Successful implantation is critical for the development of the mammalian embryo. This process involves a controlled invasion of the uterine wall by the embryonic trophoblast. The differentiation of the uterine stroma to decidua is thought to provide a barrier to invasion. The hypothesis to be tested is that the decidual extracellular matrix plays a major role in Controlling invasion by affecting the production of proteinases and proteinase inhibitors, and that this control is mediated by specific receptors found on the cell surface. The work will use mouse secondary trophoblast from day 7 ectoplacental cones (EPCs), and cultures of uterine decidual cells. The work will first complete our analyses of the cell surface rectors for laminin using affinity chromatography, and immunological and biochemical techniques. Non-integrin systems will be studied in trophoblast and decidua, and integrins in decidua. A 3- dimensional gel invasion assay will be used to study the effect of matrix composition on trophoblast invasion. Invasion will be analyzed in a Vitrogen (collagen type I) based gel containing single or multiple matrix molecules found within the implantation site. It will be determined if differences in invasive ability is related to differences in the metallo-, serine-, or cysteine proteinases produced by the trophoblast. Proteinase will be studied using zymography and inhibitor studies. These experiments will then focus on the cathepsin proteinases and will define binding sites within regulatory matrix molecules and correlate proteinase regulation to the corresponding cell surface receptor. The upregulation of cathepsins in invasive cancer cells makes the cathepsins likely candidates as an important enzyme in trophoblast invasion. Cathepsin B, D, and L expression will be studied in vivo and in vitro using cDNA probes, specific antibodies, and enzyme assays. Any study of implantation must include the contribution by the decidual cells. Decidua is thought to participate in the control of invasion by the secretion of proteinase inhibitors. The experiments will also analyze the production of cystatins, cathepsin inhibitors, during decidual differentiation in vivo and in vitro. Both cDNA probes and specific antibodies will be used to study cystatin A and C at the transcription and protein levels. This work will provide new insights on the requirements for successful implantation. Failures of this process represent the underlying defect in many infertile couples and in the development of neoplastic diseases, including chorionic destreunins and choriocarcinoma. Understanding how the body provides a natural barrier to tissue invasion could also lead to more effective therapies against malignant disease.
Showing the most recent 10 out of 11 publications
