Natural selection of human embryos: impaired decidualization of endometrium disables embryo-maternal interactions and causes recurrent pregnancy loss

BackgroundRecurrent pregnancy loss (RPL), defined as 3 or more consecutive miscarriages, is widely attributed either to repeated chromosomal instability in the conceptus or to uterine factors that are poorly defined. We tested the hypothesis that abnormal cyclic differentiation of endometrial stromal cells (ESCs) into specialized decidual cells predisposes to RPL, based on the observation that this process may not only be indispensable for placenta formation in pregnancy but also for embryo recognition and selection at time of implantation.Methodology/Principal FindingsAnalysis of mid-secretory endometrial biopsies demonstrated that RPL is associated with decreased expression of the decidual marker prolactin (PRL) but increased levels of prokineticin-1 (PROK1), a cytokine that promotes implantation. These in vivo findings were entirely recapitulated when ESCs were purified from patients with and without a history of RPL and decidualized in culture. In addition to attenuated PRL production and prolonged and enhanced PROK1 expression, RPL was further associated with a complete dysregulation of both markers upon treatment of ESC cultures with human chorionic gonadotropin, a glycoprotein hormone abundantly expressed by the implanting embryo. We postulated that impaired embryo recognition and selection would clinically be associated with increased fecundity, defined by short time-to-pregnancy (TTP) intervals. Woman-based analysis of the mean and mode TTP in a cohort of 560 RPL patients showed that 40% can be considered “superfertile”, defined by a mean TTP of 3 months or less.ConclusionsImpaired cyclic decidualization of the endometrium facilitates implantation yet predisposes to subsequent pregnancy failure by disabling natural embryo selection and by disrupting the maternal responses to embryonic signals. These findings suggest a novel pathological pathway that unifies maternal and embryonic causes of RPL.<br/

Natural selection of human embryos: decidualizing endometrial stromal cells serve as sensors of embryo quality upon implantation

BackgroundPregnancy is widely viewed as dependent upon an intimate dialogue, mediated by locally secreted factors between a developmentally competent embryo and a receptive endometrium. Reproductive success in humans is however limited, largely because of the high prevalence of chromosomally abnormal preimplantation embryos. Moreover, the transient period of endometrial receptivity in humans uniquely coincides with differentiation of endometrial stromal cells (ESCs) into highly specialized decidual cells, which in the absence of pregnancy invariably triggers menstruation. The role of cyclic decidualization of the endometrium in the implantation process and the nature of the decidual cytokines and growth factors that mediate the crosstalk with the embryo are unknown.Methodology/Principal FindingsWe employed a human co-culture model, consisting of decidualizing ESCs and single hatched blastocysts, to identify the soluble factors involved in implantation. Over the 3-day co-culture period, approximately 75% of embryos arrested whereas the remainder showed normal development. The levels of 14 implantation factors secreted by the stromal cells were determined by multiplex immunoassay. Surprisingly, the presence of a developing embryo had no significant effect on decidual secretions, apart from a modest reduction in IL-5 levels. In contrast, arresting embryos triggered a strong response, characterized by selective inhibition of IL-1?, -6, -10, -17, -18, eotaxin, and HB-EGF secretion. Co-cultures were repeated with undifferentiated ESCs but none of the secreted cytokines were affected by the presence of a developing or arresting embryo.ConclusionsHuman ESCs become biosensors of embryo quality upon differentiation into decidual cells. In view of the high incidence of gross chromosomal errors in human preimplantation embryos, cyclic decidualization followed by menstrual shedding may represent a mechanism of natural embryo selection that limits maternal investment in developmentally impaired pregnancies

The human embryo does not elicit a secretory response in undifferentiated endometrium.

<p>Undifferentiated primary ESCs were co-cultured with embryos or not (control cultures, Con). Over the 72-hour co-culture period, 15 embryos arrested (Arr) whereas 6 continued to develop normally (Dev). Co-culture with either an arrested or developing embryo had no impact on the secreted levels of the indicated factors (<i>P</i>>0.05). The concentrations of IL-5, -12, -15, -17, -18, and eotaxin in culture supernatants of undifferentiated ESCs were below the level of detection.</p

Developmentally impaired human embryos inhibit the secretion of selective implantation modulators by decidualizing ESCs.

<p>Primary ESCs were first decidualized for 5 days and then co-cultured with human embryos or not (control cultures, Con). Over the 72-hour co-culture period, 30 embryos arrested (Arr) whereas 11 continued to develop normally (Dev). Analysis of the culture supernatants revealed that the presence of an arresting embryo inhibited the secretion of the indicated factors. The letters above the box plots indicate significant differences between groups. <i>P</i><0.01 for all comparisons except for IL-6 and IL-17 (<i>P</i><0.05).</p

Secreted decidual cytokines not regulated upon embryo co-culture.

<p>Primary ESCs were first decidualized for 5 days and then co-cultured with human embryos or not (control cultures, Con). Over the 72-hour co-culture period, 30 embryos arrested (Arr) whereas 11 continued to develop normally (Dev). Analysis of the culture supernatants revealed that the presence of an arresting or developing human embryo had no significant impact on the secretion of the indicated factors (<i>P</i>>0.05).</p

Human embryo development in co-culture.

<p>(<i>A</i>) Phase contrast images of decidualizing ESCs alone (control; left) or in the presence of an arrested (middle) or developing human embryo (right) (scale bar  = 100 µm). (<i>B</i>) Arrested embryos (Arr) express significantly less hCG than developing embryos (Dev). Secreted hCG levels were assayed after 72 hours of co-culturing human embryos and decidualizing ESCs (<i>P</i><0.01). (<i>C</i>) Formation of primitive endoderm in developing human embryos co-cultured with decidual ESCs. Optical cross-sections through a day 8 embryo, cultured first on a decidualizing ESC monolayer for 72 hours, demonstrates the presence of GATA-6 positive cells aligning predominantly to the blastocoelic surface of the inner cell mass, which corresponds to the location of the primitive endoderm (top panel). The lower panel represents z-projection of all the images in the stack from a top to bottom scan through the embryo (scale bar  = 50 µm).</p