PARP调节NLRP3炎症小体介导的创伤性脑损伤固有免疫应答的机制研究

NLRP3 inflammasome is a multiprotein complex, which consists of NLRP3, apoptosis-associated speck-like protein containing CARD (ASC), and pro-caspase-1. NLRP3 inflammasome is one of key factors in innate immune response through mediating the activation of caspase-1 and the processing and release of interleukin-1β (IL-1β) and interleukin-18 (IL-18), and is reported to be involved in traumatic brain injury. Under pathological conditions, over-activation of poly(ADP-ribose) polymerase (PARP) results in the mitochondrial damage, promotes the release of high mobility group box-1 protein (HMGB1), enhances intracellular calcium, and up-regulates the expression of NLRP3 inflammasome proteins, by which, participating the activation of NLRP3 inflammasome. In vivo the activation of NLRP3 inflammasome is reported during traumatic brain injury. In previous study, we have found that PARP inhibition down-regulates the expression of NF-kB-driven inflammatory mediators, and protects brain against traumatic brain injury in mice. Therefore, we hypothesize that PARP, as a regulator of inflammatory reaction, would initiate the innate immune response and inflammatory reaction through following mechanisms: (1) protmoting the combination of NF-kB with DNA, by which, enhancing the transcriptional activity of NF-kB；(2) mediating the poly(ADP-ribosyl)ation of mitochondrial proteins, leading to the mitochondrial damage; (3) enhancing the release of HMGB1, subsequently, activating the NF-kB signal；(4) inducing the expression of TRPM2 and AMPA receptor, by which, resulting in the increased intracellular calcium and the production of reactive oxygen species. The focus of brain concussion is composed of traumatic core and surrounding penumbra. This project intends to investigate whether PARP would modulate NLRP3 inflammasome signal through influencing the mitochondrial function, intracellular calcium, reactive oxygen production, and HMGB1 and NF-kB signals in various cell types (including neuron, astrocyte, and microglia) in penumbra and core in a mouse models of traumatic brain injury by means of the approaches of pharmacology, biochemistry, molecular biology, immunology, and morphology, by which, demonstrating our hypothesis, elucidating the new mechanism of the evolution and reversion of traumatic penumbra, and providing new target and strategy for the therapy of traumatic brain injury.

NLRP3炎症小体是机体固有免疫的关键因子之一。离体研究发现PARP的过度活化能引起线粒体损伤，升高细胞内钙，促进HMGB1的释放以及NF-B介导的NLRP3炎症小体蛋白的表达等，参与NLRP3炎症小体的激活。整体动物研究证实创伤性脑损伤(TBI)能引起NLRP3炎症小体信号的激活。我们前期工作证实PARP抑制剂能下调TBI后NF-B介导的炎症介质的表达，降低TBI继发性损伤。因此，我们推测，作为一种炎症反应调节蛋白，PARP可能通过影响NLRP3炎症小体信号，启动TBI的固有免疫应答和炎症反应。本研究拟在整体动物水平，研究创伤半暗带和核心区PARP对NLRP3炎症小体信号的影响及其机制，阐述PARP信号和NLRP3炎症小体信号在TBI固有免疫应答中的作用，以更深入地理解TBI继发性损伤的病理生理机制以及创伤半暗带发生、发展和逆转的新机制，为TBI的诊治提供新靶点和新策略。

脑损伤极大地影响了患者的生活质量，同时给社会造成了严重的经济负担，然而由于继发性损伤机制的复杂性，到目前为止仍没有一种有效的治疗方式。越来越多的研究表明炎症在继发性脑损伤中发挥着重要作用，本研究基于CCI模型，结合分子细胞生物学和动物实验探讨了Nlrp3炎症小体在继发性脑损伤中的作用。研究结果表明Nlrp3炎症通路在TBI后显著性激活，大脑皮层神经元数目显著性降低以及胶质细胞激活。当敲除Nlrp3或Caspase1后，神经炎症程度减弱，而且神经元数目显著性增加。此外，我们还发现Drp1介导的线粒体分裂促进了线粒体自噬在TBI后的神经保护作用。本研究揭示了炎症通路在继发性损伤中的作用机制，为TBI的治疗提供了新的思路和理论基础。