EMILIN1 represents a major stromal element determining human trophoblast invasion of the uterine wall

Abstract

The detection of EMILIN1, a connective tissue glycoprotein associated with elastic fibers, at the level of the ectoplacental cone and trophoblast giant cells of developing mouse embryos (Braghetta et al., 2002) favored the idea of a structural as well as a functional role for this protein in the process of placentation. During the establishment of human placenta, a highly migratory subpopulation of extravillous trophoblasts (EVT), originating from anchoring chorionic villi, penetrate and invade the uterine wall. In this study we show that EMILIN1, produced by decidual stromal and smooth muscle uterine cells, is expressed in the stroma and in some instances as a gradient of increasing concentration in the perivascular region of modified vessels. This distribution pattern is consistent with the haptotactic directional migration observed in in vitro functional studies of freshly isolated EVT and of the immortalized HTR-8/SVneo cell line of trophoblasts. Function-blocking monoclonal antibodies against α4-integrin chain and against EMILIN1 as well as the use of EMILIN1-specific short interfering RNA confirmed that trophoblasts interact with EMILIN1 and/or its functional gC1q1 domain via α4β1 integrin. Finally, membrane type I-matrix metalloproteinase (MT1-MMP) and MMP-2 were upregulated in co-cultures of trophoblast cells and stromal cells, suggesting a contributing role in the haptotactic process towards EMILIN1.

An Alternative Role of C1q in Cell Migration and Tissue Remodeling: Contribution to Trophoblast Invasion and Placental Development

“…Adhesion and migration of CTBs were measured as previously reported (19). Briefly, CTBs (10 5 cells/100 ml) were labeled with the fluorescent dye FAST Dil (Molecular Probes, Invitrogen) and added to C1q-coated wells of a 96-well plate (Iwaki, Bibby Scientific Italia, Milan, Italy) for 30 min at 37˚C in an air/CO 2 incubator.…”

“…CTBs were plated at 37˚C on eight-chamber culture slides (BD Biosciences Discovery Labware, Milan, Italy) coated with C1q (10 mg/ml) or FN (10 mg/ ml) and left to adhere for various lengths of time between 15 min and 2 h. Postfixation, the cells were permeabilized and stained with N-(7-nitrobenz-2-oxa-1,3-diazol-4-y)-conjugated phallacidin (NBD-phallacidin) (Molecular Probes, Invitrogen) and mAb antipaxillin followed by Cy3-conjugated F(ab9) 2 goat anti-mouse IgG (1:300) as previously described (19).…”

“…4A, the adhesion of CTBs to C1q was inhibited almost completely by mAb 60.11, whereas the other mAb 74.5.2 to gC1qR and the rabbit IgG to cC1qR were ineffective. Based on our previous findings that trophoblast cells use a 4 b 1 integrin to interact with EMILIN1, a member of C1q family proteins sharing a gC1q signature domain (19,45), we examined the effect of neutralizing Abs to a 4 and b 1 integrin in the adhesion assay. Our results (Fig.…”

“…This protein acts as a bridging ligand for a 5 b 1 integrin expressed on EVTs and mediates their adhesion and migration at sites of anchoring villi and in the decidual tissue (17). Decidual stroma is also rich in laminins that surround the decidual cells as a distinct pericellular layer (18,19) and interact mainly with a 1 b 1 and a 6 b 1 on EVTs (20,21).…”

Adhesion Molecules in Human Trophoblast – A Review. II. Extravillous Trophoblast

“…Decidual ECM is also enriched in laminins, collagen IV [19,20] and fibronectin [19,21]. Interaction of CTB with the stromal and vascular wall fibrillar component EMILIN- 1 55 occurs by binding to integrin α4β1, leading to haptotactic migration and upregulation of matrix metalloproteinase (MMP) activity [22].…”