从内质网应激的角度探究微囊藻毒素-LR对斑马鱼离体肝细胞脂代谢的影响  

Effects of Microcystin-LR on Lipid Metabolism in Zebrafish Liver Cells via Endoplasmic Reticulum Stress Pathway

作　　者：张丹丹 杨慧 欧阳康 况宇 汤蓉[1,2,3,4] 李大鹏 李莉[1,2,3,4] Zhang Dandan;Yang Hui;Ouyang Kang;Kuang Yu;Tang Rong;Li Dapeng;Li Li(College of Fisheries,Huazhong Agricultural University,Wuhan 430070,China;Engineering Research Center of Green Development for Conventional Aquatic Biological Industry in the Yangtze River Economic Belt,Ministry of Education,Wuhan 430070,China;Hubei Provincial Engineering Laboratory for Pond Aquaculture,Wuhan 430070,China;Freshwater Aquaculture Collaborative Innovation Center of Hubei Province,Wuhan 430070,China)

机构地区：[1]华中农业大学水产学院,武汉430070 [2]长江经济带大宗水生生物产业绿色发展教育部工程研究中心,武汉430070 [3]池塘健康养殖湖北省工程实验室,武汉430070 [4]淡水水产健康养殖湖北省协同创新中心,武汉430070

出　　处：《生态毒理学报》2023年第2期410-419,共10页Asian Journal of Ecotoxicology

基　　金：国家自然科学基金资助项目(32071621);国家重点研发计划课题(2019YFD0900303);农业部现代农业产业技术体系(CARS-45-24)。

摘　　要：为了探究微囊藻毒素-LR(MC-LR)对斑马鱼离体肝细胞内质网应激(ERs)和脂质代谢的影响及机制,本实验以斑马鱼肝细胞系(ZFL)为实验材料,使用不同浓度梯度MC-LR(0、10、20、40、80和160μg·mL^(-1))分别进行24 h暴露,在明确细胞活力和半致死浓度(49.3μg·mL^(-1))的基础上,选定10μg·mL^(-1)为暴露浓度,研究细胞中总胆固醇(TC)和甘油三酯(TG)的含量及ERs信号分子、下游因子以及与脂质代谢相关的基因表达情况,并利用ERs抑制剂牛磺熊去氧胆酸(TUDCA)进行机制验证。结果表明,和对照组相比,MC-LR(10μg·mL^(-1))暴露诱导TC、TG含量显著上升,未折叠蛋白反应(UPR)途径相关基因(包括atf6、eif2ak3、ern1和xbp1s)以及下游脂质代谢相关基因(srebf1、fasn、acaca、scd、srebf2、hmgcra和hmgcs1)的mRNA表达显著上调;而TUDCA处理导致TC、TG含量显著下降,且UPR和脂类合成途径相关基因表达水平显著性下调。相对地,在TUDCA预处理组中,TC、TG含量、UPR和脂类合成途径相关基因表达相对于MC-LR处理组显著下降,但和对照组相比无显著差异。上述结果表明,MC-LR可通过影响肝脏脂质合成相关基因表达对体外ZFL细胞脂质代谢产生影响,其机制是MC-LR会诱导ERs和固醇调控元件结合蛋白(SREBP)活化(srebf1和srebf2),进而驱动下游脂质和胆固醇代谢合成基因(fasn、acaca、scd、hmgcs1和hmgcra)的上调,最终导致肝脏脂质的蓄积。TUDCA预暴露组相应检测指标的恢复进一步验证了ERs在MC-LR引起的斑马鱼肝脏脂质代谢异常中的作用。本研究的发现为MC-LR肝毒性提供了机制上的见解,并由此可外推到MC对人健康的潜在影响。To investigate the effects of MC-LR on endoplasmic reticulum stress(ERs)and lipid metabolism in vitro liver cells and its mechanism,we selected ZFL(zebrafish liver)cells as experimental model and expose them to MC-LR(0,10,20,40,80,160μg·mL^(-1))for 24 h.After determining the 24 h-LC 50 for MC-LR(49.3μg·mL^(-1)),the 10μg·mL^(-1) of MC-LR was set as exposure concentration and tauroursodeoxycholic acid(TUDCA)was used as an inhibitor to verify the role of ERs in lipid metabolism.The results showed that the contents of intracellular total cholesterol(TC)and total triglycerides(TG)were increased significantly in the MC-LR(10μg·mL^(-1))treatment group compared to the control group,and transcriptional levels of unfolded protein response(UPR)pathways related genes including atf6,eif2ak3,ern1,and xbp1s as well as downstream lipid metabolism-related genes including srebf1,fasn,acaca,scd,srebf2,hmgcra and hmgcs1 were significantly up-regulated.In contrast,the contents of TC and TG were decreased significantly in the TUDCA treatment group compared to the control group,and the expression levels of genes related to UPR pathways and lipid synthesis were significantly downregulated.In the TUDCA pretreatment group,the contents of TC and TG and expression levels of genes involved with UPR pathways and lipid synthesis returned to the control levels,but significantly decreased compared with the MC-LR treatment group.This finding indicates that MC-LR mainly affected lipid metabolism of in vitro ZFL cells by interfering with the expression of lipid synthesis-related genes(fasn,acaca,scd,hmgcs1 and hmgcra).The mechanism was that the ERs caused by MC-LR further upregulated the expression of the steroid regulatory element protein factor srebf1 and srebf2 as well as genes related to lipid synthesis.The exposure of TUDCA and the recovery of corresponding test indicators further verified the role of ERs in the abnormal lipid metabolism of zebrafish liver caused by MC-LR.In conclusion,the results of this study provided a mechanistic i

关 键 词：微囊藻毒素 ZFL细胞系 内质网应激 脂质代谢 牛磺熊去氧胆酸 

分 类 号：X171.5[环境科学与工程—环境科学]