**Resetting Cell Fate by Epigenetic Reprogramming Resetting Cell Fate by Epigenetic Reprogramming**

DOI: 10.5772/intechopen.81015

Molly Pui Man Wong and Ray Kit Ng Molly Pui Man Wong and Ray Kit Ng

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.81015

#### **Abstract**

Epigenetic modifications and their regulations govern the identity of every cell type in an organism. Cell differentiation involves a switch in gene expression profile that is accompanied by heritable changes of epigenetic signatures in the differentiated cell type. Differentiation is generally not reversible, thereby conferring cell fate decisions once an altered epigenetic pattern is set. Nevertheless, attempts have been made to reverse a differentiation cell fate to a pluripotent state by various experimental approaches, such as somatic cell nuclear transfer, cell fusion and ectopic expression of defined transcription factors. The fundamental basis of all these strategies is to mediate epigenetic reprogramming, which allows a permanent and completed conversion of cell fate. A comprehensive understanding of the dynamic of epigenetic changes during cell differentiation would provide a more precise and efficient way of reprogramming cell fate. Here we summarize the epigenetic aspects of different reprogramming strategies and discuss the possible mechanisms underlying these epigenetic reprogramming events.

**Keywords:** epigenetic, reprogramming, somatic cell nuclear transfer, cell fusion, transcription factors, pluripotency, differentiation, cell fate

#### **1. Introduction**

Development is a complex process that involves a series of cell differentiation pathways starting from totipotent embryonic cells. According to Waddington's concept of epigenetic landscape, a cell has to interact with surrounding stimuli and respond by giving a phenotype which defines its identity during development [1]. Each cell experiences different inter/ intra-cellular signals and hence has its epigenetic signature of cell identity, which in turn directs its own specific gene expression pattern without alteration of DNA sequences (with the exception of the immunoglobulin genes in B and T cells). It is now clear that the diversity of cell type specific gene expression pattern is mediated by means of epigenetic mechanisms,

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

such as DNA methylation, covalent histone modifications and chromatin remodeling. Once a cell's identity is set, it is difficult to convert it to other unrelated lineages, thus leading to stable and irreversible differentiated cell states. Nevertheless, there are a few exceptions of cell fate conversion during embryo development and adult tissue/organ regeneration, e.g. vascular endothelium to smooth muscle cells [2], which involve changing a cell's epigenetic signature into another unrelated kind.

when compared to the normal ES cells derived from fertilized embryos. Another strategy of reprogramming is achieved by fusion of a differentiated cell with a pluripotent cell in order to generate a pluripotent-like tetraploid hybrid cell [3–5]. It has been proposed that cellular components, such as transcription factors, in the pluripotent cell are able to reprogram the differentiated cell nucleus. This idea aligns with the use of cell extracts from pluripotent cell types to revert differentiated cells into a pluripotent-like state [6, 7]. Presumably the cytoplasmic "reprogramming factors" from the pluripotent cells can be isolated and concentrated to achieve a higher reprogramming efficacy. A third strategy involves ectopic expression of defined transcription factors in somatic differentiated cells to generate induced pluripotent stem (iPS) cells. Delivery of the ectopic transcription factors can be achieved by viral approaches, such as the use of retrovirus, lentivirus, adeno-associated virus or Sendai virus, or by using episomal vesicles, or by direct mRNA or protein transfection. This technique has been successfully applied to reprogram a vast number of differentiated somatic cell types. Importantly, iPS cells can also be generated by using combinations of microRNAs (miRNAs) or small chemical molecules without the needs of ectopic expression of reprogramming factors [8]. The three reprogramming strategies show different reprogramming kinetics and efficiencies, which can be associated with the distinct epigenetic mechanisms in the erasure of somatic cell epigenetic signature and re-establishment of the pluripotent one. In this review, we focus on the dynamic changes of epigenetics mediated by different reprogramming strategies and how the modulation of epigenetic status improves the reprogramming efficiency.

Resetting Cell Fate by Epigenetic Reprogramming http://dx.doi.org/10.5772/intechopen.81015 167

SCNT was first done by Briggs and Kings in 1952, who transplanted a blastula nucleus into an enucleated egg of the amphibian *Rana pipiens* [9]. Few years later, Gurdon *et al.* succeeded by using differentiated *Xenopus* intestinal epithelial donor nuclei for SCNT [10]. In 1997, the first cloned mammal, Dolly the sheep, was generated [11], and since then, more than 23 other mammalian species have now been successfully cloned [12]. Normal development of nuclear transplanted (NT) embryos requires recapitulation of the gene expression profile that supports the embryogenesis process by the differentiated donor nucleus. This involves re-activation of pluripotency genes, in particular *Oct4*, *Nanog* and *Sox2*, and repression of somatic lineage genes. In fact, the efficiency of reprogramming by SCNT is generally very low and less than 1% of NT embryos can develop into normal adults [13–15]. The cloned newborns often suffer from developmental abnormalities owing to incomplete reprogramming. It has been observed that *Oct4* was aberrantly expressed in cloned mouse blastocysts derived from cumulus donor nuclei [16, 17]. Besides, continuous expression of other somatic donor marker genes was demonstrated in some *Xenopus* NT embryos [18]. Some imprinted genes in donor cells were found to be aberrantly expressed in cloned embryos, presumably owing to the incomplete epigenetic reprogramming of the regulatory regions of imprinting loci [19–21]. Dysregulation of imprinted genes, such as *Igf2*, *Igf2r*, *H19*, and *Xist*, in cloned embryos can lead to both fetal and placental overgrowth and result in embryonic lethality or an abnormal growth condition called "large offspring syndrome", which is commonly found in cloned

**2. Epigenetic reprogramming by SCNT**

Apart from a few exceptions of natural cell fate conversion events, different strategies have been developed aiming to reprogram differentiated somatic cell fate to a pluripotent state (**Figure 1**). A historical strategy of reprogramming is by somatic cell nuclear transfer (SCNT) experiments. SCNT involves transplantation of a cell nucleus into an enucleated egg/oocyte in order to generate a "cloned" animal with an equivalent genetic composition as the donor individual. It is possible to derive embryonic stem (ES) cells from nuclear transplanted (NT) embryos (ntES cells), which shows indistinguishable pluripotent gene expression profiles

**Figure 1.** Strategies of reprogramming cell fate. Differentiated cells can be reprogrammed to pluripotent state by somatic cell nuclear transfer (SCNT), cell fusion, and ectopic expression of defined transcription factors. SCNT involves transplantation of a single differentiated cell nucleus into an enucleated egg/oocyte, which develops as a nuclear transplanted (NT) embryo. Cell fusion involves artificial fusion of a differentiated and a pluripotent cell to form a tetraploid pluripotent-like cell. Defined transcription factors (Oct4, Sox2, Klf4, c-Myc) can be ectopically expressed in differentiated cells and convert them to induced pluripotent stem (iPS) cells.

when compared to the normal ES cells derived from fertilized embryos. Another strategy of reprogramming is achieved by fusion of a differentiated cell with a pluripotent cell in order to generate a pluripotent-like tetraploid hybrid cell [3–5]. It has been proposed that cellular components, such as transcription factors, in the pluripotent cell are able to reprogram the differentiated cell nucleus. This idea aligns with the use of cell extracts from pluripotent cell types to revert differentiated cells into a pluripotent-like state [6, 7]. Presumably the cytoplasmic "reprogramming factors" from the pluripotent cells can be isolated and concentrated to achieve a higher reprogramming efficacy. A third strategy involves ectopic expression of defined transcription factors in somatic differentiated cells to generate induced pluripotent stem (iPS) cells. Delivery of the ectopic transcription factors can be achieved by viral approaches, such as the use of retrovirus, lentivirus, adeno-associated virus or Sendai virus, or by using episomal vesicles, or by direct mRNA or protein transfection. This technique has been successfully applied to reprogram a vast number of differentiated somatic cell types. Importantly, iPS cells can also be generated by using combinations of microRNAs (miRNAs) or small chemical molecules without the needs of ectopic expression of reprogramming factors [8]. The three reprogramming strategies show different reprogramming kinetics and efficiencies, which can be associated with the distinct epigenetic mechanisms in the erasure of somatic cell epigenetic signature and re-establishment of the pluripotent one. In this review, we focus on the dynamic changes of epigenetics mediated by different reprogramming strategies and how the modulation of epigenetic status improves the reprogramming efficiency.
