Selective Modulation of Neutrophils in Critical Illness

项目摘要 护理重症患者占美国所有医疗保健支出的5-10%，而住院死亡率 仍然很高（20-40%）。然而，目前除了支持性护理外，还没有有效的治疗干预措施。 和抗生素。开发新的治疗策略需要填补我们知识中的重要空白 危重病人免疫功能障碍的病理生理机制。失血性休克 诱导导致免疫功能障碍的全身反应，使宿主对严重的 继发性感染是医院ICU发病和死亡的重要原因。的重点 在这次MIRA更新中概述的研究计划是了解出血性休克如何改变 中性粒细胞的反应，急性宿主防御细菌/真菌的基本介质。当 调节失调的中性粒细胞过早地部署它们的细胞毒性蛋白酶和反应性蛋白酶的抗微生物武器库， 氧化剂，直接损伤宿主组织。失血性休克后，中性粒细胞变得“致敏”， 随后引起急性肺损伤，再加上他们发现和对抗 感染我们的长期目标是了解中性粒细胞功能障碍的机制， 出血性休克的结果，并确定纠正异常中性粒细胞功能的目标。我们的研究将 询问中性粒细胞反应的三个方面：启动、运输和抗菌功能。我们有 建立了一种快速产生嗜中性粒细胞特异性嵌合小鼠的创新方法， 条件永生化的中性粒细胞祖细胞系的移植。重要的是，这项技术 不需要动物照射，并产生稳定的供体来源的中性粒细胞嵌合体（>40%）， 在功能上等同于内源性中性粒细胞。该系统在遗传上易于处理，使得能够进行体内研究 出血性休克后中性粒细胞功能障碍的机制方面。在项目1中，我们 继续使用我们的临床相关的“两次打击”小鼠模型，通过失血性休克危重病， 继发性呼吸道感染我们将确定特定的免疫调节障碍的潜在机制， 嗜中性粒细胞效应子功能的子集（例如，反应性氧化剂生成、脱粒） 出血性休克，评估新基因在急性肺损伤和细菌清除中的作用。在项目2中， 我们将利用我们的微流控平台来研究模型中中性粒细胞隔离的机制。 肺毛细血管由于肺特别容易受到嗜中性粒细胞介导的损伤， 提供了一种方法来评估新的策略，用于调节异常中性粒细胞运输，有助于 急性肺损伤在项目3中，我们将追求几个目标，以增强 中性粒细胞而不加重对宿主组织的损伤，重点是耐甲氧西林金黄色葡萄球菌。 总之，这些综合方法将通过以下方式解决中性粒细胞病理学的主要知识缺口： 跨越分子、细胞和生物体/疾病水平的实验。

Project Summary Caring for the critically ill accounts for 5-10% of all U.S. healthcare spending, while in-hospital mortality remains high (20-40%). Yet, there are currently no effective therapeutic interventions beyond supportive care and antibiotics. The development of new therapeutic strategies requires filling important gaps in our knowledge of the pathophysiological mechanisms underlying immune dysfunction in the critically ill. Hemorrhagic shock induces a systemic response that results in immune dysfunction, rendering the host susceptible to severe secondary infections that are a significant cause of morbidity and mortality in hospital ICUs. The focus of the research program outlined in this MIRA renewal is in understanding how hemorrhagic shock alters the response of neutrophils, the essential mediators of acute host defense against bacteria/fungi. When dysregulated, neutrophils prematurely deploy their antimicrobial arsenal of cytotoxic proteases and reactive oxidants, directly injuring host tissue. Following hemorrhagic shock, neutrophils become “primed” and subsequently induce acute lung injury, coupled with a significant deficit in their capacity to find and fight infection. Our long-term goals are to understand the mechanisms of neutrophil dysfunction that occurs as a result of hemorrhagic shock and to identify targets for correcting aberrant neutrophil function. Our studies will interrogate three aspects of the neutrophil response: priming, trafficking, and antimicrobial function. We have established an innovative approach for the rapid generation of neutrophil-specific chimeric mice by transplantation of conditionally-immortalized neutrophil progenitor cell lines. Importantly, this technique does not require animal irradiation and yields robust chimerism of donor-derived neutrophils (>40%) that are functionally equivalent to endogenous neutrophils. This system is genetically tractable, enabling in vivo studies into the mechanistic aspects of neutrophil dysfunction following hemorrhagic shock. In Project 1, we will continue using our clinically relevant “two-hit” mouse model of critical illness through hemorrhagic shock and secondary respiratory infection. We will determine the mechanisms underlying the dysregulation of specific subset of neutrophil effector functions (e.g., reactive oxidant generation, degranulation) in response to hemorrhagic shock, evaluating the role of novel genes in acute lung injury and bacterial clearance. In Project 2, we will employ our microfluidic platform to investigate the mechanisms of neutrophil sequestration in model pulmonary capillaries. As the lungs are particularly susceptible to neutrophil-mediated injury, this model provides a means to evaluate new strategies for modulating aberrant neutrophil trafficking that contributes to acute lung injury. In Project 3, we will pursue several targets for enhancing the antimicrobial function of neutrophils without exacerbating injury to host tissue, focusing on methicillin-resistant Staphylococcus aureus. Together, these integrated approaches will address major knowledge gaps in neutrophil pathobiology by spanning the molecular, cellular, and organism/disease levels of experimentation.