益髓灸对氧化应激脑衰老的胰岛素信号通路的调控

Brain aging is the primary stage of Alzheimer's disease (AD) and other neurodegenerative changes, oxidative stress is an important factor to cause aging. Enhanced antioxidant capacity of brain tissue can delay brain aging, cognitive dysfunction in AD and other brain diseases with the age increased. Oxidative stress products increased block insulin signaling pathway, Which in turn cognitive ability and even the occurrence of AD are closely linked, reducing insulin signaling pathway in the brain may be a key anti-aging. The applicant will base on the previous research of the Yisui moxibustion improve brain tissue antioxidant capacity. This project uses water maze testing for memory , brain tissue immunohistochemistry for the distributing of protein of IGF-1,InR,IRS-1,AKT,FOX3a,gsk-3,klotho,MnSOD,GADD45, and the tunel in situ end-labelling to detect apoptosis in cerebral cortex and Hippocampal, western blot to test the level of protein phosphorylation of AKT,FOX3a,real time PCR method to observe the mRNA express of IGF-1、InR、IRS-1、AKT、FOX3a、gsk-3,klotho,MnSOD,GADD45, elisa to assay serum and brain tissue in T-AOC ,8-OHdG, insulin, klotho protein content. Observed on the tonifying marrow moxibustion regulating INS/IGF-1/IRS-1/PI3K/AKT/FOXOs signaling pathway in brain aging mice induced by D-galactose , whether or not the anti-aging klotho gene through inhibition of insulin signaling pathway against oxidative stress, improve learning and memory. To clear moxibustion prescription regulatory target of brain insulin signaling system ,to provide the basis for the clinical comprehensive prevention brain aging, and to alleviate the brain degenerative diseases of aging due to the social and economic pressure will play a positive role.

脑衰老是阿尔茨海默病（AD）等神经退行性改变的最初级阶段，氧化应激是导致衰老的一个重要因素。增强脑组织的抗氧化能力可以延缓脑衰老、认知功能障碍、AD等脑神经疾病。氧化应激产物增加可能使胰岛素信号通路受阻。而后者又与认知能力乃至AD的发生有密切的联系，降低脑部的胰岛素信号通路可能是延缓衰老的关键所在。基于申请者此前的益髓灸提高脑组织抗氧化能力的研究，本项目采用水迷宫行为学检测、大脑组织免疫组化、TUNEL、WB、RT-PCR等方法，观察益髓灸对D-半乳糖小鼠脑部INS/IGF-1/IRS-1/PI3K/AKT/FOXOs信号通路的调节，以及抗衰老klotho基因是否通过抑制胰岛素信号通路来抵抗氧化应激，改善学习记忆。明确益髓灸处方对脑部胰岛素信号系统的调控作用靶点，为临床综合防治脑衰老提供依据,对缓解老龄化引起的脑神经退行性疾病给社会和经济带来的压力将起到积极作用。

氧化应激是导致脑衰老的一个重要因素。内源性氧化剂水平升高和长期暴露于外源性氧化应激可能是衰老组织中INS/IGF-1信号异常的基础，直接与认知能力乃至AD的发生有关。基于申请者此前的益髓灸提高脑组织抗氧化能力的研究，本项目采用水迷宫行为学检测、大脑组织免疫组化、TUNEL、WB、RT-PCR等方法，观察益髓灸对D-半乳糖小鼠大脑的保护作用。研究结果表明：①D-半乳糖小鼠模型的脑内胰岛素信号通路变化与外周的差异——外周血清胰岛素水平升高，脑内的胰岛素水平相对不足，从而导致D-半乳糖小鼠模型的脑内INS/IGF-1信号通路异常，IGF-1mRNA及蛋白水平升高，并伴随着抑制性位点p-IRS1蛋白的降低，最终激活下游通路AKT/FOXOs。②益髓灸通过抑制D-半乳糖小鼠体内8-羟基脱氧鸟苷（8-OHdG），提高血清、脑组织中Klotho、SOD2、GADD45mRNA及其蛋白的表达，提高总抗氧化能力（T-AOC），抑制神经元细胞的坏死，改善学习记忆能力，发挥延缓脑衰老的作用。③益髓灸可上调抗衰老Klotho蛋白，负反馈于脑内的胰岛素通路。 .益髓灸处方足三里（ST36）+悬钟(GB39)、百会(DU20)+关元(RN4)可以作为临床综合防治脑衰老的措施之一，对缓解老龄化引起的脑神经退行性疾病给社会和经济带来的压力将起到积极作用。