NF-kB and Chromatin Changes in Human Sepsis

该建议继续我们的目标，阐明控制急性全身性炎症的分子事件，这是一个高度致命的过程。在之前的资助周期中，我们在先天免疫巨噬细胞和中性粒细胞中发现了Toll样受体（TLR）驱动的、暂时定义的表观遗传程序，该程序指导急性全身炎症的过程。这个过程改变染色质结构来抑制或激活不同的基因功能集，从而产生不同的表型阶段。其他人和我们已经证明，在急性全身性炎症期间，生物能量转移也发生在先天免疫细胞中。这一应用发展了统一的概念，即生物能量学的修改与表观遗传学相结合，以指导急性炎症的相移。我们提出了两个具体目标：目的1)明确生物能源与表观遗传学之间的联系。我们将使用复制tlr4依赖性相移的细胞模型来：a)描述基因特异性染色质修饰；B)确定蛋白质之间的相互作用；C)通过质谱法评估代谢谱；c)将我们的概念转化为人类正常和败血症的血液白细胞。目的2)测试修改生物能源和表观遗传转移对临床结果的影响。我们将建立小鼠脓毒症模型：a)定义生物能量和表观遗传相移，使用生化和遗传方法分析血浆和脾脏、腹膜和骨髓分离的白细胞；b)使用药理学和遗传学方法来确定进入适应期是否会改变微血管炎症和生存。我们的研究结果将对炎症领域产生广泛的影响，为急性炎症的发生提供新的见解，并为新的治疗方法提供信息。

The proposal continues our objective to elucidate the molecular events that control acute systemic inflammation, a highly lethal process. In the previous grant cycle, we discovered a Toll like receptor (TLR) - driven, temportally-defined, epigenetic program in innate immunity macrophages and neutrophis, which directs the course of acute systemic inflammation. This process modifies chromatin structure to repress or activate distinct functional sets of genes, thus generating distinct phenotypic phases. Others and we have shown that bioenergy shifts also occur in innate immunity cells during acute systemic inflammation. This application develops the unified concept that modifications in bioenergetics integrate with epigenetics to direct the phase shifts of acute inflammation. We propose two specific aims: Aim 1) To define the connections between bio-energy and epigenetics. We will use a cell model that reproduces TLR4-dependent phase shifts to: a) delineate gene-specific chromatin modifications; b) determine protein-protein interactions; c) assess metabolic profiles by mass spectroscopy; and c) translate our concept to human normal and sepsis blood leukocytes. Aim 2) To test effects of modifying bio-energy and epigenetic shift on clinical outcomes. We will us a murine model of sepsis to: a) define bio-energy and epigenetic phase shifts, using biochemical and genetic methods to analyze plasma and isolated leukocytes from spleen, peritoneum, and bone marrow; and b) use pharmacologic and genetic approaches to determine whether modifying to adaptive phase alters microvascular inflammation and survival. Our result will broadly impact the field of inflammation by providing new insight on how acute inflammation is orchestrated, and inform novel therapies.