摘要
筛选树鼩真菌性角膜炎发病过程中差异表达的微小RNA(microRNA,miRNA)和信使RNA(mRNA),分析其靶基因的生物学功能及信号通路,为阐明真菌性角膜炎的免疫机制及筛选其抗真菌治疗的潜在靶点提供线索。通过茄病镰刀菌液显微注射至树鼩角膜基质,诱导真菌性角膜炎的发生;使用高通量测序技术检测茄病镰刀菌感染树鼩第7天、14天和30天的角膜组织miRNA和mRNA表达谱,以自身未感染的眼角膜为对照;差异表达miRNAs的靶基因与差异表达mRNAs取交集构建miRNA-mRNA调控网络,采用实时荧光定量PCR进行验证;最后对靶基因进行GO和KEGG富集分析。结果表明:在茄病镰刀菌感染树鼩角膜第7天、14天和30天,分别有78、56、41个mRNAs和70、63、20个miRNAs显著性差异表达,其中有29个mRNAs和14个miRNAs在感染第7天、14天和30天均显著性差异表达,Has-miR-214-3p和Has-miR-149-5p处于调控网络的关键节点。GO和KEGG富集分析发现,显著性差异表达的mRNAs和miRNAs靶基因主要参与免疫反应和炎症反应,涉及Toll-like受体、NOD-like受体、NF-kB和TNF信号通路。研究结果明确了树鼩真菌性角膜炎存在miRNA-mRNA差异表达基因谱,Has-miR-214-3p、Has-miR-149-5p与树鼩真菌性角膜炎相关,可作为真菌性角膜炎发生和预后评价的潜在生物学标志物。
The objective of this study is to identify the differentially expressed miRNAs and mRNAs in tree shrew fungal keratitis and analyze the miRNA-mRNA regulatory network and its bioinformatics,thus providing clues for the immune mechanism of fungal keratitis as well as the potential targets for antifungal therapy.Fusarium solani solution was microinjected into the corneal stroma of tree shrew to induce the occurrence of fungal keratitis.After 7,14 and 30 days of infection,the expression levels of miRNAs and mRNAs in cornea tissues of tree shrews were detected through high-throughput RNA sequencing technology.The corneas of non-infected eyes were taken as control.The intersection of the differentially expressed mRNAs and the targeted genes of the differentially expressed miRNAs were selected to construct the miRNA-mRNA regulatory networks.Real-time PCR was used to verify the validity of sequencing.Finally,GO and KEGG enrichment analysis were performed.The results showed that at day 7,14 and 30 of infection,78,56 and 41 mRNAs and 70,63 and 20 miRNAs were differentially expressed,respectively.Among them,29 mRNAs and 14 miRNAs were differentially expressed at day 7,14 and 30 of infection.Has-miR-214-3p and Has-miR-149-5p were the key nodes in the regulatory network.GO and KEGG enrichment analysis revealed that the differentially expressed mRNAs and miRNA targeted genes were mainly involved in immune response and inflammatory response,including Toll-like receptor,NOD-like receptor,NF-kB and TNF signaling pathways.Our finding clarified that there were differentially expressed gene profiles of miRNA-mRNA in the fungal keratitis of tree shrews,and Has-miR-214-3p and Has-miR-149-5p were the related nodes which could be studied as potential biomarkers for the occurrence and prognosis evaluation of fungal keratitis.
作者
贾杰
黎晓慧
黄鑫
李明学
施梅言
邱丹丹
仝品芬
代解杰
JIA Jie;LI Xiaohui;HUANG Xin;LI Mingxue;SHI Meiyan;QIU Dandan;TONG Pinfen;DAI Jiejie(Institute of Medical Biology,Chinese Academy of Medical Science and Peking Union Medical College,The Key Laboratory of Yunnan Province for Ophthalmic Research and Disease Control,Yunnan Innovation Team of Standardization and Application Research in Tree Shrew,Kunming 650118,Yunnan,China)
出处
《陕西师范大学学报（自然科学版）》
CAS
CSCD
北大核心
2020年第5期87-97,共11页
Journal of Shaanxi Normal University：Natural Science Edition
基金
云南省眼科疾病防治研究重点实验室(2017DG008)
中央高校基本科研业务费专项资金(3332018162)
云南省科技人才和平台计划项目(2017HC019)。
