Genetic mechanisms regulating inflammation and neutrophil activity in zebrafish

描述(申请人提供)：在检测到感染或损伤后，免疫系统的吞噬细胞进行炎症反应，摧毁入侵的微生物，清除死亡细胞中的碎片，并恢复体内平衡。特别是，中性粒细胞在炎症反应的每个阶段都发挥着关键作用--它们是第一批被招募到感染或损伤部位的细胞；它们杀死微生物，吞噬碎片，并协调其他免疫细胞的活动；它们的凋亡触发炎症的消退和组织内稳态的恢复。炎症对于抵抗感染和开始愈合是必不可少的，但长期或过度活跃的炎症可能会对健康细胞造成伤害。此外，慢性炎症会加剧给社会带来巨大负担的疾病，如糖尿病、动脉粥样硬化和癌症。我们对控制炎症的启动和消退的机制的不完全理解阻碍了我们朝着治疗这些衰弱疾病的方向前进。该项目的长期目标是通过分析中性粒细胞异常的斑马鱼突变体，发现在调节炎症方面具有必要功能的新基因。中性粒细胞数量的增加和中性粒细胞向周围组织的募集是炎症的保守特征，而中性粒细胞的活跃反应是炎症的一个强有力的指标。我们建议利用斑马鱼遗传学的力量来了解炎症和中性粒细胞活动的控制。通过对一个具有高炎症表型的斑马鱼突变体的分析，我们发现了一个在没有感染或损伤的情况下预防炎症所必需的新基因。有了以下三个具体目标，我们建议分离更多扰乱炎症和中性粒细胞活性的突变，并在细胞和分子水平上研究这些基因的功能。这些研究将确定重要的炎症反应的新调节因素，从而可能为炎症的治疗控制提供新的途径。目的1：我们将使用中性粒细胞特异的转基因报告进行基因筛查，以寻找在中性粒细胞的发育、迁移和活动中具有重要功能的新基因的突变。目的：为了在细胞水平上确定突变基因的功能，我们将通过分析炎症信号，确定中性粒细胞的哪些方面的活性和发育被破坏，以及测试中性粒细胞对组织损伤的反应来表征突变的表型。目的3：克隆突变基因并分析其产物的分布和活性，以期发现突变基因的生化功能。

DESCRIPTION (provided by applicant): After detecting infection or injury, the phagocytic cells of the immune system carry out an inflammatory response, which destroys invading microorganisms, clears debris from dead cells, and restores homeostasis. Neutrophils, in particular, have a critical role at every stage of the inflammatory response-they are the first cels to be recruited to the site of infection or injury; they kill microbes, engulf debris, and orchestrte the activities of other immune cells; and their apoptosis triggers resolution of inflammation and the restoration of tissue homeostasis. Inflammation is essential to fight infection and initiate healing, but prolonged or hyperactive inflammation may cause harm to healthy cells. Furthermore, chronic inflammation exacerbates diseases that place a huge burden on society, such as diabetes, atherosclerosis, and cancer. Our incomplete understanding of the mechanisms that control the initiation and resolution of inflammation has hindered our progress toward therapies for these debilitating conditions. The long-term goal of this project is to discover new genes with essential functions in regulating inflammation by analyzing zebrafish mutants with abnormalities in neutrophils. An increase in neutrophil number and recruitment of neutrophils to peripheral tissues are conserved hallmarks of inflammation, and an active neutrophil response is a robust indicator of inflammation. We propose to exploit the power of zebrafish genetics to understand the control of inflammation and neutrophil activity. Through the analysis of a zebrafish mutant with a hyperinflammatory phenotype, we have discovered a new gene that is essential to prevent inflammation in the absence of infection or injury. With the following three Specific Aims, we propose to isolate more mutations that disrupt inflammation and neutrophil activity, and to study the functions of these genes at the cellular and molecular level. These studies will define important new regulators of the inflammatory response and may thereby provide new avenues toward therapeutic control of inflammation. Aim 1: We will conduct a genetic screen with a neutrophil-specific transgenic reporter for mutations in new genes with essential functions in development, migration, and activity of neutrophils. Aim 2: To determine the functions of the mutated genes at the cellular level, we will characterize the mutant phenotypes by analyzing inflammatory signaling, determining which aspects of neutrophil activity and development are disrupted, and testing the neutrophil response to tissue injury. Aim 3: To discover the biochemical functions of the mutated genes, we will clone the mutated genes and analyze the distribution and activities of their products.