# Pluripot Stem Cells for Transgenesis in the Rabbit: A Utopia?
## 1 Introduction  
Pluripotent stem cells (PSCs) were first derived from the inner cell mass (ICM) of mouse embryos in 1981 [1, 2]. These cells, named embryonic stem cells (ESCs), were shown to be able to colonize the epiblast of host blastocysts and, consequently, to produce chimeric mice in 1984 [3]. This ability of mouse ESCs (mESCs) has made it possible to develop transgenesis techniques in this species, facilitating substantial progress in functional genetics. Later on, PSCs were derived from the late epiblast of mouse post-implantation embryos [4, 5]. These cells, named epiblast stem cells (EpiSCs), display several different features from mESCs, especially with respect to their transcriptome and epigenome, and are notably not able to colonize host blastocysts. At the same time, mouse PSCs were obtained by reprogramming differentiated somatic cells by overexpression of four pluripotency factors, namely Oct4, Sox2, Klf4, and cMyc [6]. These cells, named induced pluripotent stem cells (iPSCs), can exhibit features of mESCs or EpiSCs depending on the medium used during their reprogramming [7, 8]. Therefore, mESCs and EpiSCs epitomize two states of pluripotency existingin vivo during early embryonic development--the naive state, corresponding to early epiblast cells from preimplantation embryos, and the primed state, corresponding to late epiblast cells from post-implantation embryos, respectively [9]. In vitro naive PSCs are sustained by the leukemia inhibitory factor (LIF)/gp130/STAT3 and bone morphogenetic protein 4 (BMP4)/ALK/ SMAD1-5-8 signaling pathways, whereas primed PSCs are supported by the fibroblast growth factor 2 (FGF2) and activin A/transforming growth factor beta (TGF\(\beta\))/SMAD2-3 signaling pathways [10]. To date, all PSC lines, ESCs and iPSCs, obtained in non-rodend mammalian species display features of primed pluripotency [11; 12]. Therefore, it seems that non-rodent PSCs are only able to stabilize in culture in the primed state of pluripotency [13]. For example, primate PSCs present a transcriptome very different from that of human early blastocyst epiblast cells [14], but is rather closer to that of post-implantation late blastocyst epiblast cells according to an analysis in monkeys [15]. The actual research challenge in the PSC field is to reprogram primed cells toward the naive state of pluripotency, in order to obtain and culture more genetically stable cells, which are easier to handle by single-cell dissociation [16; 17] and are more useful for cellular therapies or the production of disease models [18; 19]. The possibility to revert primed PSCs into naive PSCs was first demonstrated in mice by overexpressing pluripotency genes such as _Klf4_, _Nanog_, _Stat3_, _Tfcp21_, or _Prmd14_, either alone or in combination [20; 21; 22; 23; 24], and by strengthening the signaling pathways sustaining the naive state using inhibitors of mitogen-activated extracellular regulated kinase (MEK) and glycogen synthase kinase 3 beta (GSK3\(\beta\)) in a medium called 2 iLIF [25; 26]. Such strategies for reprogramming primed PSCs have been extensively studied for human PSCs and have produced naive-like cells with a heterogenous reconfiguration of their transcriptome and epigenome [27; 28; 29; 30] as well as a variable capacity to produce interspecies chimeras after microinjection into mouse blastocysts [31; 32]. These variations in the molecular and functional characteristics of primate PSCs show that embryonic cells could be stabilized in vitro at different stages along a continuum of pluripotency, the ends of which are epitomized by the naive and primed states, respectively [33]. Lagomorphs and primates share many similarities in their embryonic development [34], in particular in the timing of the embryonic genome activation at the 8/16-cell stages [35], the timing of the waves of DNA demethylation and methylation [36; 37], and the timing of the random inactivation of the second X chromosome [38].