Irx3 Promotes Gap Junction Communication Between Uterine Stromal Cells to Regulate Vascularization during Embryo Implantation

Abstract

Spontaneous abortions have been reported to affect up to 43% of parous women, with over 20% occurring before pregnancy is clinically diagnosed. Establishment of pregnancy is critically dependent on proper embryo-uterine interactions at the time of implantation. Besides oocyte abnormalities, implantation failure is a major contributor to early pregnancy loss. Previously, we demonstrated that two members of the Iroquois homeobox transcription factor family, IRX3 and IRX5, exhibited distinct and dynamic expression profiles in the developing ovary to promote oocyte and follicle survival. Elimination of each gene independently caused subfertility, but with different breeding pattern outcomes. Irx3 KO (Irx3LacZ/LacZ) females produced fewer pups throughout their reproductive lifespan which could only be partially explained by poor oocyte quality. Thus, we hypothesized that IRX3 is also expressed in the uterus where it acts to establish functional embryo-uterine interactions to support pregnancy. To test this hypothesis, we harvested pregnant uteri from control and Irx3 KO females to evaluate IRX3 expression profiles and the integrity of embryo implantation sites. Our results indicate that IRX3 is expressed in the endometrial stromal cells of the pregnant uterus. Notably, of the days evaluated, IRX3 expression expanded into the endometrial stroma starting at day 4 of pregnancy (D4) with peak expression at D5-6, and then greatly diminished by D7. This pattern corresponds to the critical window for implantation and remodeling of the vasculature network in mice. Further, histology and immunohistochemistry at D7 showed that while embryos were able to attach to the uterus, implantation sites in Irx3 KO pregnant mice exhibited impaired vascularization. In addition, our results showed significantly diminished expression of decidualization markers and disruptions in GJA1 organization in the decidual bed. These data, taken together with previous reports focused on the ovary, suggest that IRX3 promotes fertility via at least two different mechanisms: 1) promoting competent oocytes and 2) facilitating functional embryo-uterine interactions during implantation. Future research aims to tease apart the roles for IRX3 in the oocyte versus the uterus and the mechanisms by which it promotes early embryo survival and a successful pregnancy outcome.