Pericyte proteinase inhibitors and EC tube stabilization

描述（由申请人提供）：在这个更新提案中，我们继续我们的工作，证明周细胞和内皮细胞（EC）衍生的蛋白酶抑制剂在3D细胞外基质环境中控制内皮管稳定过程中的关键功能作用。我们的总体假设是，蛋白酶抑制剂TIMP-3和TIMP-2具有稳定特性，因为它们能够抑制促形态发生和促回归刺激，这是试管稳定所必需的。在最后的资助期间，我们在证明周细胞稳定EC内衬管的能力和timp在这一过程中的作用方面取得了重要进展，开发了新的EC-周细胞管共培养模型，以确定这些事件的分子需求，并确定了TIMP-3和TIMP-2的关键EC靶点，如MT1-MMP， MMP-1， MMP-10和ADAM- 15介导其影响。此外，我们在以下方面的能力也取得了重大进展：分析内皮细胞和周细胞的基因表达和功能，以确定这一过程的分子要求；分析细胞外基质重塑变化的影响，如确定EC-周细胞相互作用催化血管基底膜基质组装的需要；通过PDGF-BB和HB-EGF介导的信号传导，确定周细胞被招募到ec内衬管的机制；并确定关键的生长因子如tgf -如何影响这一过程。我们提出三个具体目标：具体目标1：确定周细胞能够催化内皮基底膜基质组装以促进血管稳定的分子机制。特定目标#2：定义控制周细胞如何特异性募集到内皮细胞壁管的机制，以及它们如何通过迁移事件动态扫描内皮基表面以调节管的稳定性。特定目标#3：定义周细胞募集到发育中的血管如何通过普遍存在的细胞因子tgf - β介导的消退刺激的衰减导致稳定。

DESCRIPTION (provided by applicant): In this renewal proposal, we continue our work demonstrating the critical functional role of pericyte and endothelial cell (EC)-derived proteinase inhibitors to control the process of endothelial tube stabilization in 3D extracellular matrix environments. Our overall hypothesis is that the proteinase inhibitors, TIMP-3 and TIMP-2, possess stabilizing properties due to their ability to suppress both promorphogenic and proregression stimuli which are necessary for tube stabilization. During this last funding period, we have made essential progress in demonstrating the ability of pericytes to stabilize EC-lined tubes and the role of TIMPs during this process, have developed novel EC-pericyte tube coculture models to identify molecular requirements for these events, and have identified critical EC targets of TIMP-3 and TIMP-2 such as MT1-MMP, MMP-1, MMP-10 and ADAM- 15 which mediate their influence. In addition, we have made significant advances in our ability to; analyze gene expression and function in ECs versus pericytes to identify the molecular requirements for this process; analyze the influence of extracellular matrix remodeling changes such as identifying a requirement for EC- pericyte interactions to catalyze vascular basement membrane matrix assembly; identify mechanisms whereby pericytes are recruited to EC-lined tubes through PDGF-BB and HB-EGF mediated signaling; and determine how critical growth factors such as TGF-beta affect this process. We propose three specific aims which are; Specific Aim #1: To determine the molecular mechanisms by which pericytes are able to catalyze endothelial basement membrane matrix assembly to facilitate vascular stabilization. Specific Aim #2: To define the mechanisms controlling how pericytes are specifically recruited to endothelial cell-lined tubes and how they dynamically scan the endothelial basal surface through migratory events to regulate tube stabilization. Specific Aim #3: To define how pericyte recruitment to developing vascular tubes leads to stabilization through attenuation of regression stimuli mediated by the ubiquitous cytokine, TGF-beta. PUBLIC HEALTH RELEVANCE: This work focuses on the ability of pericytes, a cell type surrounding small blood vessels, to support the integrity of these cell-lined tube structures. These support cells produce a series of proteins that facilitate this support mechanism. A basic understanding of the mechanisms underlying how blood vessels form and become stabilized is critical in efforts to stimulate or inhibit the process in the context of various human diseases such as cardiovascular disease, diabetes or cancer.