Early pregnancy loss is a major public health problem in reproductive women. The oviduct/fallo- pian tube and uterus play essential roles in supporting early pregnancy events, such as fertiliza- tion, preimplantation embryo development, embryo transport, and embryo implantation. RhoA is a prominent member of the Rho family of GTPases with crucial cellular functions. It is highly ex- pressed in the gestation day 3.5 (preimplantation) mouse oviduct and uterus, especially the api- cal side of uterine luminal epithelium that is the initial site of embryo-maternal interactions. Es- sential cell-type specific in vivo functions of RhoA in different systems are being revealed in mouse genetic models. The in vivo functions of RhoA in the female reproductive system are largely unknown. To investigate in vivo function of RhoA in the oviduct and uterus for supporting early pregnancy, we generated a RhoA conditional knockout mouse model (RhoAd/d) by cross- ing RhoAf/f mice with Pgr-Cre mice. Progesterone (P4) deficiency in RhoAd/d females led us to identify a novel role of RhoA in steroidogenesis in the corpus luteum. RhoAd/d females have nor- mal ovulation but implantation failure partially due to P4 deficiency. Exogenous P4 treatment could rescue embryo implantation but with significantly reduced number of implantation sites in RhoAd/d females, suggesting impaired early pregnancy in RhoAd/d females. Multiple early preg- nancy events occur in the oviduct and uterus. It is unknown which event(s) that RhoA plays a role in. Our long-term goal is to understand molecular mechanisms in regulating early preg- nancy. The defined objective of this R03 application is to identify the impaired early pregnancy event(s) leading to reduced implantation sites in P4-treated RhoAd/d females. Based on our pre- liminary data, we formulate the central hypothesis that RhoA in the oviduct and uterus supports early pregnancy. To achieve the goal of this application, we will pursue two specific aims: deter- mine spatiotemporal expression of RhoA in oviduct and uterus during early pregnancy (Aim 1) and test the hypothesis that RhoA in oviduct and uterus supports early pregnancy events (Aim 2). To overcome P4 deficiency in RhoAd/d mice and to generate the most suitable in vivo control for RhoAd/d mice, unilateral ovary transplantation will be performed in control (RhoAf/f) mice with a RhoAd/d ovary and in RhoAd/d mice with a control ovary. Unilateral ovary transplantation, realtime PCR, and immunohistochemistry are among the approaches that will be employed. The proposed work is significant because understanding functions of RhoA in early pregnancy will give novel insights into factors that affect early pregnancy thus provide information for develop- ing diagnostic and therapeutic approaches to detect and treat early pregnancy loss.
The proposed research is relevant to public health because early pregnancy loss is a major public health problem in reproductive women. This proposal aims to determine the function(s) of a prominent GTPase RhoA in supporting early pregnancy using a conditional RhoA knockout mouse model RhoAd/d (RhoAf/fPRCre/+). Understanding the functions of RhoA in early pregnancy will give novel insights into factors that affect early pregnancy thus provide information for devel- oping diagnostic and therapeutic approaches to detect and treat early pregnancy loss.
