CD80 and CD86 mediated innate immune response in sepsis

描述（由申请人提供）：脓毒症（对感染的全身性炎症反应）的发病率为75万例，是危重患者死亡的主要原因。脓毒症的治疗是有限的，在很大程度上仍然是支持性的。加深对脓毒症炎症机制的理解，并对旨在降低死亡率的干预措施进行临床前研究，是NHLBI使命的一部分。初步研究表明，共刺激分子CD80和CD86在败血症的先天免疫应答中起重要作用。CD80/86-/-小鼠在盲肠结扎和穿刺（CLP）引起的多微生物脓毒症后生存率提高，炎症细胞因子产生减少，NF-kappaB活化减少。中性粒细胞(PMN)/巨噬细胞共培养的体外模型通过CD80/86依赖途径导致巨噬细胞活化。在脓毒症的临床研究中，我们观察到脓毒症患者PMN中CD28（一种CD80/86配体）的表达增加，死亡率与可溶性CD28水平相关。我们假设巨噬细胞表达CD80/86参与了败血症的先天免疫反应。为了确定CD80和CD86的具体重要性，我们将使用先天缺乏这些分子的小鼠以及siRNA和抑制性抗体来调节巨噬细胞中CD80和CD86的表达。我们将通过流式细胞术和共聚焦显微镜研究CD80/86系统在人类败血症中的表达，并比较人和小鼠的调节，以验证CLP模型。然后，我们将在体外分析正常和败血症人类受试者的PMNs对巨噬细胞的影响，并在共培养实验中使用siRNA和阻断抗体评估CD80和CD86的作用。这是一个败血症调查和培训的建议，其中包括完成临床调查科学硕士学位。在Weiden博士的指导下完成的课程和在实验室的经历，对于培养我在模型系统中识别病理生理机制的能力至关重要。这项资助将促进一个专注于了解脓毒症炎症机制的职业，目标是开发降低这一重要疾病死亡率的干预措施。

DESCRIPTION (provided by applicant): Sepsis (systemic inflammatory response to infection) has an incidence of 750,000 cases and is the leading cause of death in critically ill patients. Treatment of sepsis has been limited and remains largely supportive in nature. Improved understanding of the mechanisms underlying inflammation in sepsis and preclinical investigation of interventions designed to reduce mortality are part of the NHLBI mission. Preliminary studies show that the costimulatory molecules, CD80 and CD86 are important in the innate immune response to sepsis. CD80/86-/- mice have improved survival, reduced inflammatory cytokine production and less NF-kappaB activation after polymicrobial sepsis produced by cecal ligation and puncture (CLP). An in vitro model using neutrophil (PMN)/macrophage co-culture leads to macrophage activation by a CD80/86 dependent pathway. During clinical investigation of sepsis we observed that PMN from septic humans have increased expression of a CD28 (a CD80/86 ligand) and mortality correlated with soluble CD28 levels. We hypothesize that macrophage expressed CD80/86 are involved in the innate immune response to sepsis. To determine the specific importance of CD80 and CD86 we will use mice congenitally deficient in these molecules as well as siRNA and inhibitory antibodies to modulate CD80 and CD86 expression in macrophages. We will investigate the expression of the CD80/86 system in human sepsis by flow cytometry and confocal microscopy and compare regulation in humans and mouse to validate the CLP model. We will then assay the effect of PMNs from normal and septic human subjects on macrophages in vitro and assess the role CD80 and CD86 using using siRNA and blocking antibodies in co-culture experiments. This is a proposal for investigation in sepsis and training which includes a completion of a Masters of Science in Clinical Investigation.The course work and the experience in the laboratory under the mentorship of Dr. Weiden are essential for developing my abilities in identifying pathophysiologic mechanisms in model systems. This grant will foster a career focused on understanding the mechanisms underlying inflammation in sepsis with the goal of developing intervensions that reduce mortality in this important disease.