MicroRNA在牛病毒性腹泻病毒感染牛外周血CD14+单核细胞调控抗病毒反应的机制研究

项目名称： MicroRNA在牛病毒性腹泻病毒感染牛外周血CD14+单核细胞调控抗病毒反应的机制研究

项目编号： No.31502104

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 农业科学

项目作者： 高闪电

作者单位： 中国农业科学院兰州兽医研究所

项目金额： 18万元

中文摘要： MicroRNA在调控抗病毒固有免疫反应方面具有重要的生物学功能，但目前并不清楚其在牛病毒性腹泻病毒（BVDV）感染宿主免疫细胞过程中的调控机制。本项目在前期利用qRT-PCR分析获得致细胞病变型(CP)和非致细胞病变型(NCP) BVDV感染导致牛外周血CD14+ 单核细胞部分microRNA差异表达信息的基础上，进一步采用高通量测序技术全面分析细胞在CP型和NCP型BVDV感染前后的microRNA差异表达谱，以双荧光素酶报告系统验证免疫调控性microRNA对相关靶基因的作用，进而采用microRNA过表达、microRNA抑制、抗病毒细胞因子定量、NF-κB和PKR活性分析、抗病毒蛋白Mx检测、病毒滴度测定等一系列鉴定技术，研究microRNA在BVDV感染牛外周血CD14+ 单核细胞调控抗病毒反应的作用机制，为新型高效抗病毒制剂及其它免疫制剂的研制提供有价值的靶标分子。

中文关键词： 牛病毒性腹泻病毒；牛外周血单核细胞；微小RNA；抗病毒反应；细胞因子

英文摘要： MicroRNAs play important bio-functional roles in the regulation of antiviral innate immunity. However, the regulatory function of microRNA in the infection of bovine viral diarrhea virus (BVDV) on host immune cells remains unknown. Our preliminary analysis using qRT-PCR demonstrated that some microRNAs in bovine peripheral blood CD14+ monocytes were differentially expressed after infected with cytopathic and noncytopathic bovine viral diarrhea virus (BVDV). In the current study, the differential microRNA expression profiles in bovine peripheral blood CD14+ monocytes after infected with cytopathic and noncytopathic BVDV will be comprehensively analyzed by high-throughput sequencing technologies. The activities of immune-regulatory microRNAs to their target genes will be validated by dual-luciferase reporter assay, and the roles of microRNAs in regulation of antiviral reaction in BVDV-infected bovine peripheral blood CD14+ monocytes will be observed via a series of assays including microRNA over-expression, microRNA suppression, antiviral cellular cytokine quantitation, NF-κB and protein kinase PKR activity analysis, antiviral protein Mx detection and virus titer determination. These findings will contribute to valuable targets for new antiviral agent and immunological preparations.

英文关键词： Bovine viral diarrhea virus;bovine peripheral blood monocytes;microRNAs;antiviral reaction;cellular cytokines