AMPK-Nrf2信号通路对炎症负性调控的分子机理

Inflammation is essential pathologic fundamentals of many diseases,but its mechanisms underlying are still unclear.It has proposed recently that the disorder of energy metabolism promots inflammation and AMP-dependent protein kinese(AMPK), a sensor enzyme for energy metaolism,negatively regulates inflammation. These understandings suggest the existence of underlying signal pathway(s) related to the role of AMPK in inflammation control. In own experiments in adavance, we found that Nrf2-mediated anti-oxidative reactions are indispensable for the inflammation supression of AMPK.Our findings show that in LPS-activated macrophages, a classic TLR receptor-mediated inflammatory cell model, metformine suppressed the LPS-elevated expression of inflammatory factors (TNFa, COX2,IL1b) as well as NO and ROS production. On the other hand, metfomine addition promoted the expression of Nrf2-down stream genes (NQO1, HO1) and Nrf2 activation. These results are novel and suggest an underlying signal transduction pathway: AMPK-Nrf2 pathway. To explore the molecular mechanisms about inflammation and to develop our understanding for AMPK-mediated inflammation control, we plan present proposal with the focus on AMPK-Nrf2 singaling network. In this proposal, we will firstly work to clarify the relationship between AMPK and Nrf2 in vivo by using LPS-induced infliammation model and Nrf2 knock out mice, then go advance to identify the AMPK binding proteins from inflammation suppressed, AMPK-activated macrophages via co-immunoprecipitation method followed by proteomics technology, such as MS analysis, bioinfomatics analysis,and other molecular biology methods. Next,we will progress to determine the crue ptotein（s）which functionally connect AMPK singnal with Nrf2 pathway during inflammation modulation by a series of fundctional assays. Finally, the crosslink between AMPK-Nrf2 pathway and the routes which pass the signals from energy control to inflammation supression will be addressed carefully, by the molecular and cellular methods including in vitro phosphorylation assay and gene modification. . Differently from previous studies which investigates how is inflammation promoted, this proposed study directly addresses how can inflammation be suppressed. Given AMPK-Nrf2 pathway connecting energy homoestasis and oxidative stress control together, we believe here the results from this project should be informative for our further understanding about energy homeostasis and inflamation suppression, also be helpful for parmacologic study and clinic application of AMPK agonists. .All of the critial materials and technic platforms necessary for this project have been set, and the members joining this project have enough ability and time to take their duty.

炎症是许多疾病的重要病理基础，迄今仍缺乏有效的控制炎症的方法和措施。近年有人提出能量代谢紊乱促进炎症的进程，而AMPK作为细胞能量代谢调节的关键酶对炎症起着负性调控作用，这提示了AMPK相关的炎症负性调控通路的存在。我们在前期研究中发现Nrf2活化的抗氧化通路对AMPK的炎症抑制作用极为重要。为进一步探讨AMPK的抗炎作用分子机制，解析仍未知晓的AMPK-Nrf2信号通路，本项目拟在前期工作基础上，利用脂多糖致炎细胞、动物模型以及Nrf2基因缺失小鼠，首先确认AMPK-Nrf2通路对体内炎症负性调控的影响，然后通过免疫共沉淀/蛋白质组学/生物信息学分析等手段，筛查影响炎症负性调节信息传递的AMPK结合蛋白，解析AMPK-Nrf2通路的关键分子节点并获悉该通路与其它通路的联系。本研究可加深我们对能量代谢与炎性反应交汇整合的认识,有助于解明炎症的负性调控机理，为新型抗炎药物的研发和应用奠定基础

代谢改变如何影响炎症发生是细胞生物学的重要和前沿问题。本项目立足探讨细胞代谢调控关键酶AMPK抑制炎症的机理，追求创新性和科学性。根据计划的研究目标和路线，我们用体内外实验方法，包括敲基因小鼠和细胞水平的遗传及化学干预，证明了AMPK活化Nrf2介导的抗氧化通路是重要的抗炎机制， 揭示了AMPK激活剂对AMPK-Nrf2信号通路的激活在控制炎症方面的有效性和应用前景，还证明了AMPK激活剂对氧化应激所致细胞衰老的缓解作用与其改善细胞自噬功能和减少炎性因子产生有关 。为进一步解析机理，应用免疫共沉淀和蛋白组学技术， 我们发现了数种未曾报道的AMPK结合蛋白，认识到AMPK对经典炎性反应通路蛋白活性的调控可能是AMPK激活Nrf2抗氧化通路的关键事件之一 。. 部分研究结果已经发表在多篇中科院分区的1区和2区SCI期刊上，并曾在国内外会议论坛演讲，为阐明AMPK抑制炎症的机理和明确代谢与炎症之间的联系提供了新的证据和视点，有明确的科学意义。本项目支持培养了共计10名博士和硕士研究生，已毕业2名博士和5硕士研究生。