Transendothelial Migration of Leukocytes: Developing New Paradigms in Health and Disease

项目摘要/摘要 经内皮细胞移位(TEM)，或渗出，是白细胞紧密挤压之间的步骤 排列在炎症部位的毛细血管后小静脉的对置内皮细胞。大多数好的，坏的， 而炎症的丑恶发生在白细胞穿过血管之后。透彻了解 因此，调节透射电子显微镜的分子和机制应该会增强我们控制这一过程的能力 从治疗上讲。因此，我的实验室已经研究这个过程30年了。我们已经制作了一些 该领域的开创性发现，包括鉴定和发现：1.血小板/内皮细胞黏附 分子-1(PECAM)和CD99作为TEM的主要选择性调节因子2.它们的下游信号转导 通往透射电子显微镜的路径；3.侧边界回收隔间(LBRC)，一个相互连接的网状结构 沿内皮细胞边界局部循环的小管泡状膜；4.表面分子 参与电子显微镜(PECAM、CD99等)在这个过程中，当白细胞通过 内皮细胞边界；5.透射电子显微镜作用促进部分单核细胞分化为树突状细胞； 白细胞的细胞旁和跨细胞的透射电子显微镜使用相同的机制和机制。 无论我们是从白细胞还是内皮细胞的角度来研究透射电子显微镜，表面 黏附/信号分子、细胞内信号通路、膜动力学或内皮细胞 超微结构，我们一致和独立地汇聚在一个最终的共同机制上，无论 无论我们是研究中性粒细胞、单核细胞还是T细胞；无论炎症条件或 研究的模型：TEM要求LBRC沿着微管定向移动到 白细胞正在迁移。我们研究的所有分子都激活了这一机制。任何有可能 抑制这种“靶向循环的LBRC”抑制了80-90%的透射电子显微镜在体外和体内。 现有抗炎疗法的最大缺点是它们也阻碍了有益的 发炎。我们已经开发了生化和遗传工具来选择性地阻止有针对性的回收 LBRC，因此在多种炎症性疾病体内模型中的透射电子显微镜。由于这些试剂和诱导剂 EC选择性基因敲除小鼠只影响EC，即先天性和获得性炎症反应的所有其他方面 保持完好无损。由于我们只能阻断80%-90%的透射电子显微镜，10%-20%的白细胞逃脱封锁 进入能够产生正常炎症反应的组织。我们的初步数据显示，我们可以减少 在不干扰寄主需要的能力的情况下发生的不适应炎症的强度 炎症反应，并保持健康。我们将在一些急性和非传染性疾病模型中检验这一假说。 慢性炎症。我们还将使用我们的能力，在选定的时间点选择性地阻止瞬变电磁，以研究其作用 透射电子显微镜在疾病的发生、发展和解决中的作用。调节的分子和机制 肺血管床中的TM尚不清楚。我们将识别它们，并与体循环进行比较。

Project Summary/Abstract Transendothelial migration (TEM), or diapedesis, is the step in which leukocytes squeeze between tightly apposed endothelial cells that line the post-capillary venules at sites of inflammation. Most of the good, the bad, and the ugly of inflammation occurs after leukocytes cross blood vessels. A thorough understanding of the molecules and mechanisms that regulate TEM should therefore enhance our ability to control the process therapeutically. Therefore, my lab has been studying this process for 30 years. We have made some of the seminal discoveries in the field, including the identification and discovery of: 1.Platelet/endothelial cell adhesion molecule-1 (PECAM) and CD99 as major selective regulators of TEM and 2. their downstream signaling pathways leading to TEM; 3.The lateral border recycling compartment (LBRC), an interconnected reticulum of tubule-vesicular membrane that recycles locally along the endothelial cell borders; 4.That the surface molecules involved in TEM (PECAM, CD99, etc.) work sequentially in the process as the leukocyte passes through the endothelial cell border; 5.That the act of TEM promotes differentiation of some monocytes into dendritic cells; 6. That paracellular and transcellular TEM of leukocytes use the same machinery and mechanisms. Whether we approached TEM from the standpoint of the leukocyte or the endothelial cell, the surface adhesion/signaling molecules, the intracellular signaling pathways, membrane dynamics, or endothelial cell ultrastructure, we consistently and independently converged on a final common mechanism regardless of whether we were studying neutrophils, monocytes, or T cells; regardless of the inflammatory conditions or models studied: TEM required the targeted movement of the LBRC along microtubules to the site at which the leukocyte was migrating. All of the molecules that we studied worked to activate this mechanism. Anything that inhibited this “targeted recycling of the LBRC” inhibited TEM by 80-90% in vitro and in vivo. The biggest shortcoming of existing anti-inflammatory therapies is that they also block beneficial inflammation. We have developed biochemical and genetic tools to selectively block targeted recycling of the LBRC and hence TEM in multiple in vivo models of inflammatory disease. Since these reagents and inducible EC-selective knockout mice only affect EC, all other aspects of the innate and adaptive inflammatory responses remain intact. Since we are only able to block TEM by 80–90%, the 10–20% of leukocytes that escape blockade enter the tissues able to mount a normal inflammatory response. Our preliminary data show that we can diminish the intensity of maladaptive inflammation without interfering with the ability of the host to mount desirable inflammatory responses and remain healthy. We will test this hypothesis in a number of models of acute and chronic inflammation. We will also use our ability to selective block TEM at selected time points to study the role of TEM in the initiation, progression, and resolution of disease. The molecules and mechanisms that regulate TEM in the pulmonary vascular bed are unknown. We will identify them and compare to the systemic circulation.