Hepatic Stabilin-2 Mechanisms of Clearance

摘要 人的肝脏清除细胞外基质物质，包括透明质酸、硫酸软骨素和 死亡的红血球，从循环中，除了作为新陈代谢的能量屋。这种拾荒者的行为 活性是由肝窦内皮细胞(LSECs)表达Stablin-2受体实现的。vt.在.的基础上 通过一组配体的激活，Stablin-2诱导了MAP激酶和NF-kB通路，所有LSECs都通过这些通路 “感觉”他们的环境，并使用像一氧化氮这样的小分子来回报。LSEC充当接口 在血液和肝细胞之间，含有调节大分子的筛板 向肝细胞灌流。LSECs的损伤通常会减少窗孔的数量和大小， 影响整体肝脏新陈代谢。脂肪肝等肝病影响了三分之一以上的成年人 这些疾病直接影响LSECs的清道夫/感应能力。受损的LSEC 导致血液中细胞外基质物质的积聚，从而对肾脏功能产生负面影响。这个 肝脏清除血液中的死亡细胞，但它也是转移性癌症的场所，两者密切相关 为LSEC的健康干杯。为了更充分地了解LSEC的清关作用，项目负责人建议 在模拟肝窦条件的剪应力条件下检查它们。这项研究将是 首次在不同流量下检测LSECs的清除活性以评估LSECs的活性 STABBILIN-2为主要受体。需要解决的问题包括：剪切力对 是简单的配体还是复杂的配体，比如细胞周围的透明质酸涂层细胞？LSEC如何对其配体作出反应 在剪切应力过程中，脂肪肝这一常见病对LSEC Stablin-2介导的LSEC有什么影响 内吞作用？根据他之前对Stablin-2的研究，项目负责人假设特定的配体 在切应力下，循环中的富含透明质酸的细胞与Stablin-2的结合增强，使它们能够 肝脏停止循环和增殖，且Stabilin-2表达水平与肝脏直接相关 清除功能。拟议研究的中心假设是配体-Stablin-2相互作用， 随后是细胞信号的内吞和激活，受血窦和 肝脏代谢状态(如脂肪肝)，影响血液清除和癌细胞向肝脏转移。 项目负责人将使用稳定的重组细胞和来自正常和 大鼠肝脏疾病模型。从这些肝脏中纯化的LSECs将在Flow细胞培养下进行评估 监测配基结合和信号事件的条件，由Stablin-2传播。为了完成这个项目 为了实现这一目标，项目负责人将追求三个具体目标：1)确定流动状态下配体的Stabilin-2结合 条件，2)确定切应力对LSECs细胞信号转导的影响，3)确定Stablin-2 正常肝和脂肪肝的表达和内吞活性。

ABSTRACT The human liver scavenges extracellular matrix material, including hyaluronic acid, chondroitin sulfate, and dead red blood cells, from circulation, in addition to serving as a metabolic power house. This scavenging activity is performed by liver sinusoidal endothelial cells (LSECs) expressing the Stabilin-2 receptor. Upon activation by a subset of ligands, Stabilin-2 induces the MAP kinase and NF-kB pathways, by which all LSECs "sense" their environment and reciprocate using small molecules like nitric oxide. LSECs serve as the interface between the blood and hepatocytes and contain fenestrae (sieve plates) that regulate macromolecules perfusing through to the hepatocytes. Damage of LSECs often reduces the number and size of the fenestrae, affecting overall liver metabolism. Liver diseases, such as fatty liver, affect more than a third of the adult population, and these diseases directly affect the scavenger/sensing capacity of LSECs. Damaged LSECs lead to a build-up of extracellular matrix material in the blood that then negatively affects kidney function. The liver clears dead cells from blood, but it is also a site for metastatic cancer, both of which are intimately related to the health of LSECs. To more fully understand the clearance role of LSECs, the project leader proposes to examine them under shear stress conditions that mimic conditions in the hepatic sinusoid. This study will be the first time that the clearance activity of LSECs has been examined under variant flow to assess activity of Stabilin-2 as the major receptor. Questions to be addressed include: What effect does shear stress have on simple ligands or complex ligands like pericellular HA coated cells? How do LSECs respond to their ligands during shear stress, and what effect does fatty liver, a common ailment, have on LSEC Stabilin-2-mediated endocytosis? From his previous studies with Stabilin-2, the project leader hypothesizes that specific ligand binding to Stabilin-2 by circulating hyaluronan-rich cells is enhanced under shear stress, allowing them to cease circulation and proliferate in liver, and that expression levels of Stabilin-2 directly correlates with liver clearance function. The central hypothesis of the proposed research is that ligand-Stabilin-2 interactions, followed by endocytosis and activation of cell signaling, are influenced by shear stress at the sinusoids and hepatic metabolic status (e.g., fatty liver), which affects blood clearance and cancer cell metastasis to liver. The project leader will test this hypothesis using stable recombinant cells and primary LSECs from normal and disease models of rat livers. Purified LSECs from these livers will be evaluated under flow cell culture conditions to monitor ligand binding and signaling events propagated by Stabilin-2. To accomplish the project goal, the project leader will pursue three specific aims: 1) Determine Stabilin-2 binding of ligands under flow conditions, 2) Determine the effects of shear stress on cellular signaling in LSECs, and 3) Determine Stabilin-2 expression and endocytic activity in normal and fatty liver.