A Panel of Embryonic Stem Cell Lines Reveals the Variety and Dynamic of Pluripotent States in Rabbits

Abstract

SummaryConventional rabbit embryonic stem cell (ESC) lines are derived from the inner cell mass (ICM) of pre-implantation embryos using methods and culture conditions that are established for primate ESCs. In this study, we explored the capacity of the rabbit ICM to give rise to ESC lines using conditions similar to those utilized to generate naive ESCs in mice. On single-cell dissociation and culture in fibroblast growth factor 2 (FGF2)-free, serum-supplemented medium, rabbit ICMs gave rise to ESC lines lacking the DNA-damage checkpoint in the G1 phase like mouse ESCs, and with a pluripotency gene expression profile closer to the rabbit ICM/epiblast profiles. These cell lines can be converted to FGF2-dependent ESCs after culture in conventional conditions. They can also colonize the rabbit pre-implantation embryo. These results indicate that rabbit epiblast cells can be coaxed toward different types of pluripotent stem cells and reveal the dynamics of pluripotent states in rabbit ESCs.

Beyond the mouse: non-rodent animal models for study of early mammalian development and biomedical research

“…Establishment of naïve pluripotent rabbit ESCs is of great interest because of their potential use in basic research, derivation of transgenic animals, and as a model in the establishment of stem-cell based therapies for human and animal medicine. Many research groups have reported derivation of rabbit ESCs or stemlike cells (Fang et al, 2006, Graves and Moreadith, 1993, Honda et al, 2008, Honda et al, 2009, Honda, 2013, Intawicha et al, 2009, Moreadith and Graves, 1992, Osteil et al, 2016, Zakhartchenko et al, 2011. Currently available rabbit ESCs are morphologically and molecularly different from mouse ESCs, however, they share many characteristics with human, primate and other non-rodent ESCs (du Puy et al, 2011, Tan et al, 2011.…”

“…To maintain their characteristics, rabbit and human ESCs require FGF2 and Activin/ Nodal, and not LIF (Ginis et al, 2004, Honda et al, 2009, Hsieh et al, 2011, while mouse ESC lines cultured without a feeder layer require LIF (Ginis et al, 2004. In a recent study, Osteil and colleagues meticulously analysed several rabbit ESC derivation conditions, including variations of dissociation methods and culture media, revealing a range of pluripotency states that cell lines acquire in vitro (Osteil et al, 2016). Similar to human ESCs and mouse EpiSCs, rabbit ESCs are rarely able to colonize the ICM in chimaeras and do not contribute to the germline.…”

“…The most successful rabbit ESC integration so far was shown by Zakharchenko and colleagues, who obtained one life born animal (out of 198 chimaeric embryos), of confirmed somatic chimaerism, which died 60 days after birth, and therefore germline transmission was never assessed (Zakhartchenko et al, 2011). Other researchers reported a rapid loss of chimaerism in rabbit preimplantation embryos after the introduction of rabbit ESCs or iPSCs (Osteil et al, 2013, Osteil et al, 2016, Tapponnier et al, 2017. Existing rabbit ESC lines are nonetheless capable of self-renewal over several passages, exhibit normal karyotype, and are well characterised by the gene expression profiling, teratoma formation and differentiation into derivatives of three germ layers in vitro (Honda et al, 2009, Osteil et al, 2013, Osteil et al, 2016.…”

“…Other researchers reported a rapid loss of chimaerism in rabbit preimplantation embryos after the introduction of rabbit ESCs or iPSCs (Osteil et al, 2013, Osteil et al, 2016, Tapponnier et al, 2017. Existing rabbit ESC lines are nonetheless capable of self-renewal over several passages, exhibit normal karyotype, and are well characterised by the gene expression profiling, teratoma formation and differentiation into derivatives of three germ layers in vitro (Honda et al, 2009, Osteil et al, 2013, Osteil et al, 2016. Several groups have also reported derivation of rabbit iPSCs, which share similar properties (Honda et al, 2010, Táncos et al, 2017, Tapponnier et al, 2017.…”

The quest for pluripotency: a comparative analysis across mammalian species

“…So far, other than human and mouse, many studies have concentrated on the derivation of PSCs from mammals such as mink, pig, bovine, rabbit, cat, dog, sheep, goat and horse (Sukoyan et al 1993, Li et al 2003, Telugu et al 2010reviewed in Ezashi et al 2016, Ogorevc et al 2016, Osteil et al 2016. In most of these cases, ESCs have either not been tested for or do not satisfy all the criteria for pluripotency (Fig.…”