TRAUMA & ORGAN FAILURE--ROLE OF LPS BINDING PROTEIN

创伤

• 批准号: 6164814
• 负责人: STEWART C WANG
• 金额: $ 10.68万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-03-01 至 2002-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6164814
• 关键词: bacterial toxins; binding proteins; endotoxins; gram negative bacteria; immune tolerance /unresponsiveness; inflammation; kidney cell; laboratory rat; lipopolysaccharides; liver cells; multiple organ failure; protein biosynthesis; protein structure function; respiratory epithelium; trauma

Sepsis and septic shock are still major causes of morbidity and mortality among critically ill surgical patients. During gram-negative sepsis, endotoxins (LPS) binds to CD14 receptors on mononuclear cells, causing release of inflammatory mediators which can cause tissue injury and organ failure. LPS-binding protein (LBP) enhances binding of LPS to the CD14 complex. When bound to LBP, many fold smaller concentrations of LPS can activate host cells. We have recently been the first to report that LBP production is induced in extrahepatic tissues in rats by injury. We have now confirmed extrahepatic LBP production in humans also. We hypothesize that locally produced LBP modulates the local immune response to LPS and Gram-negative infection, serving to localize the infecting agent and focusing host defenses at the local site. If, however, the local immune activation is too great and spills into the circulation or if the activation persists for too long, extrahepatic induction of LBP may predispose toward MSOF. We propose to study the factors controlling the production of LBP by cells other than hepatocytes and to compare them to production by hepatocytes themselves. In preparation, we have cloned rat LBP cDNA, purified rat LBP protein, generated anti-LBP antibody, defined in vitro as well as in vivo models of LBP regulation and are about to isolate and sequence the rat LBP promoter and express recombinant rat LBP. AIM I will define the factors regulating lipopolysaccharide-binding protein production by pulmonary, renal and extrahepatic cells in vitro. Studies will be done in non-hepatocytes as well as hepatocytes and contrasted to better define the mechanism behind the tissue-specific regulation of LBP. We begin by defining the cytokines which regulate LBP production. Nuclear run-on assays will be done to confirm and quantify transcriptional induciton. We will characterize the promoter DNA elements and nuclear factors controlling transcription of the LBP gene. Finally, we will determine whether LBP production is controlled by alteractions in mRNA stability or translational efficiency as elements in the 3'-UTR of its mRNA suggest. AIM II will define the regulation of lipopolysaccharide-binding protein production by extrahepatic tissues in vivo. Using two models of injury (hindlimb turpentine injection and hemorrhagic shock), we will determine whether the same cytokines that regulate LBP production in vitro also regulate LBP production in vivo. Further characterization of the cells that produce the LBP will also be done. AIM III will determine the functional role of locally produced (pulmonary) lipopolysaccharide-binding protein on host responses to LPS. Using hemorrhage, injury, or an adenovirus vector containing the LBP cDNA, we will induce increased local pulmonary production of lBP and determine to what extent increased local levels of LBP potentiates intrapulmonary immune cell activation and tissue injury by systemic or intratracheal LPS. At the completion of our studies, we will have defined how LBP is induced in extrahepatic tissues after injury as well as the pathophysiologic significance of such local LBP production. This information will provide important and previously unavailable insights into post-traumatic responses to Gram-negative sepsis and endotoxemia.

败血症和败血性休克仍然是发病率和死亡率的主要原因 在重症手术患者中。 在革兰氏阴性败血症中， 内毒素（LPS）与单核细胞上的CD14受体结合，导致 释放炎症介质会导致组织损伤和器官 失败。 LPS结合蛋白（LBP）增强了LPS与CD14的结合 复杂的。 当绑定到LBP时，许多折叠较小的LP可以 激活宿主细胞。 我们最近是第一个报告LBP的人 受伤在大鼠的肝外组织中诱导产生。 我们有 现在确认了人类的肝外LBP产生。 我们假设 局部产生的LBP调节了对LP的局部免疫反应， 革兰氏阴性感染，用于本地化感染剂和 将主机防御重点放在当地地点。 但是，如果是局部免疫 激活太大了，溢出到循环中或 激活持续时间太长，肝外诱导LBP可能 倾向于MSOF。 我们建议研究控制的因素 除肝细胞以外的其他细胞生产LBP，并将其比较 肝细胞的生产本身。 在准备中，我们克隆了老鼠 LBP cDNA，纯化的大鼠LBP蛋白，生成抗LBP抗体，定义为 体外和体内LBP调节模型，即将分离 并对大鼠LBP启动子和表达重组大鼠LBP进行测序。 目标我 将定义调节脂多糖结合蛋白的因素 在体外由肺，肾脏和肝外细胞产生。 研究 将在非羊皮细胞以及肝细胞中进行，并形成对比 更好地定义了LBP组织特异性调节背后的机制。 我们首先定义调节LBP产生的细胞因子。 核 将进行跑步测定以确认和量化转录 诱因。 我们将表征启动子DNA元素和核 控制LBP基因转录的因素。 最后，我们会的 确定LBP的产生是否受mRNA的改变控制 稳定性或翻译效率作为其mRNA的3'-UTR的元素 建议。 AIM II将定义脂多糖结合的调节 肝外组织在体内产生蛋白质。 使用两种型号 受伤（后肢松节油注射和出血性休克），我们将 确定在体外调节LBP产生的相同细胞因子是否 还调节体内LBP的生产。 进一步的表征 产生LBP的细胞也将完成。 AIM III将确定 局部生产（肺）脂多糖结合的功能作用 宿主对LPS的蛋白质反应。 使用出血，受伤或 含有LBP cDNA的腺病毒载体，我们将诱发局部增加 LBP的肺产量，并确定局部增加的程度 LBP的水平增强了肺内免疫细胞活化和组织 系统性或气管内LPS受伤。 完成研究时， 我们将定义如何在肝外组织中诱导LBP之后 损伤以及这种局部LBP的病理生理意义 生产。 这些信息将提供重要和以前 对革兰氏阴性败血症的创伤后反应的无法获得的见解 和内毒素血症。

项目成果

Thermal injury induces expression of CD14 in human skin.

• DOI: 10.1016/s0305-4179(02)00034-7
• 发表时间: 2002-05
• 期刊: Burns : journal of the International Society for Burn Injuries
• 作者: L. Steinstraesser;W. Alarcon;M. Fan;R. Klein;A. Aminlari;C. Zuccaro;G. Su;Stewart C. Wang

Burn wounds infected with Pseudomonas aeruginosa triggers weight loss in rats.

• DOI: 10.1186/1471-2482-5-19
• 发表时间: 2005-09-17
• 期刊: BMC surgery
• 影响因子: 1.9
• 作者: Steinstraesser, Lars;Burkhard, Olaf;Wang, Stewart C
• 通讯作者: Wang, Stewart C

An essential role for complement C5a in the pathogenesis of septic cardiac dysfunction.

• DOI: 10.1084/jem.20051207
• 发表时间: 2006-01-23
• 期刊: The Journal of experimental medicine
• 作者: Niederbichler AD;Hoesel LM;Westfall MV;Gao H;Ipaktchi KR;Sun L;Zetoune FS;Su GL;Arbabi S;Sarma JV;Wang SC;Hemmila MR;Ward PA
• 通讯作者: Ward PA

CD14 and lipopolysaccharide binding protein expression in a rat model of alcoholic liver disease.

酒精性肝病大鼠模型中 CD14 和脂多糖结合蛋白的表达。

• 发表时间: 1998
• 期刊: The American journal of pathology
• 作者: Su,GL;Rahemtulla,A;Thomas,P;Klein,RD;Wang,SC;Nanji,AA
• 通讯作者: Nanji,AA