Preimplantation human blastocysts release factors that differentially alter human endometrial epithelial cell adhesion and gene expression relative to IVF success

Abstract

Do human blastocysts which subsequently implant release factors that regulate endometrial epithelial cell gene expression and adhesion to facilitate endometrial receptivity? Blastocysts which subsequently implanted released factors that altered endometrial epithelial gene expression and facilitated endometrial adhesion while blastocysts that failed to implant did not. Human preimplantation blastocysts are thought to interact with the endometrium to facilitate implantation. Very little is known of the mechanisms by which this occurs and to our knowledge there is no information on whether human blastocysts facilitate blastocyst attachment to the endometrium. We used blastocyst-conditioned medium (BCM) from blastocysts that implanted (n = 28) and blastocysts that did not implant (n = 28) following IVF. Primary human endometrial epithelial cells (HEECs) (n = 3 experiments) were treated with BCM and the effect on gene expression and adhesion to trophoblast cells determined. We compared the protein production of selected genes in the endometrium of women with normal fertility (n = 40) and infertility (n = 6) during the receptive phase. We used real-time RT-PCR arrays containing 84 genes associated with the epithelial to mesenchymal transition. We validated selected genes by real-time RT-PCR (n = 3) and immunohistochemistry in the human endometrium (n = 46). Adhesion assays were performed using HEECs and a trophoblast cell line (n = 3). Blastocysts that implanted released factors that differentially altered mRNA levels for six genes (>1.5 fold) compared with blastocysts that did not implant. A cohort of genes was validated at the protein level: SPARC and Jagged1 were down-regulated (P < 0.01), while SNAI2 and TGF-B1 were up-regulated (P < 0.05) by implanted compared with non-implanted BCM. Jagged-1 (P < 0.05) and Snai-2 protein (P < 0.01) showed cyclical changes in the endometrium across the cycle, and Jagged-1 staining differed in women with normal fertility versus infertility (only) (P < 0.01). HEEC adhesion to a trophoblast cell line was increased after treatment with implanted BCM compared with untreated control (P < 0.05). This is an in vitro study and it would be beneficial to validate our findings using a physiological model, such as mouse. This new strategy has identified novel pathways that may be important for human preimplantation blastocyst-endometrial interactions and opens the possibility of examining and manipulating specific pathways to improve implantation and pregnancy success. This study was supported by the National Health and Medical Research Council of Australia (Fellowship support #550905, #611827) and project grants by Monash IVF, Australia. There are no conflicts of interest to be declared.