miR-19 在视神经脊髓炎疾病中的作用机制研究

Neuromyelitis optica (NMO) is a refractory disabling inflammatory disorder of the central nervous system (CNS) in China. The discovery of antibody targeting to aquaporin-4 (NMO-IgG or AQP4-Ab) in NMO patients leads to new methods of diagnosis and treatment for NMO. However, in some NMO patients, the AQP4-Ab was not detected. Meanwhile, the AQP4-Ab was detected in some non-NMO patients. So, it is very important to understand the mechanisms of autoimmune in CNS so that we can find a new target to diagnose and treat the NMO disease. In order to find a new target, we had sequenced the expressions of all microRNAs in NMO patients, and we found that the expressions of miR-19 cluster in NMO patients were lower than that in normal health volunteers. And then we compared the expressions of miR-19 cluster in NMO patients with MS patients. We found the miR-19 expression in NMO has a significantly difference compared with that in MS. We also analyzed the relationship between the expressions of miR-19 cluster and the courses of NMO disease. MicroRNAs belong to a regulatory class of small noncoding RNAs with fundamental roles in numerous aspects of cell biology. MiR-19 not only regulates the development of B cells, promotes Treg cells survival and regulates effector versus memory T cells differentiation, but also regulate the secrets of cytokines. Therefore we hypothesize that miR-19 may be a new target of NMO disease. In this program, we will construct the peripheral blood mononuclear cell which stable expressed miR-19 and use the neuromyelitis optica experimental animal model to investigate the targeting cells of miR-19, the roles of miR-19 in immune regulation and the mechanisms in NMO disease. And then, we will further reveal the potential targets of miR-19, and the relationship between miR-19 expressions and the process of NMO. The outcome of this project will enhance our understanding of NMO, promote appearing new diagnostic criteria of NMO, and then pave new avenues for NMO therapy.

视神经脊髓炎(NMO)是一种严重威胁人类健康的难治性疾病。水通道蛋白4抗体(AQP4-Ab)的发现为NMO的诊断和治疗开辟了新的方法，但部分患者血清内不表达AQP4-Ab，因此寻找新的靶点有助于NMO的诊断和治疗。我们对NMO患者进行miRNome基因测序，并检测80多例患者，结果发现miR-19在NMO患者中的表达较低，并发现miR-19的表达和NMO进展有密切关系。研究表明，miR-19可以调节B细胞的发育，又可以调节细胞因子的分泌，在自身免疫疾病中发挥着重要的作用，因此推测miR-19是NMO的新靶点。本申请将构建稳定表达miR-19的外周血单个核细胞，利用改良的NMO实验动物模型，探讨miR-19在NMO疾病中的作用，在此基础上探讨miR-19调节中枢神经系统免疫微环境以及NMO疾病潜在的细胞和分子机制。本项目的研究成果将为NMO的早期诊断和针对治疗提供新的靶点和策略。

视神经脊髓炎（neuromyelitis optica, NMO）是一种严重威胁人类健康的中枢神经系统炎性脱髓鞘疾病。到目前为止，对NMO的治疗并不理想，所以增加对NMO的研究有助于我们找到NMO新的诊断指标和作用靶点，为NMO的治疗提供理论基础，具有重要的社会意义和临床价值。MicroRNA是一类长度在21-22个核苷酸的调节性非编码RNA，增加对miRNA的研究，了解microRNA在神经免疫疾病中作用的机制，可以为神经免疫疾病的诊断、治疗提供理论基础和治疗靶点，具有重大的临床意义。骨髓间充质干细胞具有多重分化、免疫调节和神经营养等作用，所以应用自身来源的骨髓间充质干细胞治疗NMO被寄予厚望。但是NMO来源与健康人来源的骨髓间充质干细胞的细胞特性、生物学功能以及免疫调节是否一致，却很少有人研究。我们提取NMO患者、健康对照和MS患者外周血进行测序；然后用实时定量PCR方法对检测结果进行验证；然后利用流式细胞仪检测NMO患者各个细胞亚型中microRNA的表达高低；提取NMO患者和健康对照骨髓间充质干细胞，利用流式细胞仪和免疫组化检测骨髓间充质干细胞细胞表面分子标记和成骨、成脂分化能力，利用实时定量PCR和蛋白免疫印迹检测其细胞内分子表达差异。结果显示microRNA-17-92在NMO患者外周血中表达比正常人低，比MS患者高，而且microRNA-17-92表达差异与NMO的进程直接相关。我们用流式细胞仪检测miR-19在各个细胞中的表达。结果发现，浆细胞和辅助性T细胞内miR-19表达较高，而在Th1、Th17和调节性T细胞内miR-19表达较低。我们比较了NMO来源和健康对照来源的骨髓间充质干细胞的增殖能力和细胞凋亡，结果发现NMO来源的骨髓间充质干细胞增殖能力降低和细胞凋亡增多。而当用PDGF处理后，发现PDGF可以促进骨髓间充质干细胞增殖降低细胞凋亡，从而有助于NMO的治疗。本项目对microRNA-17-92在视神经脊髓炎中的表达情况以及应用骨髓间充质干细胞对NMO治疗的分子机制和理论基础进行了研究，对microRNA-17-92在NMO发病机制中的作用以及通过应用骨髓间充质干细胞治疗NMO的理论基础进行了初步探索。