MMP-9 and Transport through Vascular Endothelium

MMP-9 和通过血管内皮的运输

• 批准号: 6684491
• 负责人: SUSAN M LESSNER
• 金额: $ 4.64万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-12-01 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6684491
• 关键词: atherosclerotic plaque; blood lipoprotein; bone marrow; cell cell interaction; cell migration; collagenase; dextrans; flow cytometry; fluorimetry; genetically modified animals; laboratory mouse; macromolecule; mixed tissue /cell culture; monocyte; pathologic process; postdoctoral investigator; vascular cell adhesion molecule; vascular endothelium; vascular endothelium permeability; western blottings

DESCRIPTION: (provided by applicant): The proposed research is designed to test the hypothesis that increased expression of gelatinase B (MMP-9) during monocyte transmigration of vascular endothelium results in an increase in endothelial permeability to macromolecules. Such an effect may contribute to diminished endothelial barrier function and initiation of atherosclerotic lesion through facilitating the non-specific entry and accumulation of plasma molecules, including circulating lipoproteins. Specific aims include measuring permeability of endothelial monolayers to macromolecular tracers during transmigration of both wild type and MMP-9 knockout monocytes, and investigating the mechanism of MMP-9 induction during monocyte/endothelial cell interactions. We propose to investigate permeability changes during monocyte transmigration using an in vitro coculture system consisting of mouse bone marrow monocytes and mouse aortic endothelium grown on matrix-covered cell culture inserts. To investigate the specific role of MMP-9 in transmigration, we will use monocytes and endothelial cells derived both from wild type and from MMP-9 knockout mice. Endothelial permeability will be measured using flourescently labeled dextrans. Tracer concentrations will be measured fluorimetrically. Expression of MMP-9 protein and enzymatic activity will be determined by means of Western blotting and SDS-PAGE zymography, respectively. To explore the mechanism of MMP-9 induction during monocyte transmigration of endothelium, we will examine the role of specific interactions between monocyte integrins and endothelial cell adhesion molecules. Specifically, we propose to assess the role of monocyte binding to endothelial VCAM-1 using VCAM-1 knockout mouse endothelial cells in transmigration assays. These studies should further our understanding of the early events that contribute to the development of athersclerotic lesions and could provide insight into the use of MMP-9 inhibitors to treat endothelial barrier dysfunction.

描述：(申请者提供)：建议的研究旨在测试 明胶酶B(MMP9)表达增加的假说 血管内皮细胞单核细胞移位导致血管内皮细胞数量增加 内皮对大分子的渗透性。这样的影响可能有助于 内皮屏障功能减弱与动脉粥样硬化的发生 通过促进血浆的非特异性进入和积聚来损伤 分子，包括循环脂蛋白。具体目标包括测量 血管内皮细胞单层对大分子示踪剂的渗透性 野生型和基质金属蛋白酶-9基因敲除单核细胞的迁移 单核细胞/内皮细胞诱导基质金属蛋白酶-9的机制探讨 互动。我们建议研究单核细胞通透性的变化。 小鼠骨骼体外共培养体系的移植 骨髓单核细胞和小鼠主动脉内皮细胞在基质覆盖细胞上的生长 文化插页。为了研究基质金属蛋白酶-9在轮回中的具体作用， 我们将使用来自野生型和内皮型的单核细胞和内皮细胞 来自基质金属蛋白酶-9基因敲除小鼠。将使用以下方法测量内皮通透性 荧光标记的右旋糖苷。将测量示踪剂浓度 荧光计量法。基质金属蛋白酶-9蛋白的表达和酶活性 分别用Western blotting和SDS-PAGE酶谱测定。 探讨单核细胞移行过程中基质金属蛋白酶-9的诱导机制 内皮，我们将研究单核细胞之间的特异性相互作用的作用 整合素与内皮细胞黏附分子。具体来说，我们建议 用VCAM-1基因敲除评估单核细胞与内皮细胞VCAM-1结合的作用 迁移实验中的小鼠内皮细胞。这些研究应该进一步 我们对早期事件的理解有助于 动脉粥样硬化病变，并可提供对基质金属蛋白酶-9的使用的洞察 用于治疗内皮屏障功能障碍的抑制剂。