TTP PATHOGENESIS IN MODEL SYSTEMS IN VITRO AND IN VIVO

本研究建议是对NHLBI RFA HL-95-007(血栓症)的响应 血小板减少性紫癜(TTP)和艾滋病毒)。长期目标是 利用体内外研究进一步阐明TTP的发病机制 以实验模型研究可能涉及的因素 血栓性微血管病(TMA)的发病机制 易感因素、早期诊断和对治疗的洞察。 有三个具体目的：1)体外测定炎性细胞和 最有可能参与的细胞的凝血反应 TMA(内皮细胞、血小板和白细胞)的发病机制已知 如志贺样毒素(SLT)和唾液酸酶等煽动剂。它的用途 微血管内皮细胞与大血管内皮细胞的比较 在静态和剪切应力条件下，将显著增加这些 实验。正常人、HIV和TTP患者的细胞和血浆 将用于阐明体外系统中的机制；2) 用动物(小鼠和小鼠)测定SLT和唾液酸酶的体内反应 已知对SLT易感的兔)。特异性基因(P-选择素、E-选择素 选择素、P-选择素和E-选择素以及vWF)基因敲除小鼠将用于 确定我们观察结果的特殊性；3)前瞻性地 评估HIV中TMA综合征的潜在易感宿主因素 被感染的病人。 这项研究将利用细胞和分子生物学技术 研究毒素和唾液酸酶对各种感兴趣细胞的影响。这个 这项研究的设计将允许我们以一种 使用体外技术对一系列因素进行检查，然后 当我们找到相关的交互单元时，按程度聚焦实验 和分子。从体外研究中获得的知识将 在一系列有针对性的体内实验中达到高潮，寻找特定的 用生理学方法研究整个动物的分子相互作用 SLT交付。 在项目完成后，我们将更多地了解 在发展中起关键作用的因素和细胞相互作用 TMA在HIV感染患者和非感染患者中都存在。这将提供 对诊断、治疗和预防发展的见解 养生法。

This research proposal is a response to NHLBI RFA HL-95-007 (Thrombotic Thrombocytopenic Purpura (TTP) and HIV). The long term objectives are to further elucidate the pathogenesis of TTP using in vitro and in vivo experimental models to study factors that may be involved in the pathogenesis of thrombotic microangiopathy (TMA), to determine predisposing factors, early diagnosis and insights into treatment. There are three specific aims: 1) To determine in vitro inflammatory and coagulation responses of the cells most likely to be involved in the pathogenesis of TMA (endothelial cells, platelets and leukocytes) to known inciting agents such as shiga-like toxins (SLTs) and sialidase. The use of micro and macro vascular ECs and comparison of endothelial cells grown under static and shear stress conditions will add significantly to these experiments. Cells and plasma from normal subjects, HIV and TTP patients will be used to elucidate mechanisms in the in vitro system; 2) To determine in vivo responses to SLTs and sialidase using animals (mice and rabbits) known to be susceptible to SLTs. Specific gene (P-selectin, E- selectin, P- and E-selectin, and vWF) knockout mice will be used to determine the specificity of our observations; 3) To prospectively evaluate host factors potentially predisposing to TMA syndromes in HIV infected patients. The research will utilize cell and molecular biological techniques to study toxin and sialidase effects on the various cells of interest. The design of the study will allow us to initiate the experiments with an examination of a broad range of factors using in vitro techniques and then focus the experiments by degree as we find the relevant interactive cells and molecules. The knowledge gained from the in vitro studies will culminate in a series of targeted in vivo experiments looking at specific molecular interactions in whole animals utilizing physiological methods of SLT delivery. At the completion of the project we will know significantly more about the factors and cellular interactions which are critical in the development of TMA in both HIV infected and non-infected patients. This will provide insights for the development of diagnostic, therapeutic and preventative regimens.