CircRNA与系统性红斑狼疮发病相关性的分子流行病学研究

Previous studies have established the relevance of microRNA (miRNA) in the pathogenesis of systemic lupus erythematosus (SLE). Aberrant expression and function of miRNAs in T cells induce dysfunction of T cells and lead to the initiation and progression of SLE. Recent studies have shown that circular RNA (circRNA) can serve as miRNA sponge and thereby block the function of miRNA. CircRNA in nucleus can promote the transcription of its parental genes and compete with canonical splicing. Based on these evidences above, we hypothesize that aberrant expression of circRNA in T cells may affect its functions in sponging miRNA and regulating gene transcription, lead to dysfunction of T cells， and partake in the etiology of SLE. Therefore, we plan to collect circulating T cells and clinical data from SLE patients and healthy controls and then establish the expression profile of circRNA in T cells of SLE patients by virtue of microarray and Real-time PCR. As for the differentially expressed circRNA in SLE patients, we will utilize bioinformatics methods and perform overexpression and knockdown of circRNA, uncovering its role in T cell function such as cytokine secretion and the pathogenesis of SLE.

已有研究表明微小RNA（miRNA）与系统性红斑狼疮（SLE）发病密切相关。T细胞内miRNA的表达及功能异常引起SLE患者T细胞的功能异常,导致疾病的发生、发展。新近研究发现环状RNA（circRNA）可吸附miRNA，影响其功能发挥。核内circRNA具备增强其编码基因转录、与经典线性剪接竞争等功能。据此，我们假设：T细胞circRNA异常可通过影响其吸附miRNA和调控基因转录等功能，造成T细胞功能异常，从而参与SLE发病。本课题拟收集SLE患者和正常对照T细胞和相关临床资料，借助circRNA芯片和荧光定量PCR技术，建立SLE及不同临床亚型患者T细胞circRNA的表达谱；针对SLE患者T细胞异常表达circRNA，运用生物信息学、慢病毒过表达和RNA干扰等技术，探讨其在SLE患者T细胞炎症细胞因子分泌等功能异常中的作用，初步揭示circRNA在SLE发病中可能的作用机制。

本项目采用病例对照研究，使用环状RNA芯片（circRNA）和实时荧光定量聚合酶链反应（qRT-PCR）技术筛选并验证SLE患者T细胞和血浆中差异表达的circRNA，分析差异表达circRNA（hsa_circ_0045272）的表达水平与SLE患者的临床特征、实验室指标、疾病活动度间的关系；运用慢病毒过表达和RNA干扰等技术，探讨差异表达circRNA（hsa_circ_0045272）对细胞凋亡和IL-2分泌的影响；借助生物信息学方法结合专业知识，预测T细胞、血浆中差异表达circRNA（hsa_circ_0045272，hsa_circRNA_407176、hsa_circRNA_406567和hsa_circRNA_001308）的ceRNA，并构建共表达网络；选择2个T细胞中差异表达circRNA（hsa_circ_0045272）的ceRNA（NM_003466(PAX8)、NM_015177(DTX4)）进行验证，探讨潜在作用机制。结果发现，hsa_circ_0045272在SLE患者T细胞内表达降低，其表达水平与临床特征、实验室指标、疾病活动度无关；hsa_circRNA_407176和hsa_circRNA_001308在SLE患者的血浆中低表达，hsa_circRNA_407176、hsa_circRNA_406567和hsa_circRNA_001308在SLE患者的外周血单核细胞中低表达；血浆中hsa_circRNA_407176表达水平在Anti-SSB阳性组中更低，hsa_circRNA_001308表达水平与CRP呈负相关，未发现他们与其它临床特征、实验室指标、疾病活动度间的关联；血浆中hsa_circRNA_407176、hsa_circRNA_406567和hsa_circRNA_001308的表达水平与其在PBMC中的表达水平无关；hsa_circ_0045272沉默会促进细胞凋亡和IL-2的分泌；hsa_circ_0045272可能通过调控NM_003466(PAX8)和NM_015177(DTX4)的mRNA表达水平而发挥作用；hsa_circ_0045272可吸附hsa-miR-6127，但NM_003466(PAX8)和NM_015177(DTX4)不是hsa-miR-6127的靶基因；circRNA和mRNA存在强相关