MicroRNA在慢性阻塞性肺疾病急性加重粘液高分泌及预后评估中的作用

Studies have shown that MicroRNA were closely related to the severity of emphysema, lung function and muscle weakness of COPD patients. The mucus hypersecretion means poor prognosis of patients with COPD. In this study，to find the differentially expressed miRNAs, we will measured the blood microRNA levels between acute exacerbation and stable COPD patients. At the same time, patients with AECOPD were followed for 1 year, the relationship between the differentially expressed microRNAs and the prognosis of the AECOPD patients were assessed. The prognostic system bases on microRNAs for hospital-mortality,1 year-mortality and 1 year exacerbations of AECOPD patients were established. To study the role of microRNA in the hypersecretion of AECOPD, We will established a COPD rat model caused by cigarette smoke. The miRNAs of lung tissue was measured. The mRNA and the protein of MUC5AC, EGFR-ERK signaling pathway gene of lung tissue were measured. We will also explored the role of microRNA in hypersecretion of COPD in celler level. Human bronchial epithelial cells were cultured, cigarette smoke extract(CSE) were added. The microRNA mimics and the anti-microRNA were added in the culture too. The microRNAs of human bronchial epithelial cells was measured. The mRNA and the protein of MUC5AC, EGFR-ERK signaling pathway gene of human bronchial epithelial cells were measured too.

研究显示MicroRNA与COPD患者的肺气肿、肺功能、肌无力严重程度及粘液高分泌密切相关。粘液高分泌与COPD患者预后不良相关。本课题以COPD急性加重期和稳定期患者为研究对象，筛选出差异表达miRNA,同时随访观察AECOPD患者1年，观察其住院死亡、1年内急性加重及死亡情况，评估差异表达miRNA与患者预后的关系，建立基于miRNA的AECOPD患者住院死亡、1年内急性加重及死亡的预后评估系统；并在另一群AECOPD患者中加以验证。在细胞水平，以香烟烟雾提取物刺激人支气管上皮细胞，并通过miRNA模拟和抑制技术，来研究miRNA在气道粘液高分泌中的作用和信号传导机制。在动物水平，通过复制香烟COPD大鼠模型，检测其肺组织中目标miRNA,及MUC5AC、EGFR-ERK信号通路基因在RNA水平和蛋白水平的表达，研究miRNA气道粘液高分泌中的作用和机制，为COPD的防治寻找新的靶点。

研究背景.气道的粘液高分泌与COPD患者频繁急性加重、肺功能加速下降、高住院治疗率以及高死亡率密切相关。近年来研究发现，miRNA参与了COPD的发生与发展。循环血液中与粘液分泌相关miRNA的有可能成为判断患者预后的生物标记物。目前粘液分泌相关miRNA在COPD中的作用机制尚不清楚。.主要研究内容与结果.通过高通量测序分析初筛队列粘液高分泌和非粘液高分泌COPD两组患者的血浆miRNA表达谱，筛选出在所有样本均有表达，且两组间有显著差异表达的miRNA（差异表达倍数≥2，p<0.05）共有53种，根据生物信息学分析及扩大样本量验证出14种与粘液分泌相关miRNA.同时队列研究发现hsa-miR-375-3p与AECOPD患者出院后一年内死亡和急性加重再次住院住院密切相关，而hsa-miR-485-3p与AECOPD患者出院后再次急性加重和死亡不相关。在动物水平发现香烟烟雾暴露可致大鼠hsa-miR-375-3P表达下降,SP1和磷酸化ERK表达增加，粘液蛋白MUC5AC表达增加。同时,在细胞水平，以香烟烟雾提取物刺激人支气管上皮细胞，并通过miRNA模拟和抑制技术来研究hsa-miR-375-3P香烟烟雾所致COPD粘液高分泌中的作用。发现香烟烟雾提取物刺激后MUC5AC与hsa-miRNA-375-3P的表达呈负相关关系，hsa-miR-375-3P可以通过结合sp1RNA抑制sp1蛋白翻译来下调MUC5AC的表达..科学意义.本研究筛选出了粘液分泌相关的miRNA，并同时在动物水平和细胞水平发现hsa-miR-375-3p在香烟烟雾所致COPD粘液高分泌中的可能机制，将有可能为慢阻肺的诊断和治疗提供新的药物靶标。同时本研究为临床提供一个能够预测AECOPD患者1年内急性加重住院和死亡的生物标志物，对AECOPD患者的管理，降低其未来急性加重发作的风险具有一定的价值。