长茎葡萄蕨藻对斑马鱼氧化应激肝损伤的保护及基于PI3K/Akt信号通路的作用机制研究

According to recent researchs，Caulerpa lentillifera has the effect of improving glycolipid metabolism by acting on animal insulin PI3K/Akt signaling pathway, and also has the activity of alleviating oxidative stress and improving liver function. The oxidative liver injury in fish is becoming more and more serious with the increasing stress factors in fish culture. Bases on the fish physiological characteristics that persistent blood glucose elevation after stress, this study draws on the results of mammalian metabolic diseases research to discuss the pathogenesis and drug resistance mechanisms. In this project, zebrafish, a fish model organism, was used as the research object to establish a model of liver injury induced by oxidative stress with diquat stimulation. The protective effects of dietary Caulerpa lentillifera on oxidized liver injury of zebrafish under stress will be further identified. Based on the applicable effect, the mechanism of Caulerpa lentillifera on regulating glycolipid metabolism, alleviating oxidative stress and protecting liver injury through PI3K/Akt signaling pathway in zebrafish will be studied to elucidate the mechanism on protecting liver from phenotype, gene, protein, metabolism and omics levels. The results of this project will explore new scientific ideas for the research of anti-oxidative stress and the development of regulatory drugs in fish, and provide a theoretical basis for reducing production losses caused by adverse factors.

已有研究报道发现长茎葡萄蕨藻提取物通过作用于动物胰岛素信号通路PI3K/Akt来调节糖脂代谢，还具有缓解氧化应激和改善肝脏功能的活性。针对鱼类养殖中越来越多的应激因素使氧化型肝损伤日益严重的情况，本研究借鉴哺乳动物代谢性疾病的研究结果，依据鱼类受应激后血糖持续性升高的生理特点，拟以鱼类模式生物—斑马鱼为研究对象，通过敌草快（Diquat）刺激诱导建立氧化应激肝损伤模型；对被诱导过的斑马鱼投喂含长茎葡萄蕨藻的饲料，明确长茎葡萄蕨藻对斑马鱼氧化应激型肝损伤的保护作用；在此基础上，开展长茎葡萄蕨藻调控实验斑马鱼肝脏PI3K/Akt信号通路调节糖脂代谢、缓解氧化应激和防治肝脏损伤的机理研究，从表型、基因、蛋白、代谢以及组学的水平多层次全面综合阐释其保护肝脏的作用机制。本项目研究结果将为鱼类抗氧化应激研究及其调控药物的研发提供新的科学思路，为预防不良因子对养殖生产造成的损失提供理论依据。

鱼类在集约化养殖中遭受越来越多的应激因素使氧化型肝损伤问题日益严重，但是针对性的防治手段仍非常有限。本研究以模式生物—斑马鱼为研究对象，采用敌草快（Diquat）刺激诱导建立氧化应激肝损伤模型，考察长茎葡萄蕨藻对鱼类氧化型肝损伤的防治作用，开展了表型、基因、蛋白、代谢以及组学水平的研究。结果表明，补充长茎葡萄蕨藻可以有效降低敌草快造成的肝脏氧化应激，减轻肝病理组织损伤，改善氧化应激导致的血糖升高、脂质沉积现象，并且减轻对斑马鱼生长和非特异性免疫的抑制。肝脏代谢组学分析显示经长茎葡萄蕨藻治疗使肝脏的代谢轮廓接近正常对照组，转录组分析表明长茎葡萄蕨藻通过干预包括PI3K/Akt在内的多种信号通路恢复敌草快诱导的基因转录改变，补充长茎葡萄蕨藻显著逆转了氧化损伤对肝脏能量代谢、脂肪代谢、类固醇及脂肪酸合成的影响。定量RT-PCR与蛋白印迹技术显示敌草快诱导的氧化损伤影响了肝脏PI3K/Akt通路信号传导，补充长茎葡萄蕨藻调控下游mTOR分子活化Nrf2上调HO-1表达，并抑制NF-κB下调Caspase-3表达，另外，AMPK和MAPK途径可能共同参与了对mTOR的调控。长茎葡萄蕨藻可以通过斑马鱼肝脏PI3K/Akt/mTOR通路增强肝脏抗氧化能力、促进能量代谢、抑制细胞凋亡、减少脂毒性和炎症反应，发挥对肝脏氧化损伤的保护作用。本项目的研究结果为进一步深入探索氧化应激状态下鱼类肝损伤机制和新型药物研发提供了理论基础。