Abstract: Objective　To screen differentially expressed long non⁃coding RNAs (lncRNAs) in the liver of mice infected with Schistosoma japonicum during the chronic pathogenic stage and identify their functions, so as to provide insights into unraveling the role of lncRNAs in S. japonicum infection⁃induced liver disorders. Methods　Twenty 6⁃week⁃old C57BL/6 mice were randomly divided into two groups, of 10 animals each group. Each mouse in the experimental group was infected with (15 ± 2) S. japonicum cercariae via the abdomen for modeling chronic S. japonicum infection in mice, and distilled water served as controls. All mice were sacrificed 70 days post⁃infection, and mouse liver specimens were sampled for RNA extraction and library construction. All libraries were sequenced on the Illumina NovaSeq 6000 sequencing platform. Data cleaning was performed using the fastp software, and reference genome alignment and gene expression (FPKM) calculation were performed using the HISAT2 software. Potential lncRNA sequences were predicted using the software CNIC, CPC, Pfam, and PLEK, and potential lncRNAs were screened. Differentially expressed lncRNAs were screened with the DESeq2 software and subjected to Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses to identify biological processes and metabolic pathways involved in target genes of differentially expressed lncRNAs. Results　A total of 333 potential lncRNAs were screened, and 67 were identified as differentially expressed lncRNAs, including 49 up⁃regulated and 18 down⁃regulated lncRNAs. A total of 53 target genes were predicted for differentially expressed lncRNAs. GO enrichment analysis showed that these target genes were mainly enriched in biological process and molecular function, among which Sema7a, Arrb1, and Ccl21b genes may be hub target genes for positive regulation of extracellular regulated protein kinase 1 (ERK1) and ERK2 cascades and may participate in the regulation of collagen expression. KEGG enrichment analysis showed that the target genes of differentially expressed lncRNAs were mainly enriched in cytokine⁃cytokine receptor interaction, viral protein interactions with cytokines and cytokine receptors, chemokine signaling pathway, and nuclear factor kappa⁃B (NF⁃κB) signaling pathway. Conclusions　This study identifies differentially expressed lncRNAs and functional enrichment of their target genes in the liver of mice during the chronic pathogenic stage of S. japonicum infection. Up⁃regulated lncRNAs may affect biological processes of ERK1/2 cascades and chemokine signaling pathways via target genes Sema7a, Arrb1, and Ccl21b, thereby affecting collagen expression and inflammatory signal pathways, ultimately affecting the development of liver disorders.

Key words: Schistosoma japonicum, Long non?coding RNA, Hepatic fibrosis, Extracellular regulated protein kinase, Mouse

摘要： 目的　筛选日本血吸虫感染小鼠慢性致病阶段肝脏中差异表达长非编码RNA（long non⁃coding RNA, lncRNA）并进行功能分析，为探索lncRNA在日本血吸虫感染所致肝脏病变中的作用提供参考。方法　20只6周龄C57BL/6小鼠随机分为2组，每组10只。实验组每只小鼠采用腹部贴片法感染（15 ± 2）条日本血吸虫尾蚴建立日本血吸虫慢性感染小鼠模型，以蒸馏水作为对照组。两组小鼠均于感染70 d后剖杀，获取小鼠肝脏组织样本，进行RNA抽提及文库构建。通过Illumina NovaSeq 6000 测序平台对文库进行测序，采用 fastp软件进行数据清洗，使用 HISAT2软件进行参考基因组比对及基因表达量（FPKM）计算。采用CNIC、CPC、Pfam、PLEK软件对潜在lncRNA序列进行编码潜能预测，筛选出潜在lncRNA。采用DESeq2软件进行基因差异表达分析，筛选出差异表达lncRNA。通过基因本体论（Gene Ontology，GO）和京都基因和基因组百科全书（Kyoto Encyclopedia of Genes and Genomes，KEGG）富集分析，挖掘差异表达lncRNA靶基因参与的生物学过程和代谢途径。结果　共筛选出333个潜在lncRNA，67个鉴定为差异表达lncRNA，其中49个表达上调、18个表达下调。差异表达lncRNA预测靶基因共53个，GO富集分析显示这些靶基因主要富集在生物学过程和分子功能；其中Sema7a、Arrb1、Ccl21b等基因可能是细胞外调节蛋白激酶1（extracellular regulated protein kinase，ERK1）和ERK2级联正调控生物学过程的关键靶基因，可能参与调控胶原蛋白表达。KEGG富集分析显示，差异表达lncRNA靶基因主要参与细胞因子⁃细胞因子受体相互作用、病毒蛋白与细胞因子和细胞因子受体的相互作用、趋化因子信号通路和核因子κB（nuclear factor kappa⁃B，NF⁃κB）通路等信号通路。结论　本研究鉴定了日本血吸虫感染小鼠慢性致病阶段肝脏中差异表达lncRNA及其靶基因的功能富集，其中上调表达的lncRNA可能通过调控Sema7a、Arrb1、Ccl21b等靶基因影响ERK1/2级联等生物学过程以及趋化因子信号通路等，影响胶原蛋白表达及炎症相关信号通路，从而影响肝脏病变发展。 　

关键词: 日本血吸虫, 长非编码RNA, 肝纤维化, 细胞外调节蛋白激酶, 小鼠