Project 2

摘要 包括铜绿假单胞菌(PA)在内的各种病原体引起的医院感染 在美国，巨大的经济负担。已经开发出有效的感染治疗方法 由于对感染目标的了解有限而受到阻碍。研究黄褐斑形成的分子机制 肺泡巨噬细胞(AM)对抗PA感染可能有助于发现新的治疗方法 接近了。自噬是一种细胞成分被隔离到 通过泛素化降解的自噬小体。我们最近发现帕金森杆菌感染 可以诱导自噬，并随后增加细菌的降解。Toll样受体 (TLRs)与自噬和巨噬细胞吞噬有关。有趣的是，我们的 初步数据表明，Lyn与TLR-2相互作用，在促进 吞噬和杀菌活性。此外，LYN还可能在PAG-7期间与ATG-7相互作用 感染。最后，自噬抑制剂3-甲基腺嘌呤(3MA)减少了吞噬功能和 AM随后的细菌清除在这里，我们假设自噬增强了 吞噬功能，增加细菌清除，降低炎症反应。这个 本研究的目的是研究Lyn-ATG7-ATG7细胞自噬的生理意义。 基因敲除(KO)小鼠的吞噬功能调节，其炎症反应可能是 抑制导致较少的组织损伤。我们的长期目标是确定 AM对PA入侵的防御，以设计新的策略来治疗这种感染。这个 理论基础是，这项研究的实施将把自噬定义为一种新的防御机制 抗PA感染，从而提出新的治疗靶点。具有雄厚的科学基础 有了现有的数据和其他资源，我们能够很好地测试以下具体目标。 具体目的1：剖析自噬对PA吞噬效率的影响。我们 假设自噬会通过影响吞噬功能来增加细菌的清除。 小鼠和原代AM细胞将被用来研究吞噬杯中自噬的作用 形成和细菌清除。具体目标2：研究LYN在启动和释放过程中的活性 执行自噬途径。我们假设，通过传输TLR2信号，LYN是一个 PA诱导自噬的关键调控因子。这一信号通路的激活有望 在一种新发现的免疫中增加自噬小体的形成和成熟 在寄主中抵抗入侵PA的机制。具体目标3：确定ATG-7在 减轻小鼠的炎症。我们假设自噬可以控制炎症 过程，从而限制感染小鼠的组织损伤。使用一种新的动物成像技术 包括新型发光试剂，我们将区分早期和晚期的炎症反应 一个。

Summary Hospital-acquired infections due to various pathogens including P. aeruginosa (Pa) impose huge financial burdens in the USA. Development of effective therapeutics for infection has been hampered due to limited knowledge in infection targets. Studying the molecular mechanisms of alveolar macrophages (AM) in fighting Pa infection may help discover novel therapeutic approaches. Aautophagy is a mechanism by which cellular components are sequestered to autophagosomes for degradation through ubiquitination. We recently revealed that Pa infection can induce autophagy, and subsequently increasing bacterial degradation. Toll like receptors (TLRs) are implied in linking autophagy with macrophage phagocytosis. Interestingly, our preliminary data indicate that Lyn, interacting with TLR-2, plays a critical function in facilitating phagocytosis and bactericidal activity. Moreover, Lyn may also interact with Atg-7 during Pa infection. Finally, autophagy inhibitor 3-methyladenine (3MA) decreased phagocytosis and subsequent bacterial clearance by AM. Herein, we hypothesize that autophagy enhances phagocytic function, increases bacterial clearance, and lowers inflammatory responses. The objective of this proposal is to examine the physiological significance of autophagy in Lyn-Atg7- modulated phagocytosis in knockout (KO) mice, whose inflammatory responses may be suppressed leading to less tissue injury. Our long-term goal is to identify the mechanisms of AM defense against Pa invasion in order to design novel strategies to treat this infection. The rationale is that execution of this research will define autophagy as a novel defense mechanism against Pa infection, thus suggesting new therapeutic targets. With the strong basis of scientific data and other resources available, we are well positioned to test the following specific aims. Specific Aim 1: Dissect the effects of autophagy on Pa phagocytosis efficiency. We hypothesize that autophagy will increase bacterial clearance by influencing phagocytic function. Mice and primary AM cells will be used to study the role of autophagy in phagocytic cup formation and bacterial clearance. Specific Aim 2: Study the activity of Lyn in initiating and executing autophagic pathway. We hypothesize that Lyn, by transmitting TLR2 signals, is a key regulator in Pa-induced autophagy. Activation of this signaling pathway is expected to increase the formation and maturation of autophagosomes in a newly identified immunity mechanism to combat invading Pa in the host. Specific Aim 3: Identify the role of Atg-7 in alleviating inflammation in mice. We hypothesize that autophagy can control the inflammatory process, thereby limiting tissue damage of infected mice. Using a novel animal imaging that includes novel luminescent reagents, we will distinguish early inflammatory responses from late ones.