ANALYSIS OF MOUSE SECONDARY TROPHOBLAST INVASION

小鼠次级滋养层细胞侵袭分析

• 批准号: 2392364
• 负责人: BRUCE S BABIARZ
• 金额: $ 17.24万
• 依托单位: RUTGERS, THE STATE UNIV OF N.J.
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-09-01 至 1999-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2392364
• 关键词: affinity chromatography; antibody; antigen receptors; cathepsin B; cathepsin D; cell cell interaction; cell differentiation; collagen; cysteine endopeptidases; decidua; embryo /fetus tissue /cell culture; embryo implantation; genetic transcription; immunofluorescence technique; immunoprecipitation; integrins; laboratory mouse; laminin; molecular site; nucleic acid probes; protease inhibitor; protein biosynthesis; receptor binding; receptor expression; surface antigens; trophoblast; western blottings

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.

成功的植入对于哺乳动物的发展至关重要 胚胎。此过程涉及对子宫壁的受控入侵 胚胎滋养细胞。子宫基质与 Decidua被认为是入侵的障碍。假设是 测试的是，决定的细胞外基质在 通过影响蛋白酶的产生和 蛋白酶抑制剂，并且该对照是由特异性介导的 在细胞表面发现的受体。工作将使用鼠标次要 从第7天的牙术锥体（EPC）和子宫培养物从第7天开始 decIDual细胞。这项工作将首先完成我们对单元的分析 使用亲和色谱法的层粘连蛋白的表面矫正器， 免疫学和生化技术。非整合蛋白系统将是 在滋养细胞和Decidua中进行了研究，在Decidua中进行了整合素。 3- 尺寸凝胶浸润测定法将用于研究基质的效果 滋养细胞入侵的组成。 入侵将在 含有单基质或多个矩阵的基于胶原蛋白（I型I型胶原蛋白） 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差异有关 由滋养细胞产生的丝氨酸或半胱氨酸蛋白酶。蛋白酶 将使用Zymography和抑制剂研究研究。这些实验 然后将重点放在组织蛋白酶蛋白酶上，并定义结合位点 在调节基质分子中，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 侵入性癌细胞使组织蛋白酶可能候选 滋养细胞入侵中的重要酶。组织蛋白酶B，D和L表达 将使用特定的cDNA探针在体内和体外研究 抗体和酶测定。 任何植入研究都必须包括 决结细胞的贡献。被认为参与的Decidua 蛋白酶抑制剂分泌侵袭的控制。 这 实验还将分析胱抑素，组织蛋白酶的产生 抑制剂，在体内和体外分化过程中。两个都 cDNA探针和特定抗体将用于研究胱抑素A和 C在转录和蛋白质水平下。这项工作将提供新的 了解成功植入的要求。失败 过程代表许多不育夫妇中的潜在缺陷 肿瘤性疾病的发展，包括绒毛膜Destreunins 和绒毛膜瘤。了解身体如何提供自然障碍 进行组织入侵也可能导致更有效的疗法 恶性疾病。