广州管圆线虫感染大鼠脑组织转录组分析  

Transcriptome analysis of brain tissue in rat infected with Angiostrongylus cantonensis

作　　者：程东慧 蒋天哥 景一丹 杨丽敏 郭云海 方圆 李中秋 张仪 CHENG Donghui;JIANG Tiange;JING Yidan;YANG Limin;GUO Yunhai;FANG Yuan;LI Zhongqiu;ZHANG Yi(National Institute of Parasitic Diseases,Chinese Center for Disease Control and Prevention,Chinese Center for Tropical Diseases Research,National Key Laboratory of Intelligent Tracking and Forecasting for Infectious Diseases,Key Laboratory on Parasite and Vector Biology,Ministry of Health,WHO Collaborating Centre for Tropical Diseases,National Center for International Research on Tropical Diseases,Ministry of Science and Technology,Shanghai 200025,China;School of Global Health,National Center for Tropical Disease Research,Shanghai Jiao Tong University,Shanghai 200025,China)

机构地区：[1]中国疾病预防控制中心寄生虫病预防控制所(国家热带病研究中心),传染病溯源预警与智能决策全国重点实验室,国家卫生健康委员会寄生虫病原与媒介生物学重点实验室,世界卫生组织热带病合作中心,科技部国家级热带病国际研究中心,上海200025 [2]上海交通大学医学院-国家热带病研究中心全球健康学院,上海200025

出　　处：《中国寄生虫学与寄生虫病杂志》2024年第2期217-224,233,共9页Chinese Journal of Parasitology and Parasitic Diseases

基　　金：国家重点研发计划(2021YFC2300800,2021YFC2300802)。

摘　　要：目的了解广州管圆线虫感染后大鼠脑组织转录组的表达水平。方法将32只SD大鼠随机分为对照组(12只)和感染组(20只),感染组每只大鼠经灌胃感染40条广州管圆线虫Ⅲ期幼虫,对照组灌胃等体积生理盐水。感染后1、7、14、21 d,对照组、感染组分别随机解剖3、5只,取脑组织制备石蜡切片,苏木精-伊红(HE)染色后观察脑组织病理变化。TRIzol法提取感染后14 d大鼠脑组织RNA,逆转录为cDNA后测序。选取差异表达mRNA进行基因本体论(GO)富集分析和京都基因与基因组百科全书(KEGG)代谢通路分析,采用STRING数据库预测差异表达mRNA靶蛋白之间的蛋白质互作(PPI)关系。利用生物信息学方法构建差异表达基因的竞争性内源性RNA(ceRNA)调控网络。采用qPCR验证表达差异的lncRNA。结果HE染色结果显示,感染广州管圆线虫后14 d,感染组大鼠脑组织出现病理变化,脑海马区神经元胞浆固缩;感染后21 d,脑膜处可见寄生虫样组织。RNA测序结果显示,感染广州管圆线虫后14 d,大鼠脑组织中差异表达mRNA的数量为955个(890个上调、65个下调);差异表达lncRNA的数量为193个(122个上调、71个下调)。GO富集分析结果显示,差异表达mRNA主要富集在炎症反应、免疫应答等生物过程,细胞成分主要为质膜外侧的胞外空间、细胞表面等,分子功能主要为趋化因子活性、趋化因子受体结合等;KEGG代谢通路分析结果显示,差异表达mRNA主要参与细胞因子-细胞因子受体相互作用、趋化因子等信号通路。PPI分析结果显示,差异表达基因主要的靶点为趋化因子配体11、RT1-Da、丝氨酸家族E成员1等,均与免疫应答相关。ceRNA结果显示,显著富集的miRNA如mir-466b-3p、mir-1956、mir-207和mir-328a-5p等与免疫应答、细胞凋亡、血管生成等过程有关。qPCR结果显示,感染广州管圆线虫后,感染组大鼠H19的相对转录水平逐渐升高,在感染后21 d达到峰值,为15.07Objective To understand the expression levels in brain tissue transcriptome of rat infected with Angiostrongylus cantonensis.Methods Thirty-two SD rats were randomly divided into control group(12 rats)and infected group(20 rats).The rats in the infected group were infected with 40 A.cantonensis stageⅢlarvae by gavage and in the control group with the same volume of saline.8 rats(3 of the control group and 5 of the infected group)were randomly dissected to collect brain tissues,of which paraffin sections were prepared and stained with hematoxylineosin(HE)to observe the pathological changes 1,7,14 and 21 days after infection.Brain tissue RNA was extracted 14 days after infection for detection of differentially expressed mRNA and lncRNA by using RNA sequencing technique.Gene ontology(GO)enrichment analysis and Kyoto encyclopedia of genes and genomes(KEGG)metabolic pathway analysis of differentially expressed mRNA were performed.The STRING database was used to predict protein-protein interaction(PPI)between differentially expressed mRNA target proteins.Bioinformatics was utilized to construct the competitive endogenous RNA(ceRNA)regulatory network of differentially expressed genes.The differential expression of lncRNA was verified by qPCR.Results HE staining showed that pathological changes appeared in the rat brains of the infected group 14 days after infection with A.cantonensis,with cytoplasmic consolidation in the hippocampal neurons and parasite-like tissues could be seen at the meninges 21 days after infection.RNA sequencing result showed that the number of differentially expressed mRNAs in the rat brains was 955(890 up-regulated and 65 down-regulated)14 days after infection;the number of differentially expressed lncRNA was 193(122 up-regulated and 71 down-regulated).GO enrichment analysis showed that differentially expressed mRNAs were mainly enriched in biological processes such as inflammatory response and immune response,the cellular components were mainly the extracellular space outside of the plasma mem

关 键 词：广州管圆线虫 长链非编码RNA 转录组学 竞争性内源RNA 

分 类 号：R532.19[医药卫生—内科学]