(206) Functions and mechanisms of uterine epithelial estrogen receptor α and progesterone receptor in regulating uterine immunity during early pregnancy

Xiaoqin Ye^1,2, Jonathan Matthew Hancock^1,2, Taylor Elijah Martin^1,2, Yuehuan Li², Jackson Sundgren², and Wendy Watford³

¹Interdisciplinary Toxicology Program; ²Department of Physiology and Pharmacology; ³ Department of Infectious Diseases, University of Georgia, Athens, GA 30602, USA.

 

Abstract Text:

The uterus has dynamic profiles of immune cells under physiological conditions (e.g., estrous cycle and pregnancy). Immune cell trafficking and transient endometrial inflammation in response to seminal fluid post-coitus are normal uterine processes. However, chronic endometrial inflammation, such as that related to endometritis, the use of intrauterine devices, and hydrosalpinx, is correlated with increased embryo implantation failure. Therefore, a controlled uterine immune environment is essential for the success of early pregnancy. In the uterine endometrium, the dominant immune cells are those in the innate immune system, such as neutrophils, macrophages, NK cells, and dendritic cells. T cells also have a significant presence in the endometrium, and regulatory T (Treg) cells are regulated by seminal plasma and estrogen. B cells are rare in the endometrium.

            Estrogen (E2)-estrogen receptor α (ERα/Esr1) signaling and progesterone (P4)-progesterone receptor (PR/Pgr) signaling are the master controls of uterine functions, including modulating uterine immune responses. It remains to be investigated how these two master controls temporally and coordinately regulate the uterine immune environment to prepare for the passage of the allogeneic sperm as well as for the development, transport, and implantation of the semi-allogeneic embryo (to the biological mother-to-be) or allogeneic embryo (to a surrogate woman) to the uterus. It has been demonstrated that deficiency of PR in the uterine epithelium of epiPR^-/- mice does not affect sperm migration, embryo development, or embryo transport in the uterus, but it impairs embryo attachment to the uterine luminal epithelium (LE). However, deficiency of ERα in the uterine epithelium of epiERα^-/- mice disrupts the early pregnancy events from sperm migration through the uterus to the attachment of the transferred embryo (embryos from natural pregnancy could not survive in the epiERα^-/- oviduct) to the uterus.

            To study the cellular effects, we employ immunohistochemistry and flow cytometry to demonstrate enhanced neutrophils in the day 0.5 post-coitum (D0.5) epiERα^-/- uterine lumen, especially in the un-liquified semen, as well as in the D0.5 epiERα^-/- uterus. Such phenomenon is not observed in the D0.5 epiPR^-/- uterus, nor in the D3.5 epiERα^-/- uterus or D3.5 epiPR^-/- uterus. These observations reveal the essential role of uterine epithelial ERα, but not PR, in maintaining the uterine innate immunity during post-coitum.  

            To study the molecular mechanisms, we isolate the LE cells from Esr1^fl/- (control) and epiERα^-/- mice, and Pgr^fl/- (control) and epiPR^-/- mice, on D0.5 and D3.5 for mRNA-seq. We have analyzed the mRNA-seq data from Esr1^fl/- LE and epiERα^-/- LE. Thousands of differentially expressed genes (DEGs) are revealed. Most notable is the dominant pathways related to innate immune responses from the upregulated DEGs in the D0.5 epiERα^-/- LE. Foremost among these is IL-1β signaling, a quintessential proinflammatory cytokine and potent neutrophil chemoattractant. On D3.5 (prior to embryo attachment ~D4.0 in mice), the innate immune dysregulation caused by uterine epithelial ERα-deficiency diminishes. We are obtaining mRNA-seq data from Pgr^fl/- LE and epiPR^-/- LE and will compare the DEGs with those from Esr1^fl/- LE and epiERα^-/- LE. We will determine immune-related genes in the LE regulated by uterine epithelial ERα and PR, respectively, and investigate how uterine epithelial ERα and PR coordinately regulate the uterine immune environment to support early pregnancy.