Physiology and biotechnology of Embryonic stem cells

Our work aims to decipher cellular and molecular mechanisms controlling the establishment and the maintenance of pluripotency in reference and agronomic species including porcine, bovine and equine but also Aves as models of non-mammalian species. By developing and studying these pluripotent stem cells (PSCs), we articulate our research project around two main axes.

Axis 1: Derivation and characterization of pluripotent stem cells (PSCs) in different agronomic species:

We were pioneers in the isolation, in vitro establishment and characterization of chicken embryonic stem cells (ESC) (Pain et al, 1996; Lavial et al., 2007, Pain et al., 2018). This long term establishment is achieved using various culture media, which leads to different pluripotent stem cells that are then characterized at the molecular, epigenetic and developmental levels (Jean et al., 2013; Kress et al., 2016). Recently, we also demonstrated that the somatic reprogramming, defined as the process to get induced pluripotent stem cells (iPSCs) is possible but partially successful in avian species (Fuet et al., 2018). In order to improve the derivation of pluripotent cells and the reprogramming, we develop reporter genes based on endogenous retroviral elements (ERV) to enrich cell populations according to their pluripotency status.

Primordial germ cells (PGC), which are closely related to ESCs, are specified differently in avian species compared to mammals. We also isolated avian PGCs, grew them in vitro in long term cultures and compared their transcriptomes (RNAseq) and epigenomes (ChIPseq) to the ESC ones. Advances in mastering large scale 3D culture of PGCs (Chen et al., 2018) as well as in establishing duck PGC culture system (Chen et al., 2019) allows us to compare these cultured germ cells with the ones found in the gonads.

Using either the canonical Yamanaka gene cocktail or new gene combinations (Baquerre et al., 2017, EP17305082.4, patent, Aurine et al., 2018) the somatic reprogramming approach was applied to various mammalian species including human, bovine, porcine, equine and bat derived cells. Those different reprogrammed cells were characterized by immunochemistry (cell markers) and biochemistry (alkaline phosphatase and telomerase activity) and compared with the starting primary cells as well as with the early embryos by RNAseq. All together those data provide a comparison basis for establishing the pluripotency networks in all those species with a phylogenetic perspective.

Axis2: Biotechnology of pluripotent cells:

PSCs are a unique source of cell plasticity to generate specific derivatives by controlling their differentiation process. By applying various protocols of differentiation to engage the pluripotent stem cells toward different lineages, a large spectrum of differentiated progenitors could be produced (Vautherot et al., 2013, N° FR1357346, patent, Couteaudier et al., 2015; Vautherot et al., 2017). Beside the fundamental research, for which obtaining and characterizing specific differentiated cells from pluripotent stem cells is a real challenge, one of the main interests of those pluripotent and iPS cells lies in their biotechnological potential. In particular, these cells and their differentiated derivatives are useful cellular substrates for viral and bacterial studies as well as vaccine production (Giothis et al., 2019)

2018Fuet A, Montillet G, Jean C, Aubel P, Kress C, Rival-Gervier S, Pain BNANOG Is Required for the Long-Term Establishment of Avian Somatic Reprogrammed CellsStem Cell Reports
2018Chen YC, Chang WC, Lin SP, Minami M, Jean C, Hayashi H, Rival-Gervier S, Kanaki T, Wu SC, Pain BThree-dimensional culture of chicken primordial germ cells (cPGCs) in defined media containing the functional polymer FP003PLoS One
2018Chang KW, Tseng YT, Chen YC, Yu CY, Liao HF, Chen YC, Tu YE, Wu SC, Liu IH, Pinskaya M, Morillon A, Pain B, Lin SPCorrection to: Stage-dependent piRNAs in chicken implicated roles in modulating male germ cell developmentBMC Genomics
2018Chang KW, Tseng YT, Chen YC, Yu CY, Liao HF, Chen YC, Tu YE, Wu SC, Liu IH, Pinskaya M, Morillon A, Pain B, Lin SPStage-dependent piRNAs in chicken implicated roles in modulating male germ cell developmentBMC Genomics
2018Pain B, Kress C, Rival-Gervier SPluripotency in avian speciesInt J Dev Biol
2018Stadnicka K, Sławińska A, Dunisławska A, Pain B, Bednarczyk MMolecular signatures of epithelial oviduct cells of a laying hen (Gallus gallus domesticus) and quail (Coturnix japonica)BMC Dev Biol
2018Pfeuty B, Kress C, Pain BNetwork Features and Dynamical Landscape of Naive and Primed PluripotencyBiophys J
2016Kress C, Montillet G, Jean C, Fuet A, Pain BChicken embryonic stem cells and primordial germ cells display different heterochromatic histone marks than their mammalian counterpartsEpigenetics Chromatin
2015Bachelard E, Raucci F, Montillet G, Pain BIdentification of side population cells in chicken embryonic gonadsTheriogenology
2015Raucci F, Fuet A, Pain BIn vitro generation and characterization of chicken long-term germ cells from different embryonic originsTheriogenology