Maturation of IL-1beta and IL-18 in novel macrophage aggresomes

描述（由申请人提供）：白细胞介素(IL)-12和IL-18是促进炎症反应的关键介质。局部炎症反应是限制病原体感染和促进伤口愈合的重要机制。然而，过度和不受控制的IL-12和IL-18的产生以及导致的不适当的炎症与组织破坏和越来越多的炎症性疾病的衰弱症状有关。巨噬细胞中IL-12和IL-18的加工和释放是对炎症小体形成的反应。炎性小体是由淋巴结样受体（NLR）家族的细胞质模式识别受体激活，以响应病原体或宿主来源的应激信号。激活的NLRs通过必需接头蛋白ASC （PYCARD）募集前caspase-1，导致caspase-1激活并释放IL-12和IL-18。控制炎性小体形成和激活的分子机制尚不清楚，但有很高的潜力为干预IL-12和IL-18释放的新策略提供基础，以治疗炎症性疾病。我们提出的研究的中心假设是，炎症小体代表巨噬细胞中可诱导的、特化的细胞质结构，其中炎症小体成分被特异性招募来激活caspase-1。我们的假设是基于巨噬细胞炎性小体的初步成像结果，在本研究中，我们提出解决诱导炎性小体形成的机制，并将其与IL-12和IL-18释放机制联系起来。我们的假设是基于我们的初步发现，表明(1)炎性小体需要炎性小体成分的可诱导再分配；(2)炎性小体在巨噬细胞胞浆中可诱导形成；(3)炎性小体含有与特化细胞超微结构直接相关的特征性标记蛋白。我们提出了两个特定目标：特定目标#1将在巨噬细胞中建立炎性小体作为独特的结构，而特定目标#2将确定破坏这些结构形成对IL-12和IL-18成熟的影响。在这项研究完成后，我们期望提供目前难以捉摸的炎症小体在感染和应激反应中聚集的机制，这将为抑制IL-12和IL-18的成熟开辟新的途径。这项研究将有助于我们更好地理解导致炎症途径激活的先天免疫反应的分子机制，从而开发出治疗炎症和感染性疾病的改进疗法。公共卫生相关性：IL-12和IL-18的过量产生直接导致许多具有破坏性发病机制的炎症性疾病的症状，包括工业化国家的一些最常见疾病，包括关节炎、哮喘、炎症性肠病、溃疡性结肠炎、动脉粥样硬化、牙周炎、2型糖尿病、肺纤维化、多发性硬化症、阿尔茨海默病或中风。目前没有有效的治疗方法，导致患者终身症状，并对我们的社会和医疗系统造成巨大的经济和财政影响。IL-12和IL- 18需要特殊的蛋白质复合物（称为炎性小体）才能释放，但炎性小体的组装机制尚不清楚，但有望为炎症性疾病患者的治疗方案提供基础。因此，在本研究中，我们建议解决炎症小体形成的机制，并将炎症小体与现有的细胞超结构联系起来，这有望为防止IL-12和IL-18的失控释放开辟新的途径。

DESCRIPTION (provided by applicant): Interleukin (IL)-12 and IL-18 are key mediators promoting inflammatory reactions. Localized inflammatory responses are important mechanisms to limit pathogen infections and to promote wound healing. However, excessive and uncontrolled production of IL-12 and IL-18 and resulting inappropriate inflammation is linked to tissue destruction and the debilitating symptoms of the growing number of inflammatory diseases. Processing and release of IL-12 and IL-18 occurs in macrophages in response to formation of inflammasomes. Inflammasomes are activated by cytosolic pattern recognition receptors of the Nod-like receptor (NLR) family in response to pathogen or host derived stress signals. Activated NLRs recruit pro-caspase-1 via the essential adaptor protein ASC (PYCARD), resulting in caspase-1 activation and release of IL-12 and IL-18. The molecular mechanisms that control inflammasome formation and activation are poorly understood, but have a high potential to provide the basis for novel strategies to interfere with IL-12 and IL-18 release for the treatment of inflammatory diseases. Our central hypothesis for the proposed study is that inflammasomes represent inducible, specialized cytosolic structures in macrophages, where inflammasome components are specifically recruited to activate caspase-1. Our hypothesis is based on preliminary imaging results from inflammasomes in macrophages, and in this study we propose to address the mechanism of inducible inflammasome formation and link it to the IL-12 and IL-18 release mechanism. Our hypothesis is based on our preliminary findings showing that (1) inflammasomes require inducible redistribution of inflammasome components; (2) inflammasomes are inducible formed in the cytosol of macrophages; (3) inflammasomes contain characteristic marker proteins that directly link it to specialized cellular ultrastructures. We propose two specific aims: Specific aim #1 will establish inflammasomes as distinct structures in macrophages, while specific aim #2 will determine the impact on IL-12 and IL-18 maturation as a consequence of disrupting formation of these structures. At the completion of this study, we expect to provide the currently elusive mechanism by which inflammasomes assemble in response to infection and stress, which will open new avenues for inhibiting maturation of IL-12 and IL-18. This study will contribute to our long-term goal to contribute to a better understanding of molecular mechanisms of innate immune responses leading to inflammatory pathway activation, in order that improved therapies to treat inflammatory and infectious diseases can be developed. Public Health Relevance: Excessive production of IL-12 and IL-18 are directly responsible for the symptoms of a number of inflammatory diseases with destructive pathogenesis, including some of the most common diseases of industrialized nations, including arthritis, asthma, inflammatory bowel disease, ulcerative colitis, atherosclerosis, periodontitis, type 2 Diabetes, lung fibrosis, multiple sclerosis, Alzheimer's disease, or stroke. Currently there are no effective treatments available, causing patients life-long symptoms and a huge economical and financial impact on our social and medical systems. IL-12 and IL- 18 require specialized protein complexes, referred to as inflammasomes for release, but the mechanism by which inflammasomes assemble are not well understood, but are expected to provide the basis for the development of treatment options for patients suffering from inflammatory diseases. As a consequence, in this study we propose to address the mechanism of inflammasome formation, and to link inflammasomes to existing cellular ultra structures, which is expected to open new avenues for preventing uncontrolled release of IL-12 and IL-18.