巨噬细胞分泌包含GBP2的外泌体激活NLRP3炎症小体诱导II 型肺泡上皮细胞焦亡：急性肺损伤的新机制

Cell communication via exosomes has been a research highlight recently. Lipopolysaccharide (LPS) is proved to be one of the noxious substances causing cell injury and inflammatory in alveolar type II cells (AT-II) through activating macrophages, in a result, aggravating acute lung injury (ALT). However, how exosomes mediate the communication between macrophages and AT-II is largely unknown. Our previous study has confirmed that LPS was capable of increasing the expression of GBP2 in exosomes derived from macrophages, as well as activating NLRP3 inflammasomes in AT-II and lung tissues of mouse, which gave rise to the ALT. Nevertheless, the underlying mechanism remains unclear. Consequently, to explore the underlying mechanism, we hypothesize that exosomes containing GBP2 released from LPS-pretreated macrophages could directly activate NLRP3 inflammasomes via the positive and negative interactions to initiate AT-II pyroptosis, leading to the exacerbation of ALT in mouse. In this context, we will perform in vivo and in vitro experiments to explore the mechanism in which TMBIM1 regulates the GBP2-abandunt-exosomes secretion from the macrophages, as well as the pyroptosis resulted from the interaction between GBP2 and NLRP3 inflammasomes at protein level. Additionally, Angiotensin (1-7), which has been demonstrated to decrease NLRP3 inflammasomes, will be introduced to test if it could inhibit the above-mentioned pathological process of ALT induced by LPS. This project provides a novel strategy for acute lung injury prevention and therapy.

外泌体介导的细胞间对话是目前的研究热点。脂多糖（LPS）是促急性肺损伤(ALI)的重要物质，通过活化巨噬细胞（MΦ），引起II型肺泡上皮细胞（AT-II）损伤及炎性反应，促进ALI的发生。但是，外泌体如何介导上述两种细胞间的对话鲜见报道。我们的前期研究发现：LPS可诱导MΦ分泌的外泌体中GBP2含量增加；LPS诱导的MΦ外泌体可激活AT-II 及小鼠肺脏NLRP3炎症小体，并诱发小鼠ALI，然而机制不明。因此，我们设想：LPS诱导MΦ释放包含GBP2的外泌体，直接激活AT-II细胞NLRP3炎症小体，引起焦亡从而促进ALI。本研究拟从体内外层面探讨TMBIM1调控巨噬细胞分泌GBP2外泌体的机理；从蛋白结构层面分析GBP2通过正负电荷吸引与AT-II 内NLRP3炎症小体相互作用并诱导细胞焦亡；血管紧张素 (1-7) 如何抑制上述效应。本课题为ALI的防治提供新策略。

脓毒症被认为是机体面对感染时反应失调而导致的危及生命的器官功能障碍。在脓毒症期间，肺脏是最容易受到感染的器官。然而，对于脓毒症相关的急性肺损伤进展过程中炎症的分子机制研究尚不明确。深入探索脓毒症相关的急性肺损伤发病机制，对于提出新的诊疗方法具有重要意义。活化的巨噬细胞可释放大量炎症介质作用于周围的细胞及微环境，进一步加剧炎症风暴，促进脓毒症相关的急性肺损伤的发生发展。外泌体作为一种新兴的细胞间的交流方式，内含大量物质，可通过内容物引起下游受体细胞的反应，是目前研究细胞间通讯的热点。.本课题从体内、体外⑴探讨脂多糖诱导巨噬细胞释放的外泌体中的成分及其对下游Ⅱ型肺泡上皮细胞功能的影响，探索外泌体在Ⅱ型肺泡上皮细胞炎症小体激活过程中的作用机制。⑵探索鸟苷酸结合蛋白2（GBP2）在脂多糖诱导的脓毒症相关的肺损伤中的作用及机制，深入探索GBP2通过以外泌体为载体从免疫细胞释放作用于Ⅱ型肺泡上皮细胞，且可通过自身的异戊二烯化能力影响NLRP3炎症小体的激活。体外中，脂多糖诱导巨噬细胞释放的外泌体可引起Ⅱ型肺泡上皮细胞中的NLRP3炎症小体激活及细胞焦亡的发生，外泌体中的鸟苷酸结合蛋白2明显升高，且是引起Ⅱ型肺泡上皮细胞炎症小体激活的主要成分，同时鸟苷酸结合蛋白2可直接通过自身的异戊二烯化能力结合NLRP3炎症小体的PYD结构域，进而诱导Ⅱ型肺泡上皮细胞的活化。体内中，在脂多糖诱导的小鼠急性肺损伤模型中，鸟苷酸结合蛋白2可从巨噬细胞转移至Ⅱ型肺泡上皮细胞，且可明显导致脓毒症相关的急性肺损伤发生发展。综合体内体外结果，我们得出：⑴脂多糖诱导巨噬细胞释放大量的外泌体可在体内体外激活Ⅱ型肺泡上皮细胞中的NLRP3炎症小体，促进炎症的发生发展；⑵且外泌体内的鸟苷酸结合蛋白2作为主要成分直接发挥作用，促进急性肺损伤的发生发展。