LncRNA CTA-373H7.7对红斑狼疮患者Treg免疫调节的作用及机制研究

Systemic Lupus Erythematosus (SLE) is an autoimmune disease characterized as a T, B lymphocytes over-activation. Regulatory T cells (Treg) play an important role in the development of SLE. It has been proved that the frequency of Treg decreased in peripheral blood of SLE patients, and partial dysfunction of Treg activation. However, the mechanism of which remains unknown. Forkhead box protein3 (Foxp3) is a transcriptional factor specifically expressed in Treg, regulating thousands of proteins and non-coding RNAs and maintaining the function and homeostasis of Treg. Long non-coding RNA (lncRNA) has been found to express abnormally and take part in many autoimmune diseases. We have confirmed before that Foxp3 can bind the promoter region and regulate the expression of lncRNA CTA-373H7.7. And we have found that the expression of both Foxp3 and CTA-373H7.7 decreased in Treg of SLE patients. It suggested that the decrease of Foxp3 might affect the expression of CTA-373H7.7 and involves in the cell processes and immune functions. We will study the function and mechanism of CTA-373H7.7 in Treg, explore the influence of non-coding RNA regulated by Foxp3 in Treg, and thus provide a new theory for clinical therapy of SLE.

系统性红斑狼疮（SLE）是一种T、B淋巴细胞过度活化的自身免疫病，Treg在SLE发生发展中具有重要作用。研究证实SLE患者外周血Treg数量明显下降、Treg细胞功能缺陷，但具体机制尚不明确。Foxp3是Treg细胞中特异性表达的转录因子，通过调控多种蛋白质和非编码RNA的表达维持细胞功能。研究表明lncRNA在多种免疫病中表达异常并具有重要的调控功能。前期研究发现Foxp3结合lncRNA CTA-373H7.7启动子区并调控其表达，且SLE患者Treg中Foxp3和CTA-373H7.7表达均显著降低。这表明SLE患者外周血Treg中Foxp3表达下降可能抑制CTA-373H7.7的表达，并影响细胞进程及免疫抑制功能。本课题将深入研究CTA-373H7.7对Treg的调控及作用机制，揭示Foxp3调控的lncRNA对疾病发生发展的可能影响，为治疗SLE提供理论依据。

系统性红斑狼疮是一种淋巴细胞过度活化的自身免疫病，而调节性T细胞是一类调节体内自身免疫反应性的T细胞亚群，在SLE发生发展中具有重要作用。LncRNA是一类长度大于200nt的非编码RNA，能够在表观遗传修饰、转录及转录后等方面调控基因表达。本研究取HC与SLE病人CD4+ Naïve T及Treg细胞进行RNA-seq，发现在Naïve T向Treg分化过程中以及HC与SLE病人Treg细胞中lncRNA表达发生明显变化，实时定量PCR检测证实多个lncRNA表达发生显著变化并且与体外诱导Treg形成过程中的变化一致。进一步研究发现lncRNA-CTA-373H7.7在Treg分化过程中明显上调且主要分布在细胞核中。研究证实在SLE患者Treg数量明显下降、Treg细胞功能缺陷，Foxp3和CTA-373H7.7表达均显著降低。根据Foxp3-Chip及荧光素酶报告基因等实验结果发现Foxp3通过与CTA-373H7.7启动子区结合调控其表达。CD4+NaïveT细胞中过表达CTA-373H7.7，体外诱导分化发现Treg分化比率明显增加。这表明SLE患者外周血Treg中Foxp3表达下降可能抑制CTA-373H7.7的表达，并影响细胞进程及免疫抑制功能。MicroRNAs是一类长度18-25nt非编码小RNA，主要功能为基因水平的转录后调控。临床治疗SLE过程中发现治疗前后SLE患者循环miRNA表达谱变化明显，根据测序结果分析及实时定量PCR验证发现hsa-miR-196a-5p和hsa-miR-320b在治疗后显著下降，并与疾病和临床活动度密切相关，且它们共同靶基因MAP3K1的表达在治疗后明显上调，表达水平与SLEDAI评分、尿蛋白等多个临床指标密切相关。过表达miR-196a-5p和miR-320b后通过实时定量PCR、Western blotting以及荧光素酶报告基因方法确认miR-196a-5p和miR-320b调控MAP3K1。利用MRL/lpr狼疮鼠模型验证miR-196a-5p和miR-320b能够加重SLE疾病，逆转治疗效果。因此本研究深入研究CTA-373H7.7对Treg的调控作用以及hsa-miR-196a-5p和hsa-miR-320b对SLE的影响，揭示了非编码RNA对疾病发生发展的影响，为疾病诊治提供新的思路。