纳米氧化石墨烯（GO）增强自噬溶酶体功能在改善七氟烷诱导POCD中的作用机制研究

Postoperative cognitive dysfunction (POCD) is a common complication of anesthesia in elderly patients, but its mechanisms are not clear. Previous studies have shown that POCD and Alzheimer's disease (AD) may share similar molecular mechanisms. Intracellular Aβ accumulation caused by autolysosomal function damage accelerates the pathological process of AD and may lead to POCD. Applicant’s pre-experiments found that the general anesthetic sevoflurane could impair the autolysosomal function, while the nano-oxidized graphene (GO) with enhanced autophagic degradation ability was able to restore those damaged autolysosomes in cultured cells in vitro. Therefore, we speculate that GO can improve POCD under sevoflurane anesthesia by enhancing autolysosomal function. In this project, the effects of sevoflurane on autolysosomal function and Aβ clearance will be detected by immunofluorescence, western blot and enzyme-linked immunosorbent assay methods, and combining with lysosomal alkalization drug chloroquine (CQ) treatment, the relationship between autolysosomal function and POCD in aged mice will be detected by Morris water maze method. By targeting autolysosomes, the current study is designed to improve sevoflurane induced POCD through enhancing autolysosomal function by GO, and it may also provide theoretical basis and drug target for prevention and treatment of POCD.

术后认知功能障碍（POCD）是老年患者手术麻醉后的常见并发症，但其机制尚不明确。既往研究表明POCD和阿尔兹海默病（AD）可能具有某些相似的分子机制；自噬溶酶体功能受损致使细胞内Aβ积累可促进AD的病理进展，上述机制或可适用于POCD。申请人预实验发现全身麻醉药七氟烷可损害自噬溶酶体功能，而具有增强自噬降解能力的纳米氧化石墨烯（GO）在体外培养细胞中可恢复受损的自噬溶酶体功能。因此，我们推测：GO可通过增强自噬溶酶体功能改善七氟烷麻醉导致的POCD。本课题拟以七氟烷为模式麻醉药，通过IF、WB、ELISA等手段，检测七氟烷对自噬溶酶体功能、Aβ清除的影响；联合溶酶体碱化药物氯喹（CQ）处理，通过Morris水迷宫手段检测自噬溶酶体功能与老年小鼠POCD的关系；以自噬溶酶体为靶点，期望通过GO上调其功能改善七氟烷导致的POCD，为防治POCD提供理论依据和药物靶点。

目前普遍认为β-淀粉样蛋白（Aβ）与术后认知功能障碍（POCD）密切相关，但POCD仍无有效防治手段。纳米技术快速发展，但将纳米技术应用于防治POCD的研究较少。本项目以Aβ为切入点，研究了吸入麻醉药七氟烷、静脉麻醉药咪达唑仑、手术等围术期因素对体内外Aβ和小鼠认知功能的影响，并探究纳米氧化石墨烯（GO）、雷帕霉素等防治POCD的潜力和机制。研究发现：GO可通过抑制淀粉样前体蛋白（APP）的β裂解协同增强内体Aβ向溶酶体的传递，减少Aβ的生成并增强其降解，从而改善七氟烷麻醉和手术导致的海马Aβ积累与恐惧记忆受损；七氟烷暴露引发阿兹海默症（AD）模型鼠海马产生巨大自噬溶酶体，破坏细胞自噬降解功能，导致Aβ积累与空间学习记忆受损，雷帕霉素可以部分逆转上述病理变化，改善七氟烷损害的认知功能；转录因子EB（TFEB）向细胞核转运受抑制，可能参与咪达唑仑导致的溶酶体稳态失衡和Aβ积累。该项目通过基础研究，揭示了Aβ在POCD中的重要作用，提示巨大自噬溶酶体和TFEB可能是防治POCD的潜在靶点。此外，该项目创新性的提出纳米GO在改善术后动物认知功能方面具有独特作用，提示GO具有开发防治POCD纳米药物的潜力。