MATRIX METALLOPROTEINASE IN VASCULOGENESIS & ANGIOGENESIS

血管发生中的基质金属蛋白酶

• 批准号: 7720835
• 负责人: PAL GOOZ
• 金额: $ 13.5万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7720835
• 关键词: Affect; Antigen-Presenting Cells; Behavior; Biological Models; Blood Vessels; Cadherins; Catalytic Domain; Cell Adhesion Molecules; Cell Surface Proteins; Cell-Cell Adhesion; Cleaved cell; Clinical; Co-Immunoprecipitations; Complex; Computer Retrieval of Information on Scientific Projects Database; Cytoplasmic Protein; Data; Excision; Funding; Gelatinase A; Grant; Institution; Invaded; Laboratories; Ligands; Matrix Metalloproteinases; Messenger RNA; Metalloproteases; Molecular; Phosphorylation; Play; Proteins; Regulation; Research; Research Personnel; Resources; Role; Selectins; Signal Transduction; Source; TNF-alpha converting enzyme; Techniques; Testing; Tissue Engineering; Transcript; United States National Institutes of Health; Vascular Cell Adhesion Molecule-1; Vascular Endothelial Growth Factors; angiogenesis; desire; human disease; improved; in vivo; intercellular cell adhesion molecule; intermolecular interaction; knock-down; matrigel; novel; research study; response; tool; vasculogenesis

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The broad, long-term objective of our studies is to develop molecular tools that will allow us to enhance or inhibit angiogenesis in vivo. These tools can then be used in the treatment of human diseases and in the tissue engineering of blood vessels. Studies from our laboratory and others highlight ADAM-17 as a key, yet currently underappreciated, player at several levels in the regulation of angiogenesis. In Preliminary Studies, we have shown that knocking down ADAM-17 expression in HUVECs has profound effects on their behavior in model systems for angiogenesis. In particular, HUVECs require ADAM-17 to form networks on Matrigel, to invade Matrigel in response to VEGF, and to express activated ERK and MMP-2 in response to VEGF. These combined observations support our hypothesis that ADAM-17 is a central regulator of angiogenesis due to: 1) its activation by VEGF, 2) its cleavage of a variety of substrates, 3) its formation of complexes with other proteins, and 4) its global effects on ECs, accessory cells, and their interactions. Unlike most metalloproteinases that are activated by removal of a propeptide, ADAM-17 lacking a propeptide is enzymatically inactive. Because most, if not all, of the pro-angiogenic functions that we have attributed to ADAM-17 require the presence of VEGF, in our first specific aim we test the hypothesis that VEGF activates ADAM-17 and that this activation occurs via change in the phosphorylation or enzymatic cleavage of the molecule Further we will study whether the subcellular distribution of ADAM-17 in HUVEC is affected by VEGF signaling. In our second specific aim we will investigate whether ADAM-17 plays a role in angiogenesis by cleaving known substrates such as ErbB ligands, cell-cell adhesion molecules (selectins, VCAM, ICAM, cadherins). We will also attempt to identify novel substrates using the inactive catalytic domain capture technique. The possibility that the pro-angiogenic functions of ADAM-17 are independent of its catalytic activity will also be evaluated in experiments using catalytically-inactive ADAM-17. In specific aim three we will use co-immunoprecipitation to study the possibility that ADAM-17 becomes activated due to intermolecular interactions with cytoplasmic proteins or with other cell surface proteins. In the fourth specific aim we will conduct experiments using microarrays to identify mRNA transcripts whose expression is altered when ADAM-17 expression is knocked down in HUVEC cultures. While these studies may reveal additional proteins that are ADAM-17 ligands or substrates, it is more likely that these studies will reveal proteins with a variety of different, unexpected functions whose expression is regulated by ADAM-17. When such proteins are identified, plans will be made to characterize their roles in angiogenesis. The results of the proposed studies on the activation of ADAM-17, substrates for ADAM-17, ligands for ADAM-17, and proteins whose expression is regulated by ADAM-17 should provide us with a wealth of novel data that will be used to craft an R01 application to NIH. In particular, these data should suggest a variety of novel targets for treatments that will specifically enhance or inhibit angiogenesis as desired in clinical settings and should suggest improved approaches for producing tissue engineered blood vessels.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此，可以在其他清晰的条目中表示。列出的机构是 对于中心，这不一定是调查员的机构。 我们研究的广泛长期目标是开发分子工具，使我们能够增强或抑制体内血管生成。 然后，这些工具可以用于治疗人类疾病和血管组织工程。我们实验室和其他人的研究强调，亚当17是关键，但目前被低估了血管生成调节的多个级别的参与者。在初步研究中，我们已经表明，在HUVEC中击倒Adam-17的表达对它们在血管生成模型系统中的行为具有深远的影响。特别是，HUVEC要求ADAM-17在Matrigel上形成网络，以响应VEGF响应Matrigel，并表达激活的ERK和MMP-2以响应VEGF。这些合并的观察结果支持我们的假设，即Adam-17是血管生成的中心调节剂：1）vegf激活其激活，2）其对各种底物的裂解，3）与其他蛋白质形成复合物，以及4）它全球对EC，辅助细胞及其相互作用的影响。与大多数金属蛋白酶通过去除丙肽激活的大多金属蛋白酶不同，缺乏丙肽的Adam-17在酶上是非活性的。因为我们归因于ADAM-17的促血管生成功能的大多数（如果不是全部）都需要VEGF的存在，在我们的第一个特定目的中，我们测试了VEGF激活ADAM-17的假设，并且这种激活是通过变化而发生的。分子的磷酸化或酶促切割进一步研究，我们将研究HUVEC中ADAM-17的亚细胞分布是否受VEGF信号的影响。在我们的第二个特定目标中，我们将通过裂解已知底物，例如ERBB配体，细胞 - 细胞粘附分子（Selectins，VCAM，ICAM，ICAM，Cadherins）来研究ADAM-17是否在血管生成中起作用。我们还将尝试使用非活性催化域捕获技术来识别新的底物。 ADAM-17的促血管生成功能与其催化活性无关的可能性也将在使用催化活性ADAM-17的实验中评估。在特定的目标三中，我们将使用共免疫沉淀研究ADAM-17由于与细胞质蛋白或其他细胞表面蛋白的分子相互作用而被激活的可能性。在第四个特定目的中，我们将使用微阵列进行实验，以识别在HUVEC培养物中敲低ADAM-17表达时表达会改变的mRNA转录本。尽管这些研究可能揭示了其他是ADAM-17配体或底物的蛋白质，但这些研究更有可能揭示具有各种不同的意外功能的蛋白质，其表达受ADAM-17的调节。确定此类蛋白质时，将制定计划以表征其在血管生成中的作用。提出的关于ADAM-17激活，ADAM-17的底物，ADAM-17的配体的底物的结果和蛋白质的表达受ADAM-17调节的蛋白质应为我们提供大量新的数据，这些数据将用于使用这些数据，这些数据将用于将R01应用程序申请到NIH。特别是，这些数据应提出各种新型治疗靶标，这些靶标将在临床环境中特异性增强或抑制血管生成，并应提出改进的生产组织工程血管的方法。