular regulatory mechanism, chromatin characteristics, activation strategies, and the relationship of 2C-like cells with somatic reprogramming.