NLRP3/Claudin-1信号通路在蛛网膜下腔出血早期脑损伤中作用及机制研究

Claudin-1 is one of the most important components of the tight junctions. Previous studies demonstrated that Claudin-1 might play a role in blood brain barrier (BBB). However, the roles of Claudin-1 in the brain and the neuron are still unclear. Subarachnoid hemorrhage (SAH) is demonstrated to activate NLRP3 inflammasome, which further promote the secretion of interleukin-1 beta (IL-1β). We have found that melatonin improves SAH-induced early brain injury (EBI) by inhibiting the activation of the NLRP3 inflammasome. However, the downstream mechanisms after the activation of the NLRP3 inflammasome are still unclear. Notably, Claudin-1 expression is regulated by IL-1β, and we further found that ① SAH induced the down-regulation of Claudin-1 in the neurons, and the knockout of Claudin-1 exacerbated SAH-induced EBI; ② Accompanying with the upregulation of Claudin-1, inhibiting the activation of the NRLP3 inflammasome attenuated SAH-induced EBI; ③ Melatonin attenuated SAH-induced EBI, accompanying with upregulating Claudin-1. Collectively, NLRP3/Claudin-1 signaling may play a vital role in SAH-induced EBI. In this project, we will employ Claudin-1 gene knockout and transgenic mice and cell model to explore the roles of NLRP3/Claudin-1 signaling during the protection of melatonin against SAH-induced EBI. This project will provide sufficient theoretical basis for the further development of melatonin-based brain-protective treatment.

Claudin-1蛋白是维持血脑屏障完整的关键分子，但Cluadin-1在大脑实质中的作用不明。蛛网膜下腔出血（SAH）可激活NLRP3炎性小体，进而促进IL-1β分泌，我们证实褪黑素通过抑制NLRP3激活而减轻SAH后大脑实质早期脑损伤，但NLRP3作用的下游机制不清。研究证实IL-1β可以调节Claudin-1表达水平，且我们近期发现：①SAH损伤后，大脑神经元中Claudin-1表达下调，抑制Claudin-1加重早期脑损伤；②抑制NLRP3可减轻早期脑损伤，并伴Claudin-1表达上调；③褪黑素抗早期脑损伤伴随Claudin-1表达上调。以上证据强烈提示NLRP3/Claudin-1通路在SAH早期脑损伤过程中扮演重要角色。本研究将从在体、细胞和分子水平研究证实NLRP3/Claudin-1信号通路在SAH早期脑损伤中的作用及其上下游关系，同时关注该通路在褪黑素抗SAH中的作用。

Claudin-1蛋白是维持血脑屏障（blood brain barrier， BBB）完整的关键分子，但Cluadin-1在蛛网膜下腔出血(subarachnoid hemorrhage, SAH)后血脑屏障中的损伤作用不明。蛛网膜下腔出血（SAH）可激活NLRP3炎性小体，进而促进IL-1β分泌，我们证实褪黑素通过抑制NLRP3激活而减轻SAH后大脑实质早期脑损伤，但NLRP3作用的下游机制不清。研究证实IL-1β可以调节Claudin-1表达水平，我们的研究通过在体及立体SAH模型，行为学，免疫组化，基因敲减和过表达，激光共聚焦，WB等实验手段证实：①SAH损伤后，大脑神经元以及内皮细胞中Claudin-1表达下调，抑制Claudin-1加重早期脑损伤；②抑制NLRP3可减少下游caspase-1和IL1-beta的激活，减轻早期脑损伤包括神经功能缺失和血脑屏障损伤，并伴Claudin-1表达上调；③SAH后褪黑素可以减少早期脑损伤，包括神经功能缺失，脑水肿，以及血脑屏障损伤。此过程中我们也发现了一些保护血脑屏障的药物，以及血脑屏障损伤的新机制如自噬等，NLRP3激活的一些辅助分子以及血脑屏障炎性损伤和神经元凋亡的机制。以上证据强烈提示NLRP3/Claudin-1通路在SAH早期脑损伤过程中扮演重要角色。本研究从在体、细胞和分子水平研究证实NLRP3/Claudin-1信号通路在SAH早期脑损伤中的作用及其上下游关系，阐释了褪黑素抗在SAH后通过减少NLRP3激活进而阻止claudin-1降解保护血脑屏障的的作用。