MicroRNA-204-5p调控EphB2在腹泻型肠易激综合征内脏高敏感发生中的作用及机制

Irritable bowel syndrome (IBS) is a functional bowel disorder characterized by visceral hypersensitivity. Achievement of a better understanding of the causes of visceral pain will provide a basis for future treatments for IBS. MicroRNAs (miRNAs) are endogenously expressed small non-coding RNAs and have emerged as regulators involved in many human diseases. In our preliminary study, we defined a part of the miRNAs analysis in the serum of IBS-D patients and healthy controls and discovered a set of differentially expressed miRNAs. By using miRNAs target genes prediction tool, miR-204-5p was chosen as a promising candidate for inducing visceral pain via targeting EphB2 which was involved in axon guidance signaling pathway. EphB2 activation was previously confirmed to promote the reconstruction of synapse and nociceptive information transmission. This study will evaluate whether miR-204-5p modulate visceral hypersensitivity in IBS-D patients via targeting EphB2, a critical protein related to pain sensation. The results will be helpful to elucidate the pathogenesis of visceral pain and provide new targets for IBS therapy.

腹泻型肠易激综合征（IBS）是以内脏感觉异常为主要特征的肠道功能紊乱性疾病。探索降低IBS患者内脏高敏感状态的方法，对临床上IBS患者的症状缓解具有重要意义。MicroRNAs（miRNAs）是由内源基因组编码的单链小分子RNA。多项研究表明miRNAs表达异常参与了多种疾病的发生。我们在前期研究中发现IBS-D患者血清样本small RNA测序结果显示miR-204-5p显著下调。通过Targetscan等miRNA靶基因预测软件以及查阅文献报道过的已经验证的miR-204-5p靶基因，我们发现EphB2受体可直接受其调控，该受体可通过改变神经元活性以及突触可塑性参与多种疼痛的发生。因此本课题拟在前期工作基础上，探讨IBS患者异常表达的miR-204-5p能否通过调控其靶基因EphB2的表达参与内脏高敏感的发生，其结果将有助于为IBS的治疗提供新的治疗靶点。

腹泻型肠易激综合征（IBS-D）是以腹痛为主要特征的肠道功能紊乱性疾病。多数研究表明IBS患者内脏感觉异常，主要表现为引起疼痛的刺激阈值降低以及对疼痛反应的放大。因此探索降低IBS患者结肠高敏感状态的方法具有重要意义。大量研究表明miRNAs表达异常可参与多种疾病的发生。然而尚不清楚其是否在IBS内脏高敏感的发生发展中发挥调控作用。本研究中，我们通过前期small RNA测序筛选出IBS-D患者与对照组相比具有显著表达差异的miRNAs，并通过分析miRNAs靶基因相关富集通路，初步选定miR-204-5p作为研究对象，同时利用qPCR技术在TNBS内脏高敏感小鼠结肠组织中进行验证。结合相关文献报道过已验证的靶蛋白及生物信息学预测方法，我们发现EphB2可能直接受其调控。接下来我们在体外应用双荧光素酶报告基因实验以及western blots等方法证实了miR-204-5p与EphB2之间的靶向调控关系。为进一步确定miR-204-5p的来源，我们进行了肠道肌间神经丛的分离，流式分选肠胶质细胞送测序。最后我们在IBS动物模型中探讨了miR-204-5p的表达降低可能导致其靶基因蛋白EphB2上调，进而促进c-FOS蛋白表达增多，导致神经元活化，促进内脏高敏感的发生及发展。这些结果有助于我们深入认识IBS的病理生理学机制，为以后治疗提供新的方向。