Progesterone-Regulated Endometrial Factors Controlling Implantation.

During implantation, the embryo attaches to the receptive uterine epithelium to initiate pregnancy. 1 – 6 Later the embryo invades into the underlying endometrial stroma as the stromal cells are transformed into decidual cells, which support embryonic growth and survival. During establishment and maintenance of pregnancy, the steroid hormone progesterone (P) plays a central role by profoundly influencing endometrial functions. In the preimplantation phase, P acts in concert with 17β estradiol (E) to orchestrate changes in the uterine epithelium rendering it competent for embryo implantation. 4 , 6 In mice, ovarian E on days 1 and 2 of pregnancy stimulates proliferation of uterine epithelium. In this E-dominated phase, the epithelium displays distinct columnar phenotype and cell-cell contacts via intracellular tight and adherens junctions. In response to rising P levels, starting on day 3 of pregnancy, the uterine epithelium ceases to proliferate and begins to undergo differentiation. The luminal epithelium, upon differentiation, undergoes structural remodeling involving disruption of tight and adherens junctions, which facilitates embryo attachment and invasion. 7 , 8 On day 4 of pregnancy, as the embryo attaches to the luminal epithelium, the subjacent fibroblastic stromal cells undergo differentiation into unique secretory ‘decidual’ cells. P is the primary driver of this differentiation process, termed decidualization, which is a prerequisite to successful implantation. 5 , 6 In this review, we will highlight some of the recent studies that elucidate the molecular mechanisms by which P regulates the early steps leading to the acquisition of uterine receptivity for implantation and successful establishment of pregnancy ( Figure 1 ). The physiological effects of P are mediated by the intracellular progesterone receptors (PRs). There are two isoforms of PR, PR-A and PR-B, generated from alternate transcripts arising from a single gene via different promoter usage. 9 Both isoforms have the same DNA binding and ligand binding domains, but PR-B possesses an additional transactivation domain in the amino terminal region. The PRs are members of the nuclear receptor superfamily. 10 , 11 In the presence of P, PR dissociates from the heat shock chaperone proteins, undergoes dimerization, and binds to target genes via direct interaction with discrete DNA response elements or via tethering interactions with other transcription factors. At the target gene, PR-A and PR-B recruit co-regulators to control transcription. 10 , 12 They also interact with transcription factors, such as FOS, JUN, CEBPβ and STAT3, which bind to neighboring genomic sites and modulate their transcriptional activities. 13 , 14 During human endometrial stromal decidualization, PR-B controls a substantially larger cistrome and transcriptome than PR-A. 13 PR-B also directly regulates the expression of PR-A. 13 Lydon et al 15 developed the PR knockout (PRKO) mouse, which lacks both PR-A and PR-B. This model provided important insights into the crucial role played by PR in mediating P-regulated responses during early pregnancy. The PRKO mice are infertile due to ovulation defect, since PR signaling in granulosa cells of the ovary is essential for conducting the luteinizing hormone-induced ovulatory program. 15 In addition to this ovarian phenotype, the PRKO mice also exhibited hyperplastic uteri that are non-receptive to embryo implantation. Analysis of these uteri revealed impaired decidualization and hypertrophied epithelium that is infiltrated by leucocytes. 15 Interestingly, genetic manipulations in mice leading to ablation of PR-A but not PR-B expression resulted in a uterine phenotype similar to PRKO, indicating that PR-A is the major isoform involved in the regulation of uterine receptivity and decidualization in the mouse. 16 However, in the human, PR-B plays a dominant role during decidualization. 13 While these mouse models established an essential role of PR signaling in the uterus during early pregnancy, it was important to decipher the molecular pathways regulated by this receptor during the pre- and post-implantation phases. The advent of global gene expression analyses, using DNA microarrays, allowed the delineation of these pathways. Employing the well-known PR antagonist RU486, Cheon et al ., 17 identified a large repertoire of PR-regulated genes that control uterine function during implantation. The DeMayo group also identified a number of genes induced in the uterine tissue upon acute and chronic P administration in non-pregnant ovariectomized mice. 18 Collectively, these studies revealed that P-dependent pathways play critical roles in modulating the functions of both uterine epithelial and stromal cells to establish an environment necessary for successful establishment of pregnancy. In this review, we highlight the cross-talk between the factors that function downstream of PR to (1) control epithelial proliferation necessary for the acquisition of uterine receptivity and (2) stimulate stromal cell differentiation necessary for the establishment of pregnancy ( Figure 2 ).

New evidence emerging from ChIP-sequencing and gene expression profiling analyses indicates that PR-B is a critical regulator of uterine angiogenesis, an essential feature of normal endometrial development and placentation. Angiogenesis-related genes found downstream of PR-B included vascular endothelial growth factor A ( VEGFA ), angiopoietin 2 ( ANGPT2 ), angiopoietin-like 4 ( ANGPTL4 ) and fibroblast growth factor 2 ( FGF2 ). Previous reports indicated that P regulates VEGFA, a key angiogenic factor in human endometrium. 56 Consistent with this report, Kaya et al., 13 identified several PR binding sites within a ± 200 kb window of the transcription start site of the VEGF gene, leading to the concept that these regions might be bound by PR-B, which then regulates VEGFA expression through a looping mechanism. ANGPT2, ANGPTL4 and FGF2 also contained consensus PR binding sites, suggesting direct regulation by PR. Future research will address how diverse angiogenic factors regulated by PR work in concert to facilitate proper endometrial neoangiogenesis to support the implanted embryo. Maternal immune tolerance of the semiallogeneic embryo requires a delicate local modulation of immune response within the pregnant uterus during implantation. P is known to play an important role in this immuno-modulation, although the mechanisms of its action remain unclear. 57 P exerts an anti-inflammatory effect during implantation by suppressing E-induced influx of neutrophils and macrophages into the stroma. The PRKO mice exhibit an increased number of neutrophils traversing the uterine epithelium into the lumen, causing increased inflammation and edema. 58 It has been proposed that HOXA10 may mediate the immunomodulatory effects of P. This was based on the observation that HOXA10-deficiency is associated with severe immunological dysregulation during implantation. 59 Mice lacking Hoxa10 exhibit abnormal polyclonal expansion of T-lymphocytes compared to wild type mice. 59 P is also reported to suppress the proinflammatory responses of uterine dendritic cells, which play a crucial role in implantation. 60 Treatment with RU486 prevented this P-mediated suppression of proinflammatory cytokines produced by dendritic cells. 61 Further studies are needed to delineate the PR-regulated pathways that modulate the innate immune system at the implantation site to ensure survival of the embryo.

Development of genetically engineered mouse models lacking PR and its target genes has provided a wealth of information regarding the roles of P-regulated pathways in endometrial receptivity and implantation. The identification of genome-wide binding sites and downstream gene networks of PR during endometrial differentiation have offered unique insights into the role of this receptor in human uterine biology. These studies have revealed that several P-regulated pathways involved in endometrial function during implantation are conserved in mice and humans. Dysregulated P signaling has been implicated not only in recurrent miscarriage and infertility, but also in other reproductive pathologies, such as endometriosis and endometrial cancer. Endometriosis, a prevalent gynecological disease, is considered a P resistant disease, although the mechanism of this resistance is not fully understood. 62 Similarly, endometrial hyperplasia and endometrial cancer are associated with compromised P signaling that fails to oppose E signaling. 63 Undoubtedly, further studies, involving a comprehensive mouse to human translational approach, will provide important mechanistic insights regarding the role of P-regulated factors in the context of endometrial dysfunctions associated with infertility, endometriosis and endometrial cancer.