黄颡鱼甘露糖受体基因的克隆和功能分析

发布时间：2019-04-02 18:08

【摘要】：甘露糖受体(MR)隶属凝集素超家族,主要表达于巨噬细胞和未成熟的树突状细胞表面。MR不仅在先天免疫防御中发挥重要作用,还通过参与抗原呈递,激活T淋巴细胞,启动获得性免疫应答过程。本研究采用同源克隆技术获得黄颡鱼甘露糖受体(pf MR)基因,采用荧光定量PCR技术检测MR在正常黄颡鱼体内的分布情况,采用湒爱德华菌感染黄颡鱼头肾巨噬细胞和甘露聚糖封闭MR方法研究黄颡鱼MR在抗细菌感染中的作用。结果显示,pf MR同团头鲂、草鱼、斑马鱼和尼罗罗非鱼的MR聚为一支。pf MR在所检测的12个组织中均有分布,其在肾脏、脾脏和肌肉组织中表达量较高,在血液中表达量较少。湒爱德华菌感染黄颡鱼头肾巨噬细胞后,pf MR、IL-1β和TNF-a均被细菌诱导表达,超氧阴离子和一氧化氮的含量也上升,超氧阴离子在感染30 min后即显著上升,一氧化氮在感染12 h后才显著上升。甘露聚糖竞争结合MR,显著抑制巨噬细胞内化GFP标签的湒爱德华菌,加入EDTA减少内化的荧光强度,加入Ca~(2+)使内化的荧光强度回升。研究表明,黄颡鱼头肾巨噬细胞MR参与湒爱德华菌的识别和内吞过程,而且依赖Ca~(2+)。
[Abstract]:Mannose receptor (MR) belongs to the lectin superfamily and is mainly expressed on macrophages and immature dendritic cells. Mr not only plays an important role in innate immune defense, but also activates T lymphocytes by participating in antigen presentation. Initiate the process of acquired immune response. In this study, the mannose receptor (pf MR) gene of Pelteobagrus fulvidraco was obtained by homologous cloning technique, and the distribution of MR in normal yellow catfish was detected by fluorescence quantitative PCR. The role of MR in anti-bacterial infection of Pelteobagrus fulvidraco MR was studied by using Edwardian bacteria infection and mannan blocking MR method in the head kidney macrophages of Pelteobagrus fulvidagrus. The results showed that the MR of, pf MR was the same as that of snout bream, grass carp, zebrafish and Nile tilapia. The expression of pf MR was higher in kidney, spleen and muscle than in blood. The expression of pf MR,IL-1 尾 and TNF-a were induced by bacteria, the contents of superoxide anion and nitric oxide increased, and the superoxide anion increased significantly at 30 min after infection with Evodomonas fulvidracobagrus, the expression of SOD 尾 and TNF-a were induced by bacteria, and the content of superoxide anion and nitric oxide increased significantly after 30 min infection. No increased significantly after 12 hours of infection. The competition of mannan combined with MR, significantly inhibited the internalization of GFP-labeled Eduardo spp. by adding EDTA, decreased the fluorescence intensity of internalization, and increased the fluorescence intensity of internalization by adding Ca~ (2). The results showed that MR was involved in the identification and endocytosis of Edwards and depended on Ca~ (2).
【作者单位】：华中农业大学水产学院水生动物医学系;华中农业大学水产学院淡水水产健康养殖湖北省协同创新中心;华中农业大学水产学院池塘健康养殖湖北省工程实验室;华中农业大学水产学院农业部淡水生物繁育重点实验室;
【基金】：国家自然科学基金(31372563,31572657,31502199) 中央高校基本科研业务费专项(2014PY035,2662015QC019) 湖北省科技支撑计划(2015BBA228,YSF2015000255) 农业部渔用药物创制重点实验室开放课题(201401) 广东省海洋与渔业局基金(A201512C003,2015-115) 武汉市科技局基金(2016020101010089)~~
【分类号】：S943

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