MOLECULAR CELL BIOLOGICAL STUDIES ON EXPRESSION MECHANISM OF TISSUE FACTOR IN ENDOTHELIAL CELLS.

组织因子（Tissue factor, thromboplastin, factor III）是一种表达于细胞表面的受体样膜蛋白，通过与血浆因子VII/VIIa结合引发凝血级联反应。已知组织因子仅在内皮细胞、中性粒细胞和/或单核细胞受到LPS、细胞因子、应激、PAF和其他炎症介质刺激时表达。这些细胞组织因子的高表达直接导致异常的高凝血病，包括脑和/或心脏血栓形成。在本项目中，我们从分子细胞生物学的角度研究组织因子在内皮细胞、中性粒细胞和/或单核细胞中的表达机制和功能。我们还研究了组织因子介导的血管内凝血或血栓形成的体内机制。以下是本项目的研究成果。建立夹心ELISA、流式细胞术、免疫组化染色、RT-PCR等研究方法，探讨组织因子在分子细胞生物学基础上的表达机制。研究了组织因子在lps刺激的人内皮细胞中的表达及其清除机制（囊泡脱落和内吞作用）。从基因转录水平推测了剪切应力对细胞因子介导的内皮细胞组织因子表达的调控机制。发现了一种新的表达组织因子的中性粒细胞功能。这种中性粒细胞组织因子的表达是由内皮细胞和白细胞之间的相互作用介导的，体内研究表明，这可能是血管内凝血或血栓形成的一个原因。另一个新发现是组织因子在单核细胞中的信号转导功能。该功能通过与细胞表面的VII/VIIa因子结合触发，并通过组织因子相关γ链介导。

Tissue factor (thromboplastin, factor III) is a receptor-like membrane protein expressed on cell surface to initiate blood coaqulation cascade reaction through binding with plasma factor VII/VIIa. Tissue factor is known tobe expressed in endothelial cells, neutrophils and/or monocytes only upon their stimulation with LPS,cytokines, stress, PAF,and other inflammatory mediators. Hyperexpression of tissue factor of these cells directly causes abnormal hypercoagulopathy including thrombosis in brain and/or heart.In this project We investigate expression mechanism and fucntion of tissue factor in endothelial cells, neutrophils and/or monocytes on molecular cell biological bases. We also studied on in vivo mechanism of tissue factor-mediated intravascular coagulation or thrombosis. Followings are research results in this project.1.Research techniques of sandwich ELISA,flow-cytometry, immunohistochmical stain, RT-PCR were established to investigate expression mechanism of tissue factor on molecular cell biological bases.2.The expression and clearance mechanisms (vesicle shedding and endocytosis) of tissue factor in LPS-stimulated human endothelial cells were studied.3.A regulation mechanism by shear stress to cytokine-mediated tissue factor expression by human endothelial cells was deduced on gene transcriptional level.4.A new neutrophil function to express tissue factor was found. This neutrophil tissue factor expression was mediated by an interaction between endothelial cells and leukocytes and suggested to be a possible causes of intravascular coagulation or thrombosis by in vivo studies.5.Another new finding was signal transducing function of tissue factor in monocytes. This function was triggered by binding with factor VII/VIIa on cell surface and mediated by tissue factor associated gamma-chain.