背景:气道上皮充当物理屏障,保护肺气道免受病原微生物和有毒物质的侵害,如香烟烟雾(CS),细菌,和病毒。气道上皮结构完整性的破坏和功能障碍与慢性阻塞性肺疾病的发生和进展有关。
目的:本研究的目的是比较CS,肺炎克雷伯菌(KP),以及它们的组合对气道上皮屏障功能的影响。
方法:将小鼠暴露于CS,KP,和他们的组合从1到8周。在第8周停止CS和KP后,我们观察到小鼠的上皮屏障功能再恢复16周。为了比较不同组的上皮屏障功能随时间的变化,在第4、8、16和24周处死小鼠,然后收获肺部以检测肺部病理,炎性细胞因子,和紧密连接蛋白。为了确定潜在的机制,BEAS-2B细胞用表皮生长因子受体(EGFR)抑制剂(AG1478)处理.
结果:这项研究的结果表明,肺功能下降,增加支气管壁厚度(BWT),炎症因子升高,在CS诱导的小鼠中,在第8周观察到紧密连接蛋白水平降低,并且这些变化持续到第16周。在金伯利集团,仅在第8周观察到BWT增加和炎症因子升高,而在CS+KP组中,肺功能下降,肺组织损伤,炎性细胞浸润,在第4周观察到上皮屏障损伤,并持续到第24周。为了进一步确定CS的机制,细菌,以及它们对上皮屏障损伤的联合作用,我们研究了EGFR及其下游蛋白在小鼠肺组织和BEAS-2B细胞中的变化。我们的研究表明,CS,KP,或者它们的组合可以激活EGFR,可以磷酸化和激活ERK1/2,并且这种作用在CSKP组中更为明显。此外,EGFR抑制剂AG1478抑制了ERK1/2的磷酸化,随后上调了ZO-1和occludin的表达.总的来说,这些结果表明,CS和KP的组合对上皮屏障功能的损害比单独CS或KP更严重和持久,这可能与EGFR/ERK1/2信号传导有关。
结论:上皮屏障损伤发生较早,更严重,与单独暴露于CS或KP相比,CS和KP联合诱导的持续时间更长,这可能与EGFR/ERK信号有关。
BACKGROUND: The airway epithelium acts as a physical barrier to protect pulmonary airways against pathogenic microorganisms and toxic substances, such as cigarette smoke (CS), bacteria, and viruses. The disruption of the structural integrity and dysfunction of the airway epithelium is related to the occurrence and progression of chronic obstructive pulmonary disease.
OBJECTIVE: The aim of this
study is to compare the effects of CS, Klebsiella pneumoniae (KP), and their combination on airway epithelial barrier function.
METHODS: The mice were exposed to CS, KP, and their combination from 1 to 8 weeks. After the cessation of CS and KP at Week 8, we observed the recovery of epithelial barrier function in mice for an additional 16 weeks. To compare the epithelial barrier function among different groups over time, the mice were sacrificed at Weeks 4, 8, 16, and 24 and then the lungs were harvested to detect the pulmonary pathology, inflammatory cytokines, and tight junction proteins. To determine the underlying mechanisms, the BEAS-2B cells were treated with an epidermal growth factor receptor (EGFR) inhibitor (AG1478).
RESULTS: The results of this
study suggested that the decreased lung function, increased bronchial wall thickness (BWT), elevated inflammatory factors, and reduced tight junction protein levels were observed at Week 8 in CS-induced mice and these changes persisted until Week 16. In the KP group, increased BWT and elevated inflammatory factors were observed only at Week 8, whereas in the CS + KP group, decreased lung function, lung tissue injury, inflammatory cell infiltration, and epithelial barrier impairment were observed at Week 4 and persisted until Week 24. To further determine the mechanisms of CS, bacteria, and their combination on epithelial barrier injury, we investigated the changes of EGFR and its downstream protein in the lung tissues of mice and BEAS-2B cells. Our research indicated that CS, KP, or their combination could activate EGFR, which can phosphorylate and activate ERK1/2, and this effect was more pronounced in the CS + KP group. Furthermore, the EGFR inhibitor AG1478 suppressed the phosphorylation of ERK1/2 and subsequently upregulated the expression of ZO-1 and occludin. In general, these results indicated that the combination of CS and KP caused more severe and enduring damage to epithelial barrier function than CS or KP alone, which might be associated with EGFR/ERK1/2 signaling.
CONCLUSIONS: Epithelial barrier injury occurred earlier, was more severe, and had a longer duration when induced by the combination of CS and KP compared with the exposure to CS or KP alone, which might be associated with EGFR/ERK signaling.