Demonstration of uterine receptivity in vitro by co-culture of rat epithelial cells and blastocyst

Abstract

Uterine receptivity is prerequisite for the attachment of the embryo to the uterine epithelium and involves a specialized polarity-dependent property of uterine epithelial (UE) cells. These UE cells, when polarized in culture, behave like cells in utero by exhibiting apico-basal polarity. In order to develop an implantation model in vitro, UE cells were polarized on extracellular matrix (ECM), and polarity was validated by response to estradiol-17beta administered exogenously. UE cells of pregnant rats at day-3 and day-4 post-coitum (p.c.) and of non-pregnant rats were cultured on bare and extracellular-matrix-coated petri dishes until confluency. Hatched blastocysts were transferred to the cultures, and adhesion was monitored every 24 h. Although blastocysts attached to UE cells that were taken from non-pregnant rats and from rats of day-3 p.c. and cultured on bare plastic, they failed to attach to these cells polarized on ECM. However, blastocysts attached firmly to UE cells that had been taken from rats of day-4 p.c. and polarized on ECM. Receptivity of UE cells taken from non-pregnant and pregnant (day-4 p.c.) rats was quantitated by flow cytometric estimation of cellular levels of beta3 integrin. The expression of beta3 integrin in UE cells from rats of day-4 p.c. was highly significant (P<0.01) when compared with its expression in UE cells from non-pregnant rats. The expression of beta3 integrin in UE cells of day-4 p.c. confirmed the receptivity of these cells to blastocyst implantation. Uterine receptivity was also validated in vivo by inducing the decidual cell reaction in rats ovariectomized on day-3 and day-4 p.c. Whereas remarkable deciduoma was noticed in the animals of day-4 p.c., it was absent in the animals of day-3 p.c., thereby indicating that the uterus was receptive on day-4 p.c. only. Thus, blastocysts do not attach to polarized UE cells that have been obtained from a non-receptive uterus. Attachment will occur only if the cells are obtained from a receptive uterus. UE cell receptivity is therefore essential for mimicking the process of implantation in vitro.

Coculture System That Mimics In Vivo Attachment Processes in Bovine Trophoblast Cells1

“…Over the past decades, various conceptus implantation models have been developed using endometrial epithelial cells (EECs) [2][3][4][5][6][7][8][9][10][11][12], stromal and decidual cells [13][14][15][16], or extracellular matrices (ECMs) [17][18][19]. A few studies demonstrated human or rodent blastocyst invasion into the epithelial and/or stromal cell layers [20][21][22], but most of these models could support only blastocyst attachment and outgrowth on the maternal components.…”

Differentiated Human Embryonic Stem Cells Enhance the In vitro and In vivo Developmental Potential of Mouse Preimplantation Embryos

“…In general, somatic cell co-culture is considered the method of choice for overcoming the in vitro developmental block of cultured mammalian embryos. As feeder cells for embryo co-culture, cumulus cells (Malekshah and Moghaddam, 2005), oviductal cells (Khatir et al, 2004), endometrial cells (Mercader et al, 2003), vero cells (Noh et al, 2006), and other somatic cells (Hotoya et al, 2006;Srinivasan et al, 2006) have been used. All of these cells are adult somatic cells unlike our young aged d-hES cells.…”

The effect of progesterone on oxytocin-stimulated intracellular Ca2+ mobilisation and prostaglandin secretion in porcine endometrium

“…Such an effect of P 4 was not determined in epithelial cells. It is known that the effect of this steroid on epithelial cell functions may vary between experiments due to in vitro culture conditions (Glasser and Mulholland 1993, Asselin et al 1996, Srinivasan et al 2006, female reproductive status (Blitek andZiecik 2004, Srinivasan et al 2006) or species differences (Glasser andMulholland 1993, Jamshidi et al 2007). …”