细胞珠蛋白过表达对人肝细胞株LO-2脂质代谢的影响初探  

Preliminary study on the effect of cytoglobin overexpression on lipid metabolism of human liver cell line LO-2

作　　者：刘诗俐 王哲彦 李珍 王萍 董文其 LIU Shili;WANG Zheyan;LI Zhen;WANG Ping;DONG Wenqi(Department of Laboratory Medicine and Biotechnology, Southern Medical University, Guangzhou 510080, China;Molecular Imaging Center, the Fifth Affiliated Hospital of SUN YAT-SEN University, Zhuhai 519000, China)

机构地区：[1]南方医科大学检验与生物技术学院,广东广州5100802 [2]中山大学附属第五医院分子影像中心,广东珠海519000

出　　处：《暨南大学学报（自然科学与医学版）》2021年第3期242-250,共9页Journal of Jinan University(Natural Science & Medicine Edition)

基　　金：广东省科技计划项目(2013A022100027)。

摘　　要：目的:通过建立体外肝细胞株LO-2脂肪变性模型,初步探究细胞珠蛋白对非酒精性脂肪肝中脂质沉积、炎性因子表达及NF-κB通路的影响及相关分子机制.方法:通过含有浓度200μmol/L油酸、100μmol/L棕榈酸的高脂培养基诱导人正常肝细胞株LO-2脂肪沉积,以LO-2-GFP-CYGB为实验组,LO-2-GFP为对照组探究CYGB对LO-2脂质沉积的影响,通过油红O染色检测脂质沉积情况,GPO-PAP法检测细胞内甘油三酯水平,COD-PAP法检测细胞内胆固醇水平,通过RT-qPCR检测脂质生成相关基因ACACA、ACACB、FASN、ACSS1、ACSS2、ACSS3的mRNA表达水平,蛋白质免疫印迹法检测细胞炎性因子IL-1β,IL-6,TNF-α的表达水平,以及核转录因子-κB(NF-κB p65),磷酸化P65(NF-κB PP65)表达水平,用含浓度60,90μmol/L NF-κB抑制剂PDTC与高脂成分的培养基处理LO-2细胞48 h后检测细胞内甘油三酯和胆固醇水平.结果:LO-2经过高脂处理48 h后,两组细胞甘油三酯及胆固醇水平均上升,而实验组低于对照组(P<0.05),90μmol/L NF-κB抑制剂PDTC处理后,甘油三酯和总胆固醇水平与未加入PDTC相比表现下调(P<0.05);炎症相关因子IL-1β,IL-6和TNF-α在实验组中表达均低于对照组(P<0.05);与对照组相比,实验组经高脂处理24 h后NF-κB信号通路受到抑制(P<0.05).结论:CYGB下调LO-2细胞甘油三酯和总胆固醇水平,抑制炎性因子表达,抑制NF-κB信号通路,缓解LO-2脂质沉积.Objective:By establishing a steatosis model of human liver cell line LO-2 in vitro,we preliminarily explored the effects of cytoglobin(CYGB)on lipid deposition,inflammatory factor expression and NF-κB pathway in non-alcoholic fatty liver disease and their molecular mechanisms.Methods:A high fat medium containing 200μmol/L oleic acid and 100μmol/L palmitic acid was used to induce fat deposition in LO-2 cells.Take LO-2-GFP-CYGB as the experimental group and LO-2-GFP as the control group to investigate the effect of CYGB on lipid deposition in LO-2 cells.Oil red O staining was used to detect lipid deposition,GPO-PAP method to detect intracellular triglyceride content,COD-PAP method to detect intracellular cholesterol content,and RT-qPCR to detect the mRNA expression levels of lipid production-related genes ACACA,ACACB,FASN,ACSS1,ACSS2,ACSS3.Western blotting assay was used to detect the expression levels of inflammatory factors IL-1β,IL-6,TNF-α,NF-κB p65 and phosphorylated P65(P-NF-κB p65).The LO-2 cells were cultured with high fat medium containing 60 and 90μmol/L NF-κB inhibitor PDTC,respectively,for 48 h,and then the intracellular triglyceride and total cholesterol levels were detected.Results:After LO-2 cells were treated with high fat medium for 48 hours,the triglyceride and total cholesterol levels of both two groups increased,but those of experimental group was lower than those of control group(P<0.05).After treatment with 90μmol/L NF-κB inhibitor PDTC,the levels of triglyceride and total cholesterol decreased compared with those without PDTC(P<0.05).The expression of inflammation-related factors IL-1β,IL-6 and TNF-αwas lower in experimental group than in control group(P<0.05).Compared with control group,the NF-κB signaling pathway was inhibited in the experimental group after 24 hours of high fat treatment(P<0.05).Conclusion:CYGB down-regulates the content of triglyceride and total cholesterol in LO-2 cells,inhibits the expression of inflammatory factors and NF-κB signaling pathway,and relieves

关 键 词：细胞珠蛋白 非酒精性脂肪肝 LO-2 NF-ΚB 

分 类 号：Q291[生物学—细胞生物学]