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。脓毒症的治疗一直是有限的，在本质上仍然是支持性的。提高对脓毒症潜在炎症机制的理解和对旨在降低死亡率的干预措施的临床前调查是NHLBI任务的一部分。初步研究表明，共刺激分子CD80和CD86在脓毒症的先天免疫反应中起重要作用。CD80/86-/-小鼠在盲肠结扎穿孔(CLP)所致的多菌败血症后，存活率提高，炎性细胞因子产生减少，核因子-kappaB活性降低。使用中性粒细胞(PMN)/巨噬细胞共培养的体外模型通过CD80/86依赖的途径导致巨噬细胞激活。在脓毒症的临床研究中，我们观察到败血症患者的PMN增加了CD28(CD80/86配体)的表达，并且死亡率与可溶性CD28水平相关。我们推测巨噬细胞表达CD80/86参与了脓毒症的先天免疫反应。为了确定CD80和CD86的特殊重要性，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。我们将利用流式细胞仪和共聚焦显微镜研究CD80/86系统在人类脓毒症中的表达，并比较其在人和小鼠中的表达规律，以验证CLP模型的有效性。然后，我们将在体外检测正常人和脓毒症患者的PMN对巨噬细胞的影响，并在共培养实验中使用siRNA和阻断抗体来评估CD80和CD86的作用。这是一个脓毒症研究和培训的建议，包括完成临床研究的理学硕士学位。课程工作和在魏登博士指导下的实验室经验对于发展我在模型系统中识别病理生理机制的能力是必不可少的。这笔赠款将促进专注于了解脓毒症潜在炎症机制的职业，目标是开发干预措施，降低这种重要疾病的死亡率。

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.