Implantation in humans and other mammals is a critical period during which high embryonic mortality rates occur. Prostaglandins (PGs) are key mediators regulating interactions between the reproductive tract and the conceptus (embryo with extraembryonic membranes). Although the significance of PGF2α as a regulator of corpus luteum regression is well established, the role of its high amounts in the uterine lumen in most mammals, regardless of placentation type, during the implantation period remains unresolved. We hypothesized that PGF2α acting as an embryonic signal mediator contributes to pregnancy establishment. Using a porcine model, we demonstrated that the conceptus and its signal (estradiol-17β) elevated endometrial expression of PGF2α receptor (PTGFR) invivo and in vitro. PTGFR protein was expressed mainly in luminal epithelial (LE) and glandular epithelial cells and blood vessels in the endometrium. PGF2α stimulated the MAPK1/3 pathway in endometrial LE cells that coincided with elevated gene expression and secretion of endometrial vascular endothelial growth factor A (VEGFA) protein. PGF2α–PTGFR and adenylyl cyclase signaling were involved in this process. PGF2α-induced VEGFA acting through its receptors stimulated proliferation of endometrial endothelial cells. Moreover, PGF2α elevated gene expression of biglycan, matrix metalloproteinase 9, transforming growth factor β3, and interleukin 1α in the endometrium. In summary, our study indicates that PGF2α participates in pregnancy establishment by promoting angiogenesis and expression of genes involved in tissue remodeling and conceptus–maternal interactions in porcine endometrium during early pregnancy.

The conceptus and endometrium secrete large amounts of prostaglandin E2 (PGE2) into the porcine uterine lumen during the periimplantation period. We hypothesized that PGE2 acts on conceptus/trophoblast cells through auto- and paracrine mechanisms. Real-time RT-PCR analysis revealed that PGE2 receptor (PTGER)2 mRNA was 14-fold greater in conceptuses/trophoblasts on days 14–25 (implantation and early placentation period) vs preimplantation day 10–13 conceptuses (P < .05). Similarly, expression of PTGER2 protein increased during implantation. Conceptus expression of PTGER4 mRNA and protein did not differ on days 10–19. PGE2 stimulated PTGER2 mRNA expression in day 15 trophoblast cells through PTGER2 receptor signaling. PGE2 elevated aromatase expression and estradiol-17β secretion by trophoblast cells. Moreover, PGE2 and the PTGER2 agonist, butaprost, increased the adhesive capacity of both human HTR-8/SVneo trophoblast and primary porcine trophoblast cells to extracellular matrix. This PGE2-induced alteration in trophoblast cell adhesion to extracellular matrix was abolished by incubation of these cells with AH6809 (PTGER2 antagonist), ITGAVB3-directed tetrapeptide arg-gly-asp-ser or integrin ITGAVB3 antibody. PGE2 stimulated adhesion of porcine trophoblast cells via the estrogen receptor and MEK/MAPK signaling pathway. PGE2 induced phosphorylation of MAPK1/MAPK3 through PTGER2 and up-regulated expression of cell adhesion proteins such as focal adhesion kinase and intercellular adhesion molecule-1. Our study indicates that elevated PGE2 in the periimplantation uterine lumen stimulates conceptus PTGER2 expression, which in turn promotes trophoblast adhesion via integrins, and synthesis and secretion of the porcine embryonic signal estradiol-17β. Moreover, the mechanism through which PGE2 increases trophoblast adhesion is not species specific because it is PTGER2- and integrin-dependent in both porcine and human trophoblast cells.

In order to characterize the transition of the corpora lutea (CL) from acquisition of luteolytic sensitivity to rescue of luteal function: (1) the expression of 38 factors associated with steroids, prostanoids and angiogenic systems, and (2) concentrations of the main hormones responsible for maintenance of CL function in cyclic and pregnant pigs were examined. Additionally, the effect of prostaglandin (PG) E₂ and F_2α on luteal function during the estrous cycle and pregnancy was evaluated in vitro. Significantly upregulated gene expression was revealed in CL collected on day 14 of the estrous cycle (CYP19A1, ESR2, PTGS2, HIF1A, EDN1) and on days 12-14 of pregnancy (SCARB1, PGRMC1, STAR, HSD3B1, NR5A1, PTGFR, PTGER4, VEGFA). Elevated concentrations of estradiol-17β and PGE₂ occurred in CL on days 12 and 14 of pregnancy, respectively, while an increased intraluteal PGF_2α content was noted on day 14 of the estrous cycle. Both PGs increased the synthesis of progesterone by cultured luteal slices obtained on day 14 of pregnancy, in contrast to the action of PGF_2α on the corresponding day of the estrous cycle. PGE₂ stimulated cAMP production via PTGER2 and PTGER4, while PGF_2α elevated the content of cAMP-response element-binding protein in cultured luteal slices from CL of pregnant pigs. In silico analysis showed that e.g., infiltration of lymphocytes and apoptosis of microvascular endothelium were activated in CL on day 12 of the estrous cycle vs. pregnancy. Summarizing, an abundance of E₂ and PGE₂ during pregnancy regulates specific pathways responsible for steroidogenesis, the prostanoid signaling system and angiogenesis during rescue from luteolysis in porcine CL.

The amount of prostaglandin F2α (PGF2α) in the uterine lumen increases during the window of implantation in many mammals, including humans. We hypothesized that PGF2α regulates processes related to human embryo implantation. The effect of PGF2α was studied using an in vitro model of human extravillous trophoblast (EVT) cell line (HTR-8/SVneo). Adhesion, proliferation, invasion and migration assays, zymography for metalloproteinases (MMP) activity, and gene/protein expression analyses were applied. Doses of 100 nM and/or 1 μM of PGF2α and fluprostenol were used. PGF2α receptor (PTGFR), MMP9 and MMP2 proteins in the human first trimester placenta were localized by immunohistochemistry and immunofluorescence. This study is the first reporting the expression of PTGFR protein in the first trimester placenta, as well as in HTR-8/SVneo cells. PGF2α and fluprostenol increased HTR-8/SVneo cell proliferation and adhesion to extracellular matrix protein (P < 0.05). This effect was abolished by mitogen activated protein kinases (MAPK) inhibitor. PGF2α induced phosphorylation of focal adhesion kinase and MAPK1/3 (P < 0.05). PGF2α increased mRNA content and protein activity of MMP9, and gene and protein expression of interleukin-6 (P < 0.05). EVT cell migration and invasiveness were stimulated by PGF2α (P < 0.05). The PGF2α effect on cell invasion was reduced by inhibitors of MMP2, MMP9 and mTOR. In all experiments, the stimulatory effects of PGF2α were diminished by using a PTGFR antagonist. Our findings suggest a significant role for PGF2α in mechanisms associated with implantation. PGF2α acting by PTGFR in HTR-8/SVneo cells stimulates their adhesion and proliferation through the MAPK signaling pathway and increases invasiveness inducing MMP proteolytic activity and mTOR signaling.