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Expression of Syncytin 1 (HERV-W), in the preimplantation human blastocyst, embryonic stem cells and trophoblast cells derived in vitro.

Human reproduction (Oxford, England) (2016-05-14)
Bikem Soygur, Harry Moore
ABSTRACT

As Syncytin 1 (human endogenous retrovirus (HERV-W)) is crucial for human embryo placentation is it expressed during preimplantation embryo development? Syncytin 1 was expressed mainly in trophoblast cells of the blastocyst particularly in cells underlying the inner cell mass (ICM). Syncytin 1 (along with HERV-FRD or Syncytin 2) is expressed in first-trimester placenta and required for cell-cell fusion to enable formation of syncytiotrophoblast and effective placentation. Preimplantation human embryos donated for research were cultured in vitro and protein expression of Syncytin 1 at the blastocyst stage of development investigated. Comparisons were made with protein (Syncytin 1) and mRNA (Syncytin 1 and 2) expression in human embryonic stem cells (hESCs) undergoing differentiation to trophoblast-like cells in vitro. In total, 10 blastocysts (×3 or 4 replicates) were analysed and 4 hESC lines. The study was terminated after consistent observations of embryos were made. Donated embryos were thawed and cultured to blastocyst, fixed with 4% w/v paraformaldehyde. Syncytin 1 protein expression was determined by immunofluorescent localisation and confocal microscopy. Additionally, hESCs were differentiated to trophoblast-like cells in standard and conditioned culture medium with growth factors (bone morphogenetic protein 4 (BMP4) or fibroblast growth factor 4 (FGF4) and assessed for mRNA (Syncytin 1 and 2) by quantitative polymerase chain reaction (qPCR) and protein expression by immunolocalization and western blot. Syncytin 1 was expressed in cytoplasm and on the cell surface of some trophoblast cells, and consistently the trophectoderm underlying the ICM of the blastocyst. There was weak but consistent expression of Syncytin 1 in cells on the periphery of the ICM also displaying pluripotency antibody marker (Tra-1-60). Three-dimensional reconstruction of confocal slice data provided good visualization of expression. The time course of expression of Syncytin 1 was replicated in hESCs differentiated in vitro confirming the embryo observations and providing statistically significant differences in protein and mRNA level (P= 0.002) and (P< 0.05), respectively. Culture of a limited number of embryos to blastocyst in vitro may not replicate the range and quality of development in situ. Probes (antibodies, PCR) were tested for specificity, but might have non-specific reactions. Syncytin expression is a prerequisite for embryo implantation and placentation. Understanding when expression first occurs during embryo development may be informative for understanding conditions of abnormal gestations such as pre-clampsia. The study was supported partly by an ERASMUS training grant and grant G0801059 from the Medical Research Council, U.K. There were no competing interests.

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SYBR® Green JumpStart Taq ReadyMix, for quantitative PCR, MgCI2 in buffer