LPS INDUCED ENDOTHELIAL CELL ACTIVATION AND APOPTOSIS

感染性休克是革兰氏阴性菌潜在的致命后果 和阳性细菌感染，是受害者的一个重要并发症 创伤性损伤。有多种细菌产品牵涉到 致病分子包括细菌脂蛋白、脂多糖、 脂多糖、脂磷壁酸、肽聚糖、细胞壁产物等。 实验动物被证明能激活固有的细胞“自杀”程序 导致多种细胞类型的凋亡。对分子基础的洞察 对脓毒症的细胞激活/凋亡的反应处于紧张状态 进行调查，希望找到新的治疗方法。信令方式 细菌产物是通过最近描述的Toll样受体产生的 (TLR)在细胞表面。通向NFkappaB的细胞内通路 TLR-2和TLR-4(两者)的激活途径相似 对细菌产品有反应的受体)。然而，细胞凋亡途径 受到的关注较少。调查人员将检查脓毒症引起的 细胞凋亡和一种新的体外激活途径及基因的作用 导致单核细胞、淋巴细胞和 体内的内皮细胞。他们最近发现，细胞凋亡的途径 脂多糖刺激后通过FADD依赖的信号转导和 对NFkappaB的阻断不会使内皮细胞对死亡敏感。这些 观察引发了关于死亡途径和内在机制的问题 细胞保护途径。由于在内毒素抗性中发生了大量的细胞凋亡 (TLR-4缺陷)小鼠在脓毒症期间，我们还推测TLR-2提供了 TLR-4缺陷小鼠的激活信号和死亡信号。此外，他们还 观察到了导致内皮细胞激活的两条途径。第一，老鼠 缺乏功能性Fas(LPR)或FasL(GLD)会降低对内毒素的反应 他们假设Fas-FasL系统是促炎性的。第二，一个具体的 Caspase-8抑制剂降低内毒素诱导的VCAM-1的表达。他们假定 这种蛋白水解酶也是内皮细胞激活的信号。具体目标 本项目的主要内容是：1)确定MyD88、IL-1受体的作用 相关激酶和肿瘤坏死因子受体相关因子-6在脓毒症中的作用 单核/巨噬细胞和血管内皮细胞的凋亡；2)测定 Fas-FasL等分子在细胞凋亡途径中的作用 内毒素诱导的内皮细胞炎症反应；以及3)检测 单核细胞、淋巴细胞或内皮细胞中凋亡基因改变的影响 在诱导小鼠脓毒症后的存活中。

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.