Hepatocyte growth factor activator and its inhibitors in mouse uterus

小鼠子宫内肝细胞生长因子激活剂及其抑制剂

• 批准号: 7239815
• 负责人: Xuan Zhang
• 金额: $ 7.35万
• 依托单位: UNIVERSITY OF KANSAS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-04-01 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7239815
• 关键词: Biological Assay; Carcinoma; Cell Proliferation; Cells; Cleaved cell; Coculture Techniques; Condition; Data; Development; Disease; Endometrial; Endopeptidases; Epithelial; Epithelial Cells; Estrogens; Experimental Models; Functional disorder; Goals; Growth Factor; Hepatocyte Growth Factor; Immunohistochemistry; In Situ Hybridization; In Vitro; Investigation; Lead; Mediating; Methods; Molecular; Mus; Ovarian; Ovarian Steroid Hormone; Peptide Hydrolases; Phosphorylation; Physiology; Play; Process; Progesterone; Protein Tyrosine Kinase; Proteins; Proto-Oncogene Protein c-met; Purpose; Recombinants; Regulation; Research; Reverse Transcriptase Polymerase Chain Reaction; Role; Serine Protease; Signal Transduction; Steroids; Stromal Cells; System; Testing; Thrombin; Time; Transcript; Uterine Diseases; Uterus; base; cell type; design; endometriosis; extracellular; hepatocyte growth factor activator; hormone regulation; human MET protein; immunocytochemistry; in vitro Model; in vivo; inhibitor/antagonist; neutralizing antibody; novel; pro-hepatocyte growth factor; protein expression; research study; spatiotemporal

DESCRIPTION (provided by applicant): Increasing evidence indicates that hepatocyte growth factor (HGF) plays an important role in uterine development and pathological conditions such as endometriosis and carcinoma. Proteolytic conversion of single chain HGF to active heterodimeric HGF in the extracellular milieu is a critical limiting step in HGF- mediated signal transduction, however, the mechanisms controlling HGF activation in the uterus remain unexplored. The long-term goal of the current proposal is to determine the factors involved in the activation of HGF in the uterus, and elucidate the underlying regulatory mechanisms. HGF activator (HGFA) is a serine protease and is most potent in processing HGF activation in vivo, but no information is available regarding its expression in the uterus. The specific aim of this application is to examine the expression and steroid hormonal regulation of the HGF/c-Met regulatory system including HGFA and its inhibitors HAI-1 and HAI-2 in the uterus, and characterize the role of HGFA in the activation of HGF/c-Met signaling. To accomplish this goal, this study will use both in vivo and in vitro models to examine the expression of HGFA, HAI-1, HAI-2, HGF and c-Met in the mouse uterus as well as their regulation by estrogen and progesterone. The role of HGFA in HGF activation will be examined by correlating HGFA expression with HGF activation, c-Met phosphorylation and endometrial epithelial and stromal cell proliferation. Recombinant HGFA, neutralizing antibodies for HGFA and HGF will be utilized during in vitro study to confirm the specific effect of HGFA through HGF activation. Methods used in this proposal include endometrial stromal and epithelial cell culture, Western analysis, Immunohistochemistry, Real-time RT-PCR, In situ hybridization, HGF activation assay and Cell proliferation assay. Data obtained from these experiments will provide novel information on the cellular localization, regulation and function of HGFA in uterine physiology. Further more, this research will provide groundwork for thorough investigation on the molecular mechanisms controlling HGF activation in the uterus, and may even lead to the development of new strategies in treating uterine diseases characterized by abnormal expression and function of HGF, such as endometriosis. The findings from the proposed studies will provide novel information on the regulation of the uterine HGF system and may lead to the development of new therapies to target endometrial pathophysiologies such as endometriosis.

描述（由申请人提供）：越来越多的证据表明肝细胞生长因子（HGF）在子宫发育和子宫内膜异位症和癌等病理条件中起重要作用。在细胞外环境中，单链HGF蛋白水解转化为活性异源二聚体HGF是HGF介导的信号转导中的关键限制步骤，然而，控制子宫中HGF活化的机制仍未探索。目前的建议的长期目标是确定参与子宫内HGF激活的因素，并阐明潜在的调节机制。肝细胞生长因子激活剂（HGFA）是一种丝氨酸蛋白酶，在体内处理肝细胞生长因子激活方面最有效，但没有关于其在子宫中表达的信息。本申请的具体目的是检查子宫中HGF/c-Met调节系统（包括HGFA及其抑制剂HAI-1和HAI-2）的表达和类固醇激素调节，并表征HGFA在HGF/c-Met信号传导激活中的作用。为了实现这一目标，本研究将使用体内和体外模型来检测小鼠子宫中HGFA、HAI-1、HAI-2、HGF和c-Met的表达以及雌激素和孕激素对它们的调节。将通过将HGFA表达与HGF活化、c-Met磷酸化和子宫内膜上皮细胞和基质细胞增殖相关联来检查HGFA在HGF活化中的作用。在体外研究期间将使用重组HGFA、HGFA和HGF的中和抗体，以确认HGFA通过HGF活化的特异性作用。本研究采用的方法包括子宫内膜间质细胞和上皮细胞培养、Western分析、免疫组化、实时荧光定量PCR、原位杂交、肝细胞生长因子激活试验和细胞增殖试验。从这些实验中获得的数据将提供新的信息HGFA在子宫生理学中的细胞定位，调节和功能。本研究为深入研究子宫内HGF激活的分子机制奠定了基础，并可能为治疗子宫内膜异位症等HGF表达和功能异常的子宫疾病提供新的策略。这些研究结果将为调节子宫HGF系统提供新的信息，并可能导致针对子宫内膜病理生理学（如子宫内膜异位症）的新疗法的开发。