Characterization of Trophoblast and Extraembryonic Endoderm Cell Lineages Derived from Rat Preimplantation Embryos

Abstract

BackgroundPrevious attempts to isolate pluripotent cell lines from rat preimplantation embryo in mouse embryonic stem (ES) cell culture conditions (serum and LIF) were unsuccessful, however the resulting cells exhibited the expression of such traditional pluripotency markers as SSEA-1 and alkaline phosphatase. We addressed the question, which kind of cell lineages are produced from rat preimplantation embryo under “classical” mouse ES conditions.ResultsWe characterized two cell lines (C5 and B10) which were obtained from rat blastocysts in medium with serum and LIF. In the B10 cell line we found the expression of genes known to be expressed in trophoblast, Cdx-2, cytokeratin-7, and Hand-1. Also, B10 cells invaded the trophectodermal layer upon injection into rat blastocysts. In contrast to mouse Trophoblast Stem (TS) cells proliferation of B10 cells occurred independently of FGF4. Cells of the C5 line expressed traditional markers of extraembryonic-endoderm (XEN) cells, in particular, GATA-4, but also the pluripotency markers SSEA-1 and Oct-4. C5 cell proliferation exhibited dependence on LIF, which is not known to be required by mouse XEN cells.ConclusionsOur results confirm and extend previous findings about differences between blastocyst-derived cell lines of rat and mice. Our data show, that the B10 cell line represents a population of FGF4-independent rat TS-like cells. C5 cells show features that have recently become known as characteristic of rat XEN cells. Early passages of C5 and B10 cells contained both, TS and XEN cells. We speculate, that mechanisms maintaining self-renewal of cell lineages in rat preimplantation embryo and their in vitro counterparts, including ES, TS and XEN cells are different than in respective mouse lineages.

Activation of the PTHRP/adenylate cyclase pathway promotes differentiation of rat XEN cells into parietal endoderm, whereas Wnt/β-catenin signaling promotes differentiation into visceral endoderm

“…We have previously observed that the morphology of rat XEN cells varied depending on feeder versus plastic cultivation (Chuykin et al, 2010). When we cultured rat XEN cells at low density, we found that the majority of cells were rounded (Fig.…”

“…Rat XEN cells were derived as primary cultures from preimplantation embryos of different genetic backgrounds (Debeb et al, 2009;Chuykin et al, 2010). Previous studies showed that rat XEN cells expressed Oct4 and SSEA1 (Debeb et al, 2009;Chuykin et al, 2010) and contributed efficiently to both PE and VE development after blastocyst injections (Debeb et al, 2009).…”

“…Previous studies showed that rat XEN cells expressed Oct4 and SSEA1 (Debeb et al, 2009;Chuykin et al, 2010) and contributed efficiently to both PE and VE development after blastocyst injections (Debeb et al, 2009). Therefore, rat XEN cells were proposed to be at the earliest known stage observed in in vitro culture among the extraembryonic endoderm lineage (Debeb et al, 2009).…”

“…Rat XEN cell lines derived from different strains have been characterized previously (Chuykin et al, 2010). Experiments were performed on the rat XEN cell line F2 from Rattus norvegicus (strain Fisher 344) at passage 20-30.…”

“…Interestingly, although rat XEN cells exhibited similarities to the respective mouse lineages, they were found to be positive for Oct4, Rex1 and SSEA1, which are markers for undifferentiated cells (Debeb et al, 2009;Chuykin et al, 2010). After injection into the blastocyst, rat XEN cells contributed to both PE and VE formation, demonstrating a broader developmental potential compared to mouse XEN cells (Debeb et al, 2009).…”