目的 基于生物信息学分析细粒棘球蚴致敏反应机制和分子靶标。 方法 将9只BALB/c小鼠随机分为3组,每组3只,分别为感染未致敏组、感染致敏组、健康对照组。感染未致敏组小鼠经腹腔注射原头节悬液（0.2 ml/鼠）,90 d后腹腔注射生理盐水（0.2 ml/鼠）;感染致敏组小鼠经腹腔注射原头节悬液（0.2 ml/鼠）,90 d后腹腔注射粗制囊液致敏（0.2 ml/鼠）,健康对照组小鼠注射等量生理盐水。注射后1 h,取各组小鼠的脾组织提取RNA进行转录组测序,通过生物信息学分析找到差异表达基因,使用edgeR（以RNA-Seq by expectation-Maximization RSEM进行定量）软件进行表达差异显著性分析,用String数据库和Cytoscape软件对差异基因作网络互作分析,并将得分设置为> 500;使用cytoHubba获取节点的度（degree）筛选degree大于3的节点作展示;使用DAVID在线富集分析工具进行分析,分析结果使用R语言ggplot2进行可视化。 结果 感染致敏组与健康对照组有743个基因差异表达,其中568个上调表达基因和175个下调表达基因。感染未致敏组与健康对照组中有554个基因差异表达,其中393个上调表达基因和161个下调表达基因。感染致敏组与健康对照组有458个特有表达基因,感染未致敏组与健康对照组有269个特有表达基因,三组有285个共有表达基因。感染致敏组与健康对照组特有的458个基因GO富集结果显示,147个参与生物学进程,10个参与细胞成分,43个参与分子功能。共有的基因GO富集结果显示,56个参与生物学进程,23个参与细胞成分,25个参与分子功能。感染未致敏组与健康对照组特有的基因GO富集结果显示,51个参与生物学进程,13个参与细胞成分,18个参与分子功能。感染致敏组与健康对照组特有基因的功能主要集中于炎症反应、免疫应答等,共有基因的功能主要集中于神经肽信号通路、白细胞介素-1（IL-1）细胞反应等;感染未致敏组与健康对照组特有基因的功能主要集中于核小体组装、细胞外调节蛋白激酶1（ERK1）和ERK2级联的正调节等。感染致敏组与健康对照组特有基因被富集到细胞因子-细胞因子受体相互作用、JAK-STAT信号通路等32个通路;共有基因富集到磷脂酰肌醇3激酶-蛋白激酶B信号通路、补体与凝血级联反应等9个通路。感染未致敏组与健康对照组特有基因富集到酪氨酸代谢,亚油酸代谢等12个通路。感染致敏组和健康对照组特有基因PPI分析结果中,位于核心位置的包括IL-10、IL-6、前列腺素内过氧化物合酶2、CC趋化因子亚族-1、CXC趋化因子亚族-1/2/3等;感染未致敏组和健康对照组位于核心位置的包括17号染色体、19号染色体的基因、细胞色素P3a25等。感染致敏组和对照组,感染未致敏和健康对照组共有基因则包括IL-13、20号染色体、基质金属蛋白酶抑制剂-1、精氨酸酶1、触珠蛋白、纤溶酶原、17号染色体的基因等。 结论 炎症反应在棘球蚴致敏过程中发挥重要的作用;IL-6高表达促进炎症发生,IL-10作为抗炎因子对抗机体的炎症反应的发生;IL-13可能是棘球蚴感染后机体产生免疫耐受的关键基因;ERK信号通路可能在棘球蚴生长过程中起免疫调节作用,细胞因子-细胞因子受体相互作用通路和磷脂酰肌醇3激酶-蛋白激酶B信号通路与棘球蚴致敏显著相关。

Objective To analyze the sensitization mechanisms and molecular targets of Echinococcus granulosus using bioinformatics approaches. Methods Eighteen BALB/c mice were randomly grouped as infection- unsensitized (CE), infection-sensitized (ANPC), and uninfected controls (CTRL). Mice in the infected unsensitized group were injected with protoscoleces suspension (0.2 ml/mouse), followed with saline (0.2 ml/mice) after 90 days. Mice in the infected sensitized group were injected with protoscoleces suspension (0.2 ml/mice), followed with crude cyst fluid (0.2 ml/mice) 90 days after infection. Mice in the uninfected control group were injected with an equal amount of saline. Splenic tissue from three mice of each group were collected 1 h after injection for RNA extraction and transcriptome sequencing. Genes of differential expression were identified by bioinformatics analysis. Significance analysis of differential expression differences was performed using edgeR (quantified with RNA-Seq by expectation-Maximization RSEM) software. Network interaction analysis was performed using the String database and Cytoscape software with the score set as > 500. The network analysis of the genes was then performed using the Cytoscape software and use cytoHubba to obtain the degree of nodes (degree) and the nodes with degrees over 3 were filtered for display. The analysis was performed using the DAVID online enrichment analysis tool, and the results were visualized using the R language ggplot2. Results A total of 743 genes were differentially expressed in the ANPC group compared to the CTRL group, including 568 upregulated genes and 175 downregulated genes. A total of 554 genes were differentially expressed in the CE group compared to the CTRL group, including 393 upregulated genes and 161 downregulated genes. There are 458 genes specifically expressed in the ANPC group compared to the CTRL group, 269 genes specifically expressed in the CE group compared to the CTRL group and 285 genes co-expressed. GO enrichment of 458 genes, which are unique to the ANPC group, and the CTRL group showed that 147 genes are involved in the biological process, 10 genes are involved in cellular components, and 43 genes are involved in molecular functions. The GO enrichment of shared genes showed that 56 were involved in biological processes, 23 in cellular components, and 25 in molecular functions. The GO enrichment of genes that are unique to the CE group and CTRL group revealed that 51 genes are involved in biological processes, 13 genes are involved in cellular components, and 18 genes are involved in molecular functions. The functions of the ANPC group-specific genes and the CTRL group-specific genes mainly focus on inflammatory response and immune response. The functions of genes expressed in both the ANPC and CTRL groups mainly focus on the neuropeptide signaling pathway and cellular response to interleukin-1. The function of CE group-specific genes and the CTRL group-specific genes mainly focuses on the nucleosome assembly and the positive regulation of ERK1 and ERK2 cascades. The genes expressed explicitly in the ANPC group, and the CTRL group was enriched in 32 pathways, including cytokine-cytokine receptor interaction and the JAK-STAT signalling pathway. The genes commonly expressed in all groups were enriched in 9 pathways, including the phosphatidylinositol 3-kinase/protein kinase B (PI3K-Akt) signaling pathway and complement and coagulation cascades. The CE specific genes and the CTRL group-specific genes are enriched to 12 pathways, including Tyrosine metabolism and Linoleic acid metabolism, and identified key genes through the PPI network. Conclusion The inflammatory response plays an important role in metacestode sensitizing process; high expression of IL-6 promotes the occurrance of inflammation; IL-10 functions as an anti-inflammatory factor confronting the host’s inflammation response; IL-13 may be the key gene in developing immune tolerance after metacestode infection. ERK signaling pathway may play an immunomodulatory role during the growth of metacestodes. Cytokine-cytokine receptor interaction pathway and PI3K-Akt signaling pathway were significantly correlated with the sensitization induced by Einococcus infection.