ANALYSIS OF MOUSE SECONDARY TROPHOBLAST INVASION

成功的植入对哺乳动物的发育至关重要。 胚胎。这一过程涉及对子宫壁的受控侵袭 胚胎滋养层细胞。子宫间质分化为 蜕膜被认为是入侵的屏障。假设是 试验表明，蜕膜细胞外基质在 通过影响蛋白水解酶的产生来控制侵袭 蛋白水解酶抑制剂，而这种控制是由特定的 在细胞表面发现的受体。作品将使用鼠标辅助 7天外胎盘锥体滋养层细胞及子宫组织培养 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接收器，以及 免疫学和生化技术。非整合素系统将是 在滋养层和蜕膜中进行了研究，在蜕膜中进行了整合素的研究。A 3- 采用三维凝胶渗透实验研究基质的影响。 滋养层细胞侵袭的成分。入侵将在一个 含有单一或多个基质的维他命(I型胶原)凝胶 植入部位内发现的分子。将确定是否 侵袭能力的差异与金属-， 滋养层细胞产生的丝氨酸或半胱氨酸蛋白酶。水解酶 将通过酶谱和抑制剂研究进行研究。这些实验 然后将重点放在组织蛋白酶上，并将定义结合位置 在调节基质分子内，并将蛋白酶调节关联到 相应的细胞表面受体。血管内皮细胞中组织蛋白酶的上调 侵袭性癌细胞使组织蛋白有可能成为 滋养层细胞侵袭中的重要酶。组织蛋白酶B、D和L的表达 将使用cdna探针在体内和体外进行研究， 抗体和酶分析。任何关于植入的研究都必须包括 蜕膜细胞的贡献。蜕膜被认为参与了 通过分泌蛋白水解酶抑制物控制侵袭。这个 实验还将分析半胱氨酸蛋白酶、组织蛋白酶的产生 体内和体外蜕膜分化过程中的抑制剂。两者都有 将使用cDNA探针和特异性抗体来研究胱抑素A和 C在转录和蛋白水平上的表达。这项工作将提供新的 对成功植入的要求的见解。这方面的失败 这一过程代表了许多不育夫妇的潜在缺陷 包括绒毛膜促性腺激素在内的肿瘤性疾病的发展 和绒毛膜癌。了解身体是如何提供天然屏障的 对组织的侵袭也可能导致更有效的治疗 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.