ANALYSIS OF MOUSE SECONDARY TROPHOBLAST INVASION

成功的植入对于哺乳动物的发育至关重要 胚胎这一过程涉及子宫壁的控制性侵入， 胚胎滋养层子宫间质的分化， 蜕膜被认为是入侵的屏障。假设是 测试的是，蜕膜细胞外基质发挥了重要作用， 通过影响蛋白酶的产生来控制入侵， 蛋白酶抑制剂，这种控制是由特定的 细胞表面的受体。这项工作将使用鼠标辅助 来自第7天的外胎盘锥（EPCs）的滋养层和子宫内膜的培养物， 蜕膜细胞这项工作将首先完成我们对细胞的分析 使用亲和层析的层粘连蛋白的表面反应物，和 免疫学和生物化学技术。非整合素系统将 在滋养层和蜕膜中进行了研究，并在蜕膜中进行了整合素研究。A 3- 三维凝胶侵袭试验将用于研究基质的影响 组成对滋养层侵袭的影响。 入侵将在一个 含有单一或多种基质的Vitrogen（I型胶原）凝胶 在植入部位发现的分子。将确定是否 侵入能力的差异与金属-， 由滋养层产生的丝氨酸或半胱氨酸蛋白酶。蛋白酶 将使用酶谱和抑制剂研究进行研究。这些实验 然后将集中在组织蛋白酶蛋白酶，并将确定结合位点 在调节基质分子内，并将蛋白酶调节与 相应的细胞表面受体。组织蛋白酶的上调， 侵袭性癌细胞使组织蛋白酶成为可能的候选者， 滋养层侵入中重要酶。组织蛋白酶B、D和L表达 将在体内和体外使用cDNA探针进行研究， 抗体和酶分析。 任何植入研究必须包括 蜕膜细胞的贡献。蜕膜被认为参与了 通过分泌蛋白酶抑制剂来控制入侵。 的 实验还将分析半胱氨酸蛋白酶抑制剂、组织蛋白酶 抑制剂，在体内和体外的蜕膜分化。两 cDNA探针和特异性抗体将用于研究胱抑素A和 C在转录和蛋白质水平的表达。这项工作将提供新的 了解成功植入的要求。失败的原因 过程代表了许多不孕夫妇的潜在缺陷， 肿瘤性疾病的发展，包括绒毛膜溃结 和绒毛膜癌。了解身体如何提供天然屏障 也可能导致更有效的治疗方法， 恶性疾病

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.