白介素-27作用于树突状细胞CD39/ATP轴参与调控哮喘气道炎症和气道高反应性的机制研究

The polarization of Th2 and Th17 subsets of T helper cells plays an important role in the development of asthma, but the regulatory mechanism is still unknown. It has been demonstrated that extracellular ATP (eATP) can lead to the activation of NLRP3 inflammasome, inducing the release of large amount of interleukin (IL)-1β and IL-18, augmenting Th2 and Th17 immune response. Our previous studies showed that the serum IL-27 levels in asthmatic mice were lower than that of control, and IL-27 receptor knock-out (IL-27R-/-) mice exhibited severer airway hyper-responsiveness and airway inflammation. Meanwhile, we found that IL-27 functional deficiency induced lower expression of CD39, an eATP hydrolase, on dendritic cells (DCs). JAK/STAT signaling pathway mediates the function of IL-27. And bioinformatics analysis recently identified several putative STAT-binding elements in upstream of the transcription start site in the CD39 gene (Entpd1) promoter. Therefore, we assume that IL-27 suppresses airway inflammation and airway hyper-responsiveness in asthma by acting on dendritic cells CD39/ATP axis through JAK/STAT signaling pathway. For this purpose, we take advantage of IL-27R-/- and CD39-/- mice to establish asthmatic animal models and demonstrate that the dysregulation of IL-27 on dendritic cells CD39/ATP axis in vivo. Then, we confirm that dendritic cells CD39/ATP axis is a key junction in the function of IL-27 in vitro. Ultimately, we clarify the mechanism of JAK/STAT signaling pathway in the regulation of IL-27 on CD39 expression, which contributes to developing new approaches for therapeutic intervention.

辅助性T细胞（Th）极化是哮喘发生的重要机制，但尚未完全阐明。研究证实，危险信号ATP能激活NLRP3炎性小体，大量释放白介素（IL）-1β和IL-18，促进Th2与Th17极化。我们前期发现，①哮喘小鼠血清IL-27水平降低，IL-27受体敲除的哮喘小鼠气道炎症加重；②IL-27功能缺陷下调树突状细胞ATP水解酶CD39表达。JAK/STAT通路介导IL-27信号，新近发现CD39基因调控序列也存在STAT结合位点。据此假设，IL-27可能通过JAK/STAT信号通路作用于树突状细胞CD39/ATP轴并抑制哮喘气道炎症和气道高反应性。故本项目拟证实哮喘小鼠体内存在IL-27对树突状细胞CD39/ATP轴的异常调控，体外研究阐明树突状细胞CD39/ATP轴是IL-27功能调控的关键枢纽，最后明确JAK/STAT信号通路介导IL-27调控CD39表达的分子机制，为临床防治哮喘提供新思路。

辅助性T细胞（Th）极化是哮喘发生的重要机制。研究证实，危险信号ATP能激活NLRP3炎性小体，大量释放IL-1β和IL-18，促进Th2与Th17细胞极化。我们前期研究发现，（1）哮喘小鼠体内IL-27表达降低；（2）IL-27功能缺陷下调树突状细胞ATP水解酶CD39表达。JAK/STAT信号通路介导IL-27信号，新近发现CD39基因调控序列也存在STAT结合位点。据此假设，IL-27可能通过JAK/STAT信号通路作用于树突状细胞CD39/ATP轴参与调控哮喘小鼠辅助性T细胞极化。本项目研究证实，（1）IL-27Rα基因敲除加重哮喘小鼠气道高反应性和气道炎症特征；（2）IL-27Rα-/-哮喘小鼠肺组织CD39表达水平较野生型（WT）哮喘小鼠下降，NLRP3炎性小体组分NLRP3、ASC和Caspase-1表达增加，BALF中ATP浓度和IL-1β、IL-18含量升高。（3）IL-27Rα-/-哮喘小鼠肺树突状细胞CD39表达水平较WT哮喘小鼠下降。与WT小鼠骨髓诱导的树突状细胞（WT-BMDC）比较，IL-27Rα-/-小鼠骨髓诱导的树突状细胞（IL-27Rα-/--BMDC）CD39表达水平降低，而培养液上清中ATP含量显著增加。IL-27Rα-/--BMDC NLRP3炎性小体组分表达也明显高于WT-BMDC。（4）外源性rmIL-27能促进WT-BMDC表达CD39，降低培养液中ATP含量，诱导WT-BMDC NLRP3炎性小体组分表达水平下调。（5）rmIL-27干预WT-BMDC能引起JAK/STAT信号通路中磷酸化的JAK1、JAK2、STAT1和STAT3表达增加。据此推论，IL-27Rα基因敲除加重哮喘小鼠气道炎症和气道高反应性，其可能的机制是IL-27功能缺陷导致哮喘小鼠树突状细胞CD39表达水平降低，胞外ATP含量剧增，引起树突状细胞NLRP3炎性小体活化，大量释放促炎因子IL-1β和IL-18，加剧Th2和Th17反应偏倚。上述研究结果为今后开发IL-27基因修饰的树突状细胞疫苗或靶向RNA干扰树突状细胞JAK/STAT信号通路的药物奠定理论基础。