Bacterial Membrane Proteins in Gram-Negative Sepsis

革兰氏阴性脓毒症中的细菌膜蛋白

基本信息

批准号：
7151973
负责人：
H Shaw Warren
金额：
$ 35.25万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-03-01 至 2008-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7151973
关键词：
Affinity Chromatography Agonist Antibiotics Apolipoprotein A-I Bacteria Biological Biology Blood Circulation Cell Adhesion Molecules Cell Wall Cells Complex Confocal Microscopy Cytokine Activation Endothelial Cells Enzyme-Linked Immunosorbent Assay Equilibrium Centrifugation Escherichia coli Infections Functional disorder Goals Grant High Pressure Liquid Chromatography Human Immunoglobulin G In Vitro Infection Inflammation Inflammatory Inflammatory Response Infusion procedures Kinetics Laboratories Leukocytes Ligands Ligation Lipoproteins Location Lung Mediating Membrane Membrane Proteins Modeling Monoclonal Antibodies Mus Natural Immunity Organ Pathogenesis Pattern recognition receptor Peptidoglycan Peritoneal Play Process Production Proteins Puncture procedure Receptor Signaling Relative (related person)Research Personnel Role Sepsis Serum Signal Transduction Splenocyte System TLR2 gene TLR4 gene Techniques Time Tissues Toll-like receptors Up-Regulation Work cytokine density design in vivo macrophage particle peptidoglycan-associated lipoprotein polyclonal antibody programs prototype research study response time use

项目摘要

DESCRIPTION (provided by applicant): During infection, membrane components are shed from bacteria in complexes that include LPS and two bacterial membrane lipoproteins: murein lipoprotein (MLP), and peptidoglycans associated lipoprotein (PAL). This renewal proposal seeks to study the physical interactions that each of these bacterial membrane components have with each other and with host lipoproteins, and the relative contribution of each in activation of cellular and systemic inflammatory responses that are mediated through Toll-like receptors (TLRs). Our central hypothesis is that MLP and PAL contribute to the pathogenesis of sepsis, and that their activity in vivo may be altered by interactions with each other, with LPS, and with host proteins and lipoproteins. Whereas most previous work has studied chemically purified or synthetic TLR agonists, this work will study natural forms of MLP and PAL wherever possible. The first specific aim is to study the kinetics of release of MLP and PAL from the bacterial cell wall into bacterial membrane complexes and into low density forms in serum. In vitro studies will utilize human serum. In vivo studies will be performed in two models of infection in mice: peritoneal E. coli infection and cecal ligation and puncture. The physical association of MLP and PAL with subsets of lipoprotein particles containing LPS and apoA 1 and the distribution of MLP and PAL in organs and cells in tissues will be studied over time. The second specific aim is to compare the effects of serum released forms of MLP and PAL with chemically purified forms of each in vitro and in vivo. Effects will be evaluated using both macrophages and endothelial cells because these cells integrate the inflammaory response in sepsis. Bacterial strains deficient in MLP and PAL, and mice deficient in TLR2 and TLR4, will be used to distinguish the contributions of cellular activation by each bacterial lipoprotein and by LPS. Our third specific aim is to study the effect of anti-MLP IgG on the processes above and on survival in the two models. The experiments are designed to evaluate the importance that physiologically relevant forms of MLP and PAL play in the pathophysiology of sepsis during infection.

描述（由申请人提供）：在感染过程中，膜成分是从包括LPS和两个细菌膜脂蛋白的复合物中的细菌中脱离的：Murein脂蛋白（MLP）和肽聚糖相关的脂蛋白（PAL）。该更新提案旨在研究这些细菌膜成分彼此和宿主脂蛋白中的每种物理相互作用，以及每个细菌膜成分在激活细胞和全身性炎症反应中的相对贡献，这些反应是通过Toll样受体（TLR）介导的。我们的中心假设是MLP和PAL有助于败血症的发病机理，并且它们在体内的活性可能会因与LPS，LPS以及宿主蛋白和脂蛋白的相互作用而改变。尽管大多数以前的工作都研究了化学纯化或合成的TLR激动剂，但这项工作将尽可能研究MLP和PAL的自然形式。第一个具体目的是研究MLP和PAL从细菌细胞壁释放到细菌膜复合物中的动力学，并在血清中的低密度形式中。体外研究将利用人类血清。体内研究将在小鼠的两种模型中进行：腹膜大肠杆菌感染以及盲肠结扎和穿刺。 MLP和PAL与含有LPS和APOA 1的脂蛋白颗粒亚集的物理关联以及组织中器官和细胞中MLP和PAL的分布将随着时间的流逝而进行研究。第二个具体目的是将释放的MLP和PAL形式的影响与每个体外和体内的化学纯化形式进行比较。将使用巨噬细胞和内皮细胞来评估效果，因为这些细胞整合了败血症中的炎症反应。细菌菌株缺乏MLP和PAL，而缺乏TLR2和TLR4的小鼠将使用每种细菌脂蛋白和LPS来区分细胞活化的贡献。我们的第三个具体目的是研究抗MLP IgG对这两个模型中上述过程和生存过程的影响。该实验旨在评估MLP和PAL在感染过程中败血症的病理生理学中的生理相关形式的重要性。