中国兽医杂志2025，Vol.61Issue(5)：22-32,11.DOI:10.20157/j.cnki.zgsyzz.2025.05.003
基于转录组分析CSFV感染猪睾丸支持细胞对屏障功能相关基因的影响
Transcriptomic Analysis of the Effects of CSFV Infection on Barrier Function-Related Genes in ST Cells
摘要
Abstract
To investigate the impact of classical swine fever virus(CSFV)infection on the mRNA expression profile of swine testicular Sertoli cells(ST),ST cells infected with CSFV for 24 h and 48 h,along with uninfected control cells,were used as models.Western blotting(WB)and indirect immunofluorescence(IIF)staining were performed to detect CSFV E2 protein expression in ST cells.Four cDNA libraries were constructed for transcriptome sequencing,and differentially expressed genes(DEGs)were annotated using Gene Ontology(GO)and enriched via Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis.DEGs associated with the blood-testis barrier(BTB)were further screened and validated by real-time fluorescence quantitative PCR(qPCR),WB,and IIF.The results showed that E2 protein was expressed in ST cells at both 24 h and 48 h post-CSFV infection.Compared with Control 24 h,a total of 86 significant DEGs(P<0.05)were identified in the CSFV 24 h,and 1 481 significant DEGs(P<0.05)were identified in the CSFV 48 h compared with Control 48 h.These DEGs were significantly enriched in 27 GO terms,including biological processes,cellular components,and molecular functions(P<0.01).The most significantly enriched KEGG pathways included TNF signaling,AGE-RAGE signaling,IL-17 signaling,and MAPK signaling pathways(P<0.01).qPCR results showed that,at 24 h,CLDN11,H2AFX,and CDH2 were significantly upregulated(P<0.001 or P<0.01),while TJP1,OCLN,and GJA1 were significantly downregulated(P<0.05).At 48 h,FASLG,TGFB1,IL-6,and H2AFX were significantly upregulated(P<0.001 or P<0.01),whereas TJP1,OCLN,GJA1,CLDN11,and CDH2 were significantly downregulated(P<0.01).WB validation showed that protein levels of TJP1,CLDN11,and GJA1 were significantly downregulated at 24 h(P<0.001 or P<0.01),with CLDN11 showing significant upregulation(P<0.001).At 48 h,TJP1,CLDN11,OCLN,and GJA1 protein levels were all significantly downregulated(P<0.001 or P<0.01).IIF results at 48 h showed that the relative fluorescence area of TJP1 was significantly decreased(P<0.01),and CLDN11,OCLN,and GJA1 were also significantly reduced(P<0.05).These findings were consistent with transcriptome sequencing results,indicating that CSFV infection at both 24 h and 48 h alters the expression of barrier function-related genes in ST cells.This study provides data support for further exploration of how CSFV affects the barrier function of ST cells.关键词
猪瘟病毒(CSFV)/猪睾丸支持细胞/mRNA差异表达/基因本体论(GO)功能注释/京都基因与基因组百科全书数据库(KEGG)富集分析Key words
classical swine fever virus(CSFV)/sertoli cells/differential mRNA expression/Gene ontology(GO)function annotation/Kyoto encyclopedia of genes and genomes(KEGG)enrichment analysis分类
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马尧丹,俞晖,陈海恩,杨阳开心,葛书君,吴江,康恺..基于转录组分析CSFV感染猪睾丸支持细胞对屏障功能相关基因的影响[J].中国兽医杂志,2025,61(5):22-32,11.基金项目
广东省自然科学基金(2020A1515110362) （2020A1515110362）
广东省科技厅海外名师项目(K23438) （K23438）
湛江市科学技术局海洋生物方向基金(2021E05028) （2021E05028）
