LPS INDUCED ENDOTHELIAL CELL ACTIVATION AND APOPTOSIS

LPS 诱导内皮细胞活化和凋亡

基本信息

批准号：
6519351
负责人：
ROBERT K WINN
金额：
$ 23.41万
依托单位：
UNIVERSITY OF WASHINGTON
依托单位国家：
美国
项目类别：
财政年份：
1990
资助国家：
美国
起止时间：
1990-04-01 至 2004-05-31
项目状态：
已结题

项目摘要

Septic shock is a potentially lethal consequence of gram negative and positive bacterial infection and is a significant complication in victims of traumatic injury. There are multiple bacterial products implicated as pathogenic molecules including bacterial lipoproteins, lipopolysaccharide, (LPS), lipoteichoic acid, peptidoglycans, cell wall products, etc. Sepsis in experimental animals was shown to activate the intrinsic cell "suicide" program leading to apoptosis in multiple cell types. Insights into the molecular basis of cellular activation/apoptosis in response to sepsis are under intense investigation in the hope of finding new approaches to therapy. Signaling by bacterial products occurs through the recently described Toll-like receptors (TLR) on the surface of cells. Intracellular pathways leading to NFkappaB activation proceed along similar pathways for TLR-2 and TLR-4 (the two receptors shown to respond to bacterial products). However, apoptosis pathways have received less attention. The investigators will examine sepsis-induced apoptosis and a novel activation pathway in vitro as well as the effect of gene alterations that lead to decreased apoptosis in monocytes, lymphocytes and endothelial cells in vivo. They have recently shown that the apoptotic pathway following stimulation with LPS proceeds through FADD dependent signaling and that blockade of NFkappaB does not sensitize endothelial cells to death. These observation lead to questions regarding the death pathway and intrinsic cyto-protective pathways. Since considerable apoptosis occurs in LPS resistant (TLR-4 deficient) mice during sepsis, we also speculate that TLR-2 provides both activation signals and death signals in TLR-4 deficient mice. Also, they have observed two pathways leading to endothelial cell activation. First, mice lacking functional Fas (lpr) or FasL (gld) have reduced responses to LPS, and they postulate that the Fas-FasL system is pro-imflammatory. Second, a specific caspase-8 inhibitor reduces LPS-induced VCAM-1 expression. They postulate that this protease also signals for endothelial cell activation. The specific aims of this project are: 1) To determine the role of MyD88, Il-1 receptor- associated kinase and TNF receptor- associated factor-6 in sepsis-induced apoptosis of monocytes/macrophages and endothelial cells; 2) To determine the contribution of Fas-FasL and other molecules in that apoptosis pathway in LPS-induced inflammatory response of endothelial cells; and 3) To examine the effects of apoptotic gene alterations in monocytes, lymphocytes or endothelial in survival following induction of sepsis in mice.

感染性休克是革兰氏阴性菌的潜在致命后果和阳性细菌感染，是受害者的严重并发症的外伤。有多种细菌产物涉及致病分子包括细菌脂蛋白、脂多糖、 (LPS)、脂磷壁酸、肽聚糖、细胞壁产物等。脓毒症实验动物被证明可以激活细胞内在的“自杀”程序导致多种细胞类型凋亡。深入了解分子基础脓毒症反应中的细胞活化/凋亡受到强烈影响进行调查，希望找到新的治疗方法。发信号由细菌产物通过最近描述的 Toll 样受体产生 (TLR) 位于细胞表面。导致 NFkappaB 的细胞内途径 TLR-2 和 TLR-4 的激活沿着相似的途径进行（这两个途径显示对细菌产物有反应的受体）。然而，细胞凋亡途径受到的关注较少。研究人员将检查败血症引起的细胞凋亡和体外新的激活途径以及基因的作用导致单核细胞、淋巴细胞和细胞凋亡减少的改变体内的内皮细胞。他们最近发现细胞凋亡途径 LPS 刺激后通过 FADD 依赖性信号传导进行， NFkappaB 的阻断不会使内皮细胞对死亡敏感。这些观察引发了有关死亡途径和内在的问题细胞保护途径。由于 LPS 抗性中发生大量细胞凋亡（TLR-4 缺陷）小鼠在败血症期间，我们还推测 TLR-2 提供 TLR-4 缺陷小鼠的激活信号和死亡信号。还有，他们观察到两条导致内皮细胞激活的途径。一、老鼠缺乏功能性 Fas (lpr) 或 FasL (gld) 会降低对 LPS 的反应，并且他们推测 Fas-FasL 系统具有促炎作用。二、具体 caspase-8 抑制剂可降低 LPS 诱导的 VCAM-1 表达。他们假设这种蛋白酶还发出内皮细胞激活信号。具体目标该项目的主要内容是： 1) 确定 MyD88、Il-1 受体的作用- 脓毒症诱导的相关激酶和 TNF 受体相关因子 6 单核细胞/巨噬细胞和内皮细胞的凋亡； 2）确定 Fas-FasL 和其他分子在细胞凋亡途径中的贡献 LPS诱导的内皮细胞炎症反应； 3) 检查单核细胞、淋巴细胞或内皮细胞凋亡基因改变的影响诱导小鼠败血症后的存活率。