The Role of Necroptosis in Aging

项目总结/摘要 研究目标是表征坏死性凋亡诱导的炎症对衰老的作用。慢性的， 低度炎症（炎症）是衰老的标志，并且是“衰老的七大支柱”之一。 炎症是老年人发病率和死亡率的一个非常重要的危险因素，因为 各种年龄相关疾病（如2型糖尿病、心血管疾病、癌症和神经退行性疾病） 疾病）共享强炎性表型。尽管炎症，衰老和年龄之间存在联系- 与相关疾病有关，目前我们对炎症在衰老中的作用的理解存在两个主要差距： (1)负责慢性低度炎症的分子机制/途径，以及（2）是否 炎症是衰老的一个致病因素或继发于衰老。损伤相关分子模式 DAMPs在年龄相关的慢性炎症中起作用，坏死性凋亡是一种新发现的DAMPs通路。 程序性坏死在DAMP的产生中起主要作用。当坏死性下垂发生时， 刺激依次激活受体相互作用蛋白激酶1（RIPK 1）、RIPK 3和混合谱系激酶 结构域样（MLKL）蛋白。磷酸化MLKL结合并破坏血浆 细胞膜，释放DAMPs。DAMPs反过来又引发慢性低度炎症， 先天免疫细胞产生的炎性细胞因子如TNF α增加，可激活RIPK 1 在其他细胞中形成正反馈回路。研究表明，通过敲除Ripk 3来抑制坏死性凋亡， 坏死性凋亡以及炎症。我们的初步数据提供了第一个证据 显示坏死性凋亡可能在衰老中起作用，即，在野生型（WT）小鼠中，坏死性凋亡随年龄增长而增加 在加速老化模型（Sod 1-/-小鼠）中，减少/阻断坏死性凋亡（遗传和 在Sod 1-/-小鼠中减少炎症。根据我们的初步数据，我们假设， 坏死性凋亡在慢性低度炎症中起作用，这种炎症随着年龄的增长而发生， 坏死性凋亡将减弱炎症，从而延长寿命并改善健康状况。测试 基于这一假设，我们将在Aim 1中通过以下方式确定坏死性凋亡在年龄相关慢性炎症中的作用： 确定青年、中年和老年男性各种组织中坏死性凋亡和炎症的进展 和雌性WT小鼠，并确定在遗传上（Ripk 3 +/-和Ripk 3-/-）减少坏死性凋亡的作用。 小鼠）或RIPK（necrostatin-1 s，一种RIPK 1抑制剂）对炎症的影响;在Aim 2中，我们将确定 坏死性凋亡介导年龄相关性炎症的机制， 炎症通路的年轻和老年WT小鼠的组织和细胞，并确定遗传的影响， 抑制坏死性凋亡对炎症通路的影响;在Aim 3中，我们将确定坏死性凋亡在衰老中的作用 通过比较Ripk 3 +/-和Ripk 3-/-小鼠与WT小鼠的寿命、健康期和年龄相关病理学。

Project Summary/Abstract The research objective is to characterize the role of necroptosis-induced inflammation on aging. Chronic, low-grade inflammation (inflammaging) is a hallmark of aging and is one of the ‘seven pillars of aging’. Inflammaging is a highly significant risk factor for both morbidity and mortality in the elderly people because a variety of age-related diseases (e.g. type 2 diabetes, cardiovascular diseases, cancer, and neurodegenerative diseases) share a strong inflammatory phenotype. Despite the link between inflammation, aging and age- associated diseases, two major gaps currently exist in our understanding of the role inflammation plays in aging: (1) the molecular mechanism(s)/pathway(s) responsible for the chronic, low-grade inflammation and (2) whether inflammaging is a causative factor in aging or occurs secondary to aging. Damage-associated molecular patterns (DAMPs) play a role in age-associated chronic inflammation and necroptosis is a newly identified pathway of programmed necrosis that plays a major in the generation of DAMPs. Necroptosis is initiated when necroptotic stimuli sequentially activate the receptor-interacting protein kinase 1 (RIPK1), RIPK3, and mixed lineage kinase domain like (MLKL) protein through phosphorylation. Phosphorylated MLKL binds to and disrupts the plasma membrane of cells, releasing DAMPs. The DAMPs in turn trigger chronic low-grade inflammation through increased production of inflammatory cytokines such as TNF by innate immune cells, which can activate RIPK1 in other cells in a positive feedback loop. Studies show that inhibiting necroptosis by knocking out Ripk3 reduces necroptosis as well as inflammation in several mouse models. Our preliminary data provides the first evidence showing that necroptosis might play a role in aging, i.e., necroptosis increases with age in wild type (WT) mice and in a model of accelerated aging (Sod1-/- mice), and reducing/blocking necroptosis (both genetically and pharmacologically) reduces inflammation in Sod1-/- mice. Based on our preliminary data, we hypothesize that necroptosis plays a role in chronic, low-grade inflammation, which occurs with age, and preventing necroptosis will attenuate inflammation, leading to increased lifespan and improved healthspan. To test this hypothesis, in Aim1 we will determine the role of necroptosis in age-associated chronic inflammation by identifying progression of necroptosis and inflammation in various tissues of young, middle-aged and old male and female WT mice and determine the effect of reducing necroptosis either genetically (Ripk3+/- and Ripk3-/- mice) or pharmacologically (necrostatin-1s, a RIPK1 inhibitor) on inflammation; in Aim2 we will determine the mechanism by which necroptosis mediates age-associated inflammation by assessing the activation of inflammatory pathways in tissues and cells of young and old WT mice and determine the effect of genetic inhibition of necroptosis on inflammatory pathways; in Aim3, we will determine the role of necroptosis in aging by comparing the lifespan, healthspan and age-associated pathology of Ripk3+/- and Ripk3-/- mice to WT mice.