Milk fat globule-EGF-factor 8 (MFG-E8), a protein linking apoptotic cells to phagocytes, is expressed in human endometrium during the window of implantation

ANAHÍ FRANCHI; SILVINA BOCCA; JESSICA H. ZARET; SERGIO OEHNINGER

Washington DC, USA

Congreso; American Society for Reproductive Medicine 63rd Annual Meeting; 2007

American Society for Reproductive Medicine

OBJECTIVE: In a previous study using microarray we demonstrated 49 genes up-regulated in the receptive vs. the pre-receptive human endometrium of natural cycles. MFG-E8, a novel gene not previously linked to implantation, was up-regulated 2.6-fold during the window of implantation. MFG-E8, a membrane glycoprotein containing two EGF domains, is expressed abundantly in mammary glands. MFG-E8 has been demonstrated to bind phosphatidylserine to link apoptotic cells to phagocytes via avb3 integrin. These results indicate that MFG-E8 is involved in the elimination of apoptotic epithelial cells during mammary gland involution. In addition, there is increasing evidence that apoptosis may be important during decidualization and implantation.We hypothesized that MFG-E8 and avb3 might be involved in endometrial reorganization during the receptive phase. We evaluated the expression of MFG-E8 and avb3 in human mid-secretory phase endometrium and in a well-differentiated steroid-responsive endometrial cell line (Ishikawa)in order to use this system as a model to study the function and regulationof the studied proteins. DESIGN: Protein and mRNA expression analysis in human endometriumand in Ishikawa cells. MATERIALS AND METHODS: Immunohistochemistry with mouseonoclonal antibodies was used to detect MFG-E8 and avb3 proteins in endometrial biopsies and Ishikawa cells.Western Blot (WB) with the same antibody was used to analyse MFG-E8 in Ishikawa cells lysate. MFG-E8 mRNAwas identified by reverse transcription-real time polymerase chain reaction (RT-PCR). RESULTS: MFG-E8 protein was strongly present in endometrial epithelial cells from biopsies as well as in Ishikawa cells. This was confirmed by the presence of a 45?50 kDa band on WB and an 88 bp band on RT-PCR. To the contrary, MFG-E8 was absent from stromal cells. avb3 integrin was detected in both endometrial compartments (though stronger in epithelial cells) as well as in Ishikawa cells. CONCLUSIONS: MFG-E8 and its ligand avb3 integrin were localized in epithelial cells in mid secretory-phase endometrial biopsies as well as in Ishikawa cells. These results show for the first time MFG-E8 expression in the endometrium suggesting its possible role in the endometrial reorganization during the receptive phase. In addition, MFG-E8 expression in Ishikawa cells suggests that these cells can be used as a model to study the MFG-E8 biological function and regulation during the endometrium remodeling events associated to the implantation process.