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G-protein coupled receptor 64 is required for decidualization of endometrial stromal cells.

ABSTRACT: Although GPR64 has an important role for male fertility, its physiological roles in the female reproductive system are still unknown. In the present study, immunohistochemical analysis reveals a spatiotemporal expression of GPR64 in the uterus during early pregnancy. Observation of remarkable induction of GPR64 expression in uterine decidual cells points to its potential physiological significance on decidualization. The decidualization of uterine stromal cells is a key event in implantation. Progesterone (P4) signaling is crucial for the decidualization of the endometrial stromal cells for successful pregnancy. Therefore, we examined ovarian steroid hormone regulation of GPR64 expression in the murine uterus. P4 induced GPR64 expression in the epithelial and stromal cells of the uterus in ovariectomized wild-type mice, but not in PRKO mice. ChIP analysis confirmed that PGR proteins were recruited on progesterone response element of Gpr64 gene in the uteri of wild-type mice treated with P4. Furthermore, the expression of GPR64 was increased in human endometrial stromal cells (hESCs) during in vitro decidualization. Interestingly, small interfering RNA (siRNA)-mediated knockdown of GPR64 in hESCs remarkably reduced decidualization. These results suggest that Gpr64 has a crucial role in the decidualization of endometrial stromal cells.

SUBMITTER: Yoo JY

PROVIDER: S-EPMC5503986 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

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G-protein coupled receptor 64 is required for decidualization of endometrial stromal cells.

Yoo Jung-Yoon JY Ahn Jong Il JI Kim Tae Hoon TH Yu Sungryul S Ahn Ji Yeon JY Lim Jeong Mook JM Jeong Jae-Wook JW

Scientific reports 20170710 1

Although GPR64 has an important role for male fertility, its physiological roles in the female reproductive system are still unknown. In the present study, immunohistochemical analysis reveals a spatiotemporal expression of GPR64 in the uterus during early pregnancy. Observation of remarkable induction of GPR64 expression in uterine decidual cells points to its potential physiological significance on decidualization. The decidualization of uterine stromal cells is a key event in implantation. Pr ...[more]

PMID: 28694502

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Project description:Steroid receptor coactivator-3 (SRC-3; also known as NCOA3 or AIB1) is a member of the multifunctional p160/SRC family of coactivators, which also includes SRC-1 and SRC-2. Clinical and cell-based studies as well as investigations on mice have demonstrated pivotal roles for each SRC in numerous physiological and pathophysiological contexts, underscoring their functional pleiotropy. We previously demonstrated the critical involvement of SRC-2 in murine embryo implantation as well as in human endometrial stromal cell (HESC) decidualization, a cellular transformation process required for trophoblast invasion and ultimately placentation. We show here that, like SRC-2, SRC-3 is expressed in the epithelial and stromal cellular compartments of the human endometrium during the proliferative and secretory phase of the menstrual cycle as well as in cultured HESCs. We also found that SRC-3 depletion in cultured HESCs results in a significant attenuation in the induction of a wide-range of established biomarkers of decidualization, despite exposure of these cells to a deciduogenic stimulus and normal progesterone receptor expression. These molecular findings are supported at the cellular level by the inability of HESCs to morphologically transform from a stromal fibroblastoid cell to an epithelioid decidual cell when endogenous SRC-3 levels are markedly reduced. To identify genes, signaling pathways and networks that are controlled by SRC-3 and potentially important for hormone-dependent decidualization, we performed RNA-sequencing on HESCs in which SRC-3 levels were significantly reduced at the time of administering the deciduogenic stimulus. Comparing HESC controls with HESCs deficient in SRC-3, gene enrichment analysis of the differentially expressed gene set revealed an overrepresentation of genes involved in chromatin remodeling, cell proliferation/motility, and programmed cell death. These predictive bioanalytic results were confirmed by the demonstration that SRC-3 is required for the expansion, migratory and invasive activities of the HESC population, cellular properties that are required in vivo in the formation or functioning of the decidua. Collectively, our results support SRC-3 as an important coregulator in HESC decidualization. Since perturbation of normal homeostatic levels of SRC-3 is linked with common gynecological disorders diagnosed in reproductive age women, this endometrial coregulator-along with its new molecular targets described here-may open novel clinical avenues in the diagnosis and/or treatment of a non-receptive endometrium, particularly in patients presenting non-aneuploid early pregnancy loss.

Project description:Nucleotide-binding oligomerization domain (NACHT), leucine rich repeat (LRR) and pyrin domain (PYD) 7 containing protein, NLRP7, is a member of the NLR family which serves as innate immune sensors. Mutations and genetic variants of NLRP7 have been found in women with infertility associated conditions, such as recurrent hydatidiform mole, recurrent miscarriage, and preeclampsia. Decidualization of endometrial stromal cells is a hallmark of tissue remodeling to support embryo implantation and proper placental development. Given defective decidualization has been implicated in miscarriage as well as preeclampsia, we aimed to explore the link between the NLRP7 gene and decidualization.Endometrial samples obtained from pregnant women in the first trimester and non-pregnant women were used to study NLRP7 expression pattern. The human telomerase reverse transcriptase (hTERT)-immortalized human endometrial stromal cells (T-HESCs) were used to study the effect of NLRP7 on decidualization. Decidualization of T-HESCs was induced with 1 μM medroxyprogesterone acetate (MPA) and 0.5 mM 8-bromoadenosine 3':5'-cyclic monophosphate (8-Br-cAMP). siRNA was used to knock down NLRP7 while lentiviral vectors were used to overexpress NLRP7 in cells. NLRP7 expression was detected by immunofluorescence, qRT-PCR, and Western blotting. Decidualization markers, Insulin-like growth factor-binding protein 1 (IGFBP-1) and prolactin (PRL), were detected by qRT-PCR and ELISA. Nuclear translocation of NLRP7 was detected by the subcellular fractionation and confocal microscopy. The effect of NLRP7 on progesterone receptor (PR) activity was evaluated by a reporter system.NLRP7 was up-regulated in the decidual stromal cells of human first-trimester endometrium. After in vitro decidualization, T-HESCs presented with the swollen phenotype and increased expressions of IGFBP-1 and PRL. Knockdown or over-expression of NLRP7 reduced or enhanced the decidualization, respectively, according to the expression level of IGFBP-1. NLRP7 was found to translocate in the nucleus of decidualized T-HESCs and able to promote PR activity.NLRP7 was upregulated and translocated to the nucleus of the endometrial stromal cells in an in vitro decidualization model. Overexpressed NLRP7 promoted the IGFBP-1 expression and PR reporter activation. IGFBP-1 expression decreased with the knockdown of NLRP7. Therefore, we suggest that NLRP7 contributes to in vitro decidualization of endometrial stromal cells.

Project description:Infertility is constantly increasing in Canada, where 16% of Canadian couples are experiencing difficulty conceiving. It is thought that infertility can emanate from the dysregulated communication between the embryo and the maternal endometrium. In order to allow for this window of implantation to be open at the right moment, endometrial stromal cells proliferate and differentiate by a mechanism called decidualization. Intracellular and molecular mechanisms involved in the regulation of apoptosis and cell proliferation during decidualization of the endometrium are yet to be fully understood. It has been well demonstrated previously that Akt is importantly involved in cell survival and glycogen synthesis. Akt1, Akt2 and Akt3 isoforms have distinct physiological roles; this could also be the case during decidualization and pregnancy. The aim of this study is to investigate the regulation of PI3K/Akt pathway during the decidualization process of endometrial stromal cells. Expression of Akt isoforms, Akt activity (phospho-Akt), pIκB and substrates of Akt during decidualization were measured. To our knowledge, these results are the first to suggest a decrease in levels of Akt isoforms as well as a downregulation of Akt activity in the process of decidualization of human endometrial stromal cells. We also uncovered that decidualization induced nuclear localization of p65 through the phosphorylation of IκB, its inhibitory subunit; however, Par-4, a recently uncovered regulator of cell differentiation, was displaced from the nucleus upon decidualization. Our results also suggest that HIESC cells exhibit decreased motility during decidualization and that PI3K pathway inhibition could be involved in this process. Finally, we demonstrate that specific Akt isoforms present unique effects on the successful induction of decidualization. Further analyses will involve investigations to understand the precise signaling mechanisms by which this pathway is regulated.

Dataset Information

G-protein coupled receptor 64 is required for decidualization of endometrial stromal cells.

Publications

G-protein coupled receptor 64 is required for decidualization of endometrial stromal cells.

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