内质网应激介导微囊藻毒素诱导草鱼肝细胞凋亡的机制及GRP78的调控研究

项目名称： 内质网应激介导微囊藻毒素诱导草鱼肝细胞凋亡的机制及GRP78的调控研究

项目编号： No.31460146

项目类型： 地区科学基金项目

立项/批准年度： 2015

项目学科： 生物科学

项目作者： 隗黎丽

作者单位： 江西农业大学

项目金额： 58万元

中文摘要： 鄱阳湖是我国最大的淡水湖，日益面临着富营养化的威胁。水体富营养化可产生微囊藻毒素(MCs)，其高度的肝毒性已引起科学界广泛关注。诱导细胞凋亡是MCs重要毒性机制之一，但由内质网应激(ERS)介导的凋亡调控机制尚不清楚。我们前期研究初步表明MCs可致鱼类细胞凋亡并激活ERS相关基因TRAF2和CHOP等，我们推测两条ERS信号通路ASKl-JNK和CHOP可能参与了MCs诱导草鱼肝细胞凋亡。基于以上原因，本项目拟以鄱阳湖区草鱼作为研究对象，采用流式细胞术、免疫印迹等方法进一步研究ERS介导MCs诱导细胞凋亡的具体途径，揭示ERS介导凋亡的作用机制；同时采用过表达、RNA干扰和高通量测序技术研究ERS的信号分子GRP78对MCs诱导细胞凋亡的调控作用，揭示其调控机制。该项目的完成将诠释MCs肝毒性的新机制，为MCs的防治提供参考资料，亦为鄱阳湖水体污染的生物监测及渔业生产安全评估提供可靠依据。

中文关键词： 内质网应激；微囊藻毒素-LR；草鱼；细胞凋亡；GRP78

英文摘要： As the largest fresh water lake in china, Poyang Lake is being threatened by the progressive eutrophication of water bodies. The occurrence of cyanobacterial bloom can produce microcystins which is highly hepatotoxic.Induction of apoptosis is one of the important mechanisms of its toxicity, however, the apoptosis pathway of endoplasimic reticulum stress (ERS) is still unkown. On the base of our research, ERS was observed in hepatocyte from grass carp, and some other apoptosis-related genes,such as TRAF2, CHOP, were activated by MCs. Taking into consideration the mechanisms of apotosis caused by microcystins are different with exposure to the different concentration and time, so, we presumed the apoptotic pathway could be ASKl-JNK and CHOP. Glueose-regulated protein 78 is the key gene during the ERS, which could repress and activate the related genes to control the occurrence of apoptosis. In this study, grass carp from Poyang Lake and hepatocyte cultured in vitro will be exposed to microcystin-LR,one of the most hepatatoxin of microcystins, to build experimental model. The pathways and mechanisms of apoptosis mediated role of ERS will be studied by using flow cytometry, western blot and so on. Futhermore, in order to study the roles of regulating ERS-mediated apoptosis threatened by microcystin-LR and its signaling pathway, overexpression and short interfering RNA will be used to interfere the expression of GRP78. Meanwhile, high-throughput sequencing will be used to screen the downstream genes of GRP78 so as to understand its regulatory pathways deeply. This work can provide not only better understanding of the hepatotoxic mechanisms and prevention of microcystin-LR, but also more scientific basis for monitoring of MC-LR of water in Poyang Lake and security evaluation of fishery production.

英文关键词： Endoplasimic Reticulum Stress;Microcystin-LR;Grass carp;Apoptosis;GRP78