Determinants of sinusoidal endothelial cell phenotype

描述（由申请人提供）：酒精性肝病中毛细血管化先于纤维化。在毛细血管化过程中，窦状隙内皮细胞（SEC）失去窗孔并形成基底膜，这是一种以前未研究过的表型丧失。我们建议在一系列的体外和体内实验中检查SEC表型的决定因素。在体外，将在同型培养物和与其他肝细胞共培养物中检查分离的细胞，以确定SEC表型是否受旁分泌调节或异型接触调节。体外研究将利用从正常大鼠和肝病模型分离的细胞，特别是脂肪变性、酒精性肝病和使用硫代乙酰胺的纤维化模型。在肝脏疾病模型中进行的SEC研究将检查异型细胞对正常SEC表型决定因素的反应变化。对肝病模型中肝细胞和星状细胞的研究将寻找决定SEC表型的因素的变化。蛋白质组学的方法将被用来确定和检查参与旁分泌调节的蛋白质。体内研究将通过监测肝组织学的变化，在正常大鼠或来自上述肝病模型的大鼠中使用可溶性因子或其抑制剂的连续门静脉内输注来证实作为SEC表型决定因素的可溶性因子的相关性。将表征来自正常和肝脏疾病模型大鼠的SEC之间的基因表达差异，以了解是什么调节SEC表型。这些研究将用于鉴定在模型中改变的SEC标志物，以确认蛋白质组学研究的结果，并鉴定调节在正常SEC中表达的基因表达的共同途径，但不在从肝病模型分离的SEC中表达。

DESCRIPTION (provided by applicant): Capillarization precedes fibrosis in alcoholic liver disease. In capillarization, sinusoidal endothelial cells (SEC) lose fenestrae and form a basement membrane, a loss of phenotype that has not been examined previously. We propose to examine determinants of SEC phenotype in a series of in vitro and in vivo experiments. In vitro, isolated cells will be examined in homotypic culture and in co-culture with other liver cells to determine whether SEC phenotype is regulated by paracrine regulation or by heterotypic contact. The in vitro studies will utilize cells isolated from normal rats and from liver disease models, notably models of steatosis, alcoholic liver disease and fibrosis using thioacetamide. Studies in SEC from liver disease models will examine changes in response to determinants of normal SEC phenotype by heterotypic cells. Studies in hepatocytes and stellate cells from liver disease models will look for changes in factors that determine SEC phenotype. A proteomic approach will be used to identify and examine proteins involved in paracrine regulation. In vivo studies will provide confirmation of the relevance of the soluble factors implicated as determinants of SEC phenotype using continuous intraportal infusions of the soluble factors or their inhibitors in normal rats or rats from the above mentioned liver disease models by monitoring changes in liver histology. Differences in gene expression between SEC from normal and liver disease model rats will be characterized to understand what regulates SEC phenotype. These studies will be used to identify markers of SEC that are altered in the models, to confirm the findings of the proteomic studies and to identify common pathways that regulate expression of genes that are expressed in normal SEC, but not in SEC isolated from liver disease models.