Wnt4 Acts Downstream of BMP2 and Functions via Canonical Beta-Catenin-Dependent Signaling Pathway to Regulate Human Endometrial Stromal Cell Differentiation.

Abstract

Differentiation of endometrial stromal cells into decidual cells is a prerequisite for successful embryo implantation. Our previous studies have shown that bone morphogenetic protein 2 (BMP2), a morphogen belonging to the TGFβ superfamily, is markedly induced in primary cultures of human endometrial stromal cells as they undergo differentiation in response to a hormonal cocktail containing progesterone, estrogen, and a cyclic AMP analog. Using loss-of-function and gain-of-function approaches we further showed that BMP2 plays a key role during human endometrial stromal cell differentiation. The present study was undertaken to identify the downstream targets of BMP2 in human endometrium during decidualization. Gene expression profiling experiments identified Wnt4 as a downstream target of BMP2 regulation during differentiation of human endometrial stromal cells. To determine the function of Wnt4 during this differentiation process, we employed siRNA targeted to its mRNA. Our study showed that siRNA-mediated down-regulation of Wnt4 expression in the stromal cells efficiently blocked their differentiation, as indicated by the loss of expression of well-known biomarkers of decidualization such as prolactin and insulin-like growth factor binding protein 1. Conversely, adenovirus-mediated overexpression of Wnt4 in human endometrial stromal cells markedly advanced the differentiation program. Interestingly, the stimulatory effect of Wnt4 was accompanied by the accumulation of an active form of β-catenin in the nuclei of decidualizing stromal cells, indicating the involvement of the canonical Wnt signaling pathway in the differentiation process. Furthermore, overexpression of Dickkopf-1 (Dkk1), a known inhibitor of the β-catenin-dependent canonical Wnt signaling pathway, led to a marked suppression of BMP2- and Wnt4-mediated decidualization. We also performed gene expression profiling to identify downstream targets of Wnt4 in human endometrial stromal cells. Our studies identified Foxo1, a member of the Forkhead/Winged helix family of transcription factors, as a downstream target of Wnt4 in decidualized human endometrial stromal cells. Collectively, these studies uncovered a unique pathway involving BMP2, Wnt4, and Foxo1 that operates in the human endometrial stromal cells and is critical for their transformation into decidual cells. (Supported by NIH grants to RNT, FJD, MKB, and ICB). (poster)