高白鲑鱼油通过抑制核因子κB、信号传导转录激活因子1和琥珀酸盐/缺氧诱导因子-1α信号通路的抗炎机制  

Anti-inflammatory Mechanism of Coregonus peled Oil via the Inhibition of Nuclear Factor Kappa-B(NF-κB),Signal Transducer and Activator of Transcription 1(STAT1),and Succinate/Hypoxia-Inducible Factor-1α(HIF-1α)Signaling Pathways

作　　者：刘宏旭 宋国库 王珍 夏效东 秦宁波 LIU Hongxu;SONG Guoku;WANG Zhen;XIA Xiaodong;QIN Ningbo(College of Food Science and Technology,Dalian Polytechnic University,Dalian 116034,China)

机构地区：[1]大连工业大学食品学院,辽宁大连116034

出　　处：《食品科学》2023年第17期67-78,共12页Food Science

基　　金：国家自然科学基金青年科学基金项目(32101963);辽宁省教育厅高校基本科研项目(面上项目)(LJKMZ20220890)。

摘　　要：目的:探讨高白鲑鱼油的化学成分和抗炎作用机制。方法:采用匀浆萃取超声法从高白鲑中提取鱼油,并对其主要化学成分进行检测。采用气相色谱-质谱联用仪对高白鲑鱼油进行脂肪酸成分分析。构建巨噬细胞RAW264.7炎症细胞模型,用脂多糖(lipopolysaccharide,LPS)刺激后,通过MTT法检测细胞毒性,采用液相色谱串联质谱联用仪对琥珀酸盐含量进行测定;采用试剂盒法测定上清液中一氧化氮(nitric oxide,NO)含量和一氧化氮合酶(inducible nitric oxide synthase,iNOS)活力;采用酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)测定细胞上清液中白细胞介素-6(interleukin-6,IL-6)、白细胞介素-1β(interleukin-1β,IL-1β)和肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)质量浓度。采用实时荧光定量聚合酶链式反应法测定细胞上清液中CD68、CD86、Arg1、CD206、YM1、HIF-1α基因的相对表达量,并通过蛋白免疫印迹法测定其蛋白的相对表达量。结果表明:高白鲑鱼油的主要化学成分为棕榈油酸、油酸、二十碳五烯酸和二十二碳六烯酸。其可以显著抑制NO、iNOS、IL-6、TNF-α、IL-1β的产生及相关基因的表达。鱼油可通过抑制LPS诱导RAW264.7细胞中的核因子-κB(nuclear factor kappa-B,NF-κB)抑制剂α的降解和减少p65核易位来抑制NF-κB的激活。此外,高白鲑鱼油可通过抑制信号传导转录激活因子1(signal transducer and activator of transcription 1,STAT1)的磷酸化,将巨噬细胞从M1型极化为M2型。另外,通过测定琥珀酸盐的含量、缺氧诱导因子-1α(hypoxia-inducible factor-1α,HIF-1α)基因和蛋白的表达量发现,高白鲑鱼油可以通过抑制琥珀酸盐的积累来抑制IL-1β的产生。结论:高白鲑鱼油可能通过抑制NF-κB、STAT1和琥珀酸盐/HIF-1α信号通路抑制炎症。Objective:To investigate the chemical composition and anti-inflammatory mechanism of Coregonus peled oil(CPO).Methods:CPO was extracted by ultrasonic homogenization and its major chemical components were detected.In addition,its fatty acid(FA)composition was analyzed by gas chromatography-mass spectrometry(GC-MS).Macrophage RAW264.7 cells were cultured as an inflammation model and stimulated with lipopolysaccharides(LPS).Cytotoxicity was evaluated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)assay.The accumulation of succinate was detected by liquid chromatography tandem mass spectrometry(LC-MS/MS).The concentration of nitric oxide(NO)and the activity of inducible nitric oxide synthase(iNOS)in the supernatant were measured by commercial kits.Enzyme-linked immunosorbent assay(ELISA)was used to determine the levels of interleukin-6(IL-6),interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α)in the supernatant.The expression levels of CD68,CD86,Arg1,CD206,YM1 and HIF-1αwere detected by quantitative real-time polymerase chain reaction(qPCR),and their protein expression levels were determined by Western blotting.Results:The major chemical constituents of CPO were palmitoleic acid,oleic acid,eicosapntemacnioc acid(EPA),and docosahexaenoic acid(DHA).CPO significantly inhibited the production of NO,iNOS,IL-6,TNF-α,and IL-1βas well as their gene expression levels.CPO blocked NF-κB activation by inhibiting the degradation of inhibitorαof NF-κB(IκBα)and decreasing p65 nuclear translocation in LPS-induced RAW264.7 cells.In addition,CPO promoted macrophage polarization from M1 to M2 types by inhibiting the phosphorylation of signal transducer and activator of transcription 1(STAT1).The content of succinate and the expression levels of hypoxia-inducible factor-1α(HIF-1α)gene and protein indicated that CPO could inhibit the production of IL-1βby inhibiting the accumulation of succinate.Conclusion:The anti-inflammatory mechanism of CPO may be due to the inhibition of the NF-κB,STAT1

关 键 词：高白鲑 鱼油 炎症 极化 琥珀酸盐/缺氧诱导因子-1α 

分 类 号：TS201.4[轻工技术与工程—食品科学]