褪黑素通过Nrf2通路对糖尿病大鼠颈动脉损伤的围术期保护研究

Diabetic vascular diseases derive from oxidative damage, especially during the perioperative period, for that hyperglycemia can induce a large number of reactive oxygen species (ROS). Previous studies show that ROS inhibits endothelial eNOS function and transforms the normal contractile phenotype of smooth muscle into pathological synthesis phenotype. As the protector in oxidative stress, varieties of antioxidant enzymes are the key factor of scavenging ROS, and their gene transcriptions are promoted by nuclear factor erythroid 2-related factor 2 (Nrf2). Being activated by phosphorylation signaling pathway, the activated p-Nrf2 translocates from cytoplasm to nucleus, and then up-regulates the transcription of antioxidant enzyme genes. Therefore, activating Nrf2 may effectively protect vascular vessels from diabetes-related oxidative damage during the perioperative period. Known as the biological rhythm factor, melatonin also participates in the process of vascular healing and anti-inflammation through activating the phosphorylation signaling pathway. Besides, our study group has found that melatonin can promote the functional expression of Nrf2 in blood vessels. Based on the above statements, the project intends to study the beneficial effects of melatonin on the promotion of Nrf2 signaling/ mechanism relationship between melatonin and Nrf2 activation through Nrf2 overexpressed/knockdown cell lines, Nrf2(-/-) rat diabetes and carotid artery balloon surgery model. Moreover, the antioxidant efficiency of melatonin is further demonstrated through the binding ability of Nrf2 to antioxidant response element (ARE) and the increase in the expression and activity of antioxidant enzyme enzymes. Following our previous national natural science foundation research, this study is the first to demonstrate that melatonin may serve as a therapeutic strategy to ameliorate the vascular ROS-injury in diabetic rats received carotid artery balloon surgery through Nrf2 signaling pathway.

糖尿病是重大公共卫生问题，其核心是血管病变，其关键是糖尿病血管围术期不良事件。活性氧簇(ROS)的氧化损伤是糖尿病血管围术期风险的始动点。机体清除ROS的因子是抗氧化酶。核因子NF-E2相关因子(Nrf2)是抗氧化酶表达的启动因子，Nrf2信号是重要的抗氧化损伤通路。因此激活Nrf2信号通路可有效降低糖尿病血管围术期风险。褪黑素是调节生物节律的关键因子，申请人前期研究发现其与MT1/MT2受体结合后，参与血管愈合过程，并与Nrf2信号通路有关。本课题拟通过Nrf2过表/降低细胞系、Nrf2敲降大鼠和颈动脉球囊手术模型，研究Nrf2信号通路的激活机制；并检测Nrf2与抗氧化酶基因顺式作用元件ARE的结合，探讨褪黑素对抗氧化酶表达的促进作用。本课题在前期基础上深入分析Nrf2信号通路在糖尿病血管围术期氧化损伤中的保护作用，为糖尿病血管围术期安全以及术后快速愈合提供新的研究靶点和治疗策略。

随着社会经济发展、寿命和老年人口的增加，需要手术的患者数量增加，并且许多人一生中至少要做一次手术。如何保证围术期安全是重要的关注点，在手术监护中，除了心率、血压、呼吸、氧合之外，体温是另外一个重要的生命体征。因此，围术期体温监测已经成为国内外共识。此外，在围术期患者安全的议题中，糖尿病提高了手术风险。其中，糖尿病血管病变是造成围术期高风险的重要因素，其可造成心脑等重要组织器官供血不足和功能丧失，极大地影响糖尿病患者围术期安全和术后恢复。. 围术期的手术创伤和应激，可归结于炎症和氧化损伤。本研究以褪黑素的抗炎、抗氧化作用为切入点，分析其在围术期中的血管保护作用，以及激活Nrf2信号通路的抗氧化损伤机制。并且，监测围术期体温对于血管功能的影响，尤其是脑血管的功能变化，进而揭示术中体温对于术后快速康复和认知功能恢复的影响。. 研究表明，褪黑素可显著降低血管壁的氧化损伤。通过免疫荧光分析，应用褪黑素的糖尿病大鼠血管壁的过氧化物ROS明显降低；血管壁功能分析表明，动脉粥样硬化标记分子骨桥蛋白osteopontin表达明显降低和血管壁弹力纤维降解率减少。通过进一步分析围术期的体温监测研究，围术期保温可促进大鼠术后认知功能的快速康复，表现为Morris水迷宫和Y迷宫实验的认知记忆得分均显著提高。通过脑血管的功能分析，研究发现血管内皮保护因子eNOS和VEGF表达明细增加，并且血管平滑肌细胞合成表型标记物RBP的表达减低，而收缩表型标记物SM-MHC的表达增加，这表明血管平滑肌仍表现为正常的收缩表型。. 综上所述，褪黑素发挥抗氧化损伤的作用，在糖尿病血管围术期保护方面具有治疗前景。体温作为术中重要的生理指标，围术期体温保护可维持术中正常的脑血管功能，并且促进术后认知功能的快速康复。通过围术期安全和术后恢复的研究，可显著降低住院日、减轻经济负担，具有重要的临床及社会价值；并为围术期患者快速康复提供新的研究靶点和防治策略。