The BCL2L1- and BCL2-overexpressing hESCs maintained their pluripotent state and could differentiate into embryoid bodies (EBs) expressing marker genes of all three germ layers ( Supplementary information, Figure S1C-S1F). We used a doxycycline (DOX)-inducible system for transient induction of human anti-apoptotic genes, BCL2L1 or BCL2, together with the DsRed reporter gene, in hESCs carrying a constitutively expressed reporter gene GFP ( Supplementary information, Figure S1A and S1B). We hence hypothesize that inhibition of apoptosis may enable the primed hESCs to form interspecies chimeras upon injection into mouse pre-implantation stage embryos. Recently, a non-stage-matching approach has been established to generate mouse-rat interspecies chimeras by inhibiting apoptosis of the primed ESCs 10. For example, primed hPSCs were engrafted into in vitro cultured gastrula-stage mouse embryos to form chimeras, and the primed hPSCs were also converted into a naïve-like state and then were injected into pre-implantation embryos to form mouse or pig chimeras 7, 8, 9. Several studies have reported the generation of human-animal interspecies chimeras using the stage-matching hESCs. Intriguingly, although derived from the ICM, the human ESCs (hESCs) are similar to the mouse EpiSCs in many aspects such as the morphology and self-renewal pathways, hence they are considered to represent the primed pluripotent state. Only naïve state ESCs can colonize the early embryos before the blastocyst stage to form adult chimeras and transmit to the germline, while the primed state EpiSCs can only integrate into the post-implantation embryos 5, 6. Although both being pluripotent, the ESCs and EpiSCs are considered to represent two distinct states of pluripotency, the naïve and primed pluripotent state. In rodents, embryonic stem cells (ESCs) and epiblast stem cells (EpiSCs) have been derived from inner cell mass (ICM) of the blastocyst and post-implantation epiblast respectively 3, 4. In addition, the interspecies chimeras formed between hPSCs and large animal embryos would open new avenues to generate human tissues and organs for regenerative medicine 2. Recently, interspecies chimera formation has been established in rodents by injection of rat pluripotent stem cells (PSCs) into mouse early embryos, and such a system provides an in vivo assay to test the developmental potential of human PSCs (hPSCs) 1.

Embreyo typ o postive