Interspecies chimera between primate embryonic stem cells and mouse embryos: Monkey ESCs engraft into mouse embryos, but not post-implantation fetuses

Abstract

Unequivocal evidence for pluripotency in which embryonic stem cells contribute to chimeric offspring has yet to be demonstrated in human or nonhuman primates (NHPs). Here, rhesus and baboons ESCs were investigated in interspecific mouse chimera generated by aggregation or blastocyst injection. Aggregation chimera produced mouse blastocysts with GFP-nhpESCs at the inner cell mass (ICM), and embryo transfers (ETs) generated dimly-fluorescencing abnormal fetuses. Direct injection of GFP-nhpESCs into blastocysts produced normal non-GFP-fluorescencing fetuses. Injected chimera showed >70% loss of GFP-nhpESCs after 21 h culture. Outgrowths of all chimeric blastocysts established distinct but separate mouse- and NHP-ESC colonies. Extensive endogenous autofluorescence compromised anti-GFP detection and PCR analysis did not detect nhpESCs in fetuses. NhpESCs localize to the ICM in chimera and generate pregnancies. Because primate ESCs do not engraft post-implantation, and also because endogenous autofluorescence results in misleading positive signals, interspecific chimera assays for pluripotency with primate stem cells is unreliable with the currently available ESCs. Testing primate ESCs reprogrammed into even more naïve states in these inter-specific chimera assays will be an important future endeavor.

Interspecific in vitro assay for the chimera-forming ability of human pluripotent stem cells

“…Interspecies in vitro chimera assay for monkey cells Primate PSCs have characteristics similar to human PSCs: they do not form chimeras in allogeneic settings (Tachibana et al, 2012) or xenogeneic settings (Simerly et al, 2011). We applied this in vitro chimera assay to cynomolgus monkey (Macaca fascicularis) ESCs (Suemori et al, 2001) (line CMK6; supplementary material Fig.…”

Interordinal Chimera Formation Between Medaka and Zebrafish for Analyzing Stem Cell Differentiation

“…On the one hand, human ES cells can engraft into mouse blastocysts, where they proliferate and differentiate in vitro and persist in chimeric embryos that implant and develop in the uterus of pseudo-pregnant foster mice [29]. On the other hand, in interspecific chimeras similarly produced between monkey ES cells and mouse embryos, the donor ES cells engraft into mouse preimplantation embryos but not postimplantation fetuses [27]; and production of intergeneric chimeras between mice and voles did not succeed [45]. These studies suggest that the phylogenetic distance does not prevent ISCF in general.…”

“…Several other factors may also be responsible for donorhost compatibility. A difference in cell cycle length, for instance, has been proposed to preclude primate ES cells from participation in development after implantation [27]. In this case, the development speed also differs significantly between rodents and primates: Blastocysts develop within 3.5-4 days in mice but 5-6 days in humans and a week or more in monkeys.…”

“…Since the report of chimera formation between sheep and goats in 1949 [24], interspecific chimera formation (ISCF) has attracted considerable attention in developmental biology [25,26]. ISCF remains the only option for testing the pluripotency of stem cells in vivo in certain cases, where host embryos are rarely available as in nonhuman primates [27] or even not allowed for chimera experiments as in humans [28,29]. Recently, ISCF has become an emerging reproductive biotechnology for animal production in conservation biology [16,22].…”

Four decades of leading‐edge research in the reproductive and developmental sciences: The Infant Primate Research Laboratory at the University of Washington National Primate Research Center

“…This has been particularly useful for creating gene targeted disease models and testing cell transplant therapies in the mouse [Coffman et al, 1993;Huijbers et al, 2011;Kiel et al, 2008;Kolodziejska et al, 2008;Stillwell et al, 2009]. Significant emphasis is currently being placed on advancing the development of mammalian stem cell chimeras in the rat, pig and nonhuman primate to unequivocally demonstrate pluripotency and enhance our understanding of the biomedical utility of embryonic stem/induced pluripotent stem (ES/iPS) cell lines in these species [Simerly et al, 2011;Vassiliev et al, 2010;Yamamoto et al, 2012]. The physiological, morphological, and genetic similarities between the human and nonhuman primate make the nonhuman primate a singularly exceptional model system for some human diseases.…”