摘要：

This study investigated the role of milk fat globule-epidermal growth factor-factor 8 (MFGE8) in TGF-β-induced epithelial– mesenchymal transition (EMT) of endometrial epithelial cells. These were in vitro studies using human endometrial epithelial cells and mouse blastocysts. We investigated the ability of TGF-β to induce EMT in endometrial epithelial cells (HEC-1A) by assessment of cytological phenotype (by light and atomic force microscopy), changes in expression of the markers of cell adhesion/differentiation Eand N-cadherin, and of the transcription factor Snail (by immunofluorescence and immunoblotting), and competence to support embryo attachment in a mouse blastocyst outgrowth assay. We also studied the effects of E-cadherin expression in cells transfected by retroviral shRNA vectors specifically silencing MFGE8. Results demonstrated that TGF-β induced EMT as demonstrated by phenotypic cell changes, by a switch of cadherin expression as well as by upregulation of the expression of the mesenchymal markers Snail and Vimentin. Upon MFGE8 knockdown, these processes were interfered with, suggesting that MFGE8 and TGF-β together may participate in regulation of EMT. This study demonstrated for the first time that endometrial MFGE8 modulates TGF-β-induced EMT in human endometrium cells. Reproduction (2016) 152 225–233 10.1530/REP-15-0585 Introduction Milk fat globule-epidermal growth factor-factor 8 (MFGE8), originally found in milk and mammary epithelial cells (Stubbs et al. 1990), is also known as lactadherin (Taylor et al. 1997) or SED1 (Ensslin & Shur 2003). Our group was the first to report the expression of MFGE8 in human endometrium and its upregulation during the window of implantation (Mirkin et al. 2005). Our data showed that MFGE8 protein is upregulated by prolactin in primary endometrial epithelial cell cultures, indicating paracrine epithelial–stromal cells interaction (Franchi et al. 2011). In addition, MFGE8 protein is highly expressed in human chorionic villi at all trimesters of gestation and in murine implantation sites (Bocca et al. 2012), and participates in trophoblast adhesion with its receptor integrin ανβ3 in an in vitro model of human implantation (Schmitz et al. 2014). We have also demonstrated that endometrial MFGE8 gene expression, as well as the expression of other inflammatory factors, such as IL-6 and IL-8, are significantly regulated by TNFα (Yu et al. 2014) and hCG (Sarhan A et al. 2013) respectively. These studies strongly suggested that endometrial MFGE8 performs an important role in physiological conditions during menstrual endometrium remodeling and implantation, and dysfunctions of its expression may be associated with endometrial pathological conditions. In extra-uterine tissues, this secreted phosphatidylserine binding protein, MFGE8, has been reported to have functions in apoptosis control, cell remodeling, neovascularization and immunomodulation (Morris et al. 1983, Silvestre et al. 2005). In human melanoma cells, MFGE8 promoted disease progression through coordination with integrin ανβ3 in tumor microenvironment; it also increased cell resistance to apoptosis, triggered epithelial– mesenchymal transition (EMT), and stimulated invasion (Jinushi et al. 2008). However, melanoma cells with MFGE8 deficiency showed the ability to attenuate Akt

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