Dickkopf相关蛋白1抑制肺炎支原体P1-C诱导小鼠肺上皮细胞过度分泌MUC5AC

Dickkopf-1 inhibits the secretion of MUC5AC induced by Mycoplasma pneumoniae P1-C in mouse lung epithelial cells<

摘要:

肺炎支原体(Mycoplasma pneumoniae)是儿童和成人最常见的呼吸道感染病原体。临床观察肺炎支原体感染会引起呼吸道黏液大量分泌，给患者呼吸造成困难，已有研究表明肺炎支原体感染会引起大量黏蛋白5AC (mucin 5AC, MUC5AC)的分泌。肺炎支原体P1黏附素通过介导病原体与宿主细胞的黏附在肺炎支原体感染的发病机制中发挥重要作用，其中P1的C-末端残基(P1-C)具有免疫原性。本研究探讨了Wnt (Wingless, Wnt)/β-catenin信号通路抑制因子Dickkopf-1 (Dickkopf-1, DKK1)在肺炎支原体P1-C诱导的肺上皮细胞分泌黏蛋白MUC5AC的分子机制。利用扫描电镜(scanning electron microscope, SEM)、苏木精-伊红(hematoxylin-eosin, HE)染色观察肺炎支原体P1-C对小鼠肺上皮细胞(mouse airway epithelial cells, MAECs)黏液分泌的影响；利用蛋白芯片技术检测肺炎支原体P1-C对小鼠气道上皮细胞炎症因子分泌及对相关信号通路的富集分析；采用糖原染色(periodic acid schiff stain, PAS)、Tunel染色、Masson染色检测肺炎支原体P1-C对小鼠肺的损伤情况；采用免疫组化检测黏蛋白MUC5AC的分泌情况，采用蛋白免疫印迹检测DKK1调控肺炎支原体P1-C蛋白诱导小鼠肺上皮细胞分泌黏蛋白MUC5AC的分子机制。结果表明，肺炎支原体P1-C能够引起小鼠原代上皮细胞大量黏液和炎性因子的分泌，在肺炎支原体P1-C感染中，DKK1能下调JAK激酶2 (janus kinase 2, JAK2)、磷酸化信号传导与转录激活因子1 (phosphorylation signal transducer and activator of transcription 1, p-STAT1)和磷酸化信号传导与转录激活因子3 (phosphorylation signal transducer and activator of transcription 3, p-STAT3)蛋白的表达；同时，DKK1过表达显著上调MUC5AC抑制转录因子叉头框蛋白A2 (fork-head box A2, FOXA2)的表达，从而显著抑制了肺炎支原体P1-C诱导的MUC5AC的表达。通过该研究推测DKK1通过抑制JAK/STAT1-STAT3信号通路以及上调FOXA2的表达有效地减少肺炎支原体P1-C诱导的小鼠肺上皮细胞MUC5AC的分泌。

关键词:肺炎支原体;P1黏附素;黏蛋白5AC;Dickkopf相关蛋白1

Abstract:

Mycoplasma pneumoniae is the most common pathogen of respiratory tract infection in children and adults. Clinical observation shows that M. pneumoniae infection can cause massive mucus secretion in the respiratory tract, which makes the breathing of patients difficult. Studies have shown that M. pneumoniae infection can cause massive secretion of mucin 5AC (MUC5AC). Adhesin P1 plays an important role in the pathogenesis of M. pneumoniae infection by mediating the adhesion of pathogens to host cells, and the C-terminal residues of P1 (P1-C) are immunogenic. This study investigated the molecular mechanism of Wnt/β-catenin signaling pathway inhibitor Dickkopf-1 (DKK1) in the secretion of MUC5AC in mouse airway epithelial cells (MAECs) induced by P1-C. Scanning electron microscope and hematoxylin-eosin staining were used to observe the effect of P1-C on mucus secretion of MAECs. Protein chip was used to detect the secretion of cytokines and analyse the enrichment of related signaling pathways induced by P1-C in MAECs. Periodic acid schiff stain (PAS) staining, Tunel staining and Masson staining were used to detect the damage of the lungs of mouse exposed to P1-C. Immunohistochemistry was used to detect the secretion of MUC5AC expression, and Western blotting was used to reveal the molecular mechanism of DKK1-regulated secretion of MUC5AC induced by P1-C protein in MACES. The results showed that P1-C induced the massive secretion of mucus and inflammatory factors in MAECs. During P1-C infection, DKK1 down-regulated janus kinase 2 (JAK2), phosphorylation signaling and transcription activator 1 (p-STAT1) and phosphorylation signaling and activator of transcription 3 (p-STAT3) expression. Overexpression of DKK1 significantly up-regulated the expression of MUC5AC repressor transcription factor fork-head box protein A2 (FOXA2). At the same time, the expression of MUC5AC induced by P1-C was inhibited significantly. It is speculated that DKK1 can effectively reduce the secretion of MUC5AC in MAECs induced by P1-C by inhibiting the JAK/STAT1-STAT3 signaling pathway and up-regulating the expression of FOXA2.

Key words:Mycoplasma pneumoniae;P1 adhesin;mucin 5AC (MUC5AC);Dickkopf-1