Slater Mariel, Torr Elizabeth, Harrison Tim, Forrester Doug, Knox Alan, Shaw Dominick, Sayers Ian
Division of Respiratory Medicine, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom.
Division of Respiratory Medicine, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom
Physiol Rep. 2016 Sep;4(18). doi: 10.14814/phy2.12960.
Macrolides including azithromycin (AZM) can improve clinical symptoms in asthma regardless of infection status. The mechanisms underlying these beneficial effects are yet to be elucidated. The aim of this study was to determine the effect of AZM on the airway epithelial barrier both in an in vitro model and in patients with asthma. Primary human bronchial epithelial cells (HBEC) were grown at air liquid interface (ALI) and challenged using lipopolysaccharides from Pseudomonas aeruginosa AZM was added at various stages and barrier integrity assessed using transepithelial electrical resistance (TEER) and permeability to FITC-dextran. MMP-9 levels were measured using ELISA AZM enhanced barrier integrity (TEER/FITC-dextran), increased thickness, suppressed mucin production, and MMP-9 release during the formation of a normal epithelial barrier in vitro. MMP-9 levels inversely correlated with TEER AZM also enhanced maintenance of the barrier and facilitated repair post-LPS challenge. To provide translation of our findings, 10 patients with moderate-severe asthma were recruited and received 250 mg AZM o.d for 6 weeks. Bronchial biopsies taken pre- and post-AZM treatment did not show evidence of increased epithelial barrier thickness or decreased mucin production. Similarly, bronchial wash samples did not show reduced MMP-9 levels. Overall, our data show that AZM can significantly improve the development of a normal bronchial epithelial barrier in vitro, mimicking reepithelization postinjury. AZM also suppressed MMP-9 release which correlated with barrier integrity, suggesting a putative mechanism. However, these effects were not observed in biopsy samples from asthma patients treated with AZM, possibly due to small sample size.
包括阿奇霉素（AZM）在内的大环内酯类药物无论感染状况如何，均可改善哮喘的临床症状。这些有益作用的潜在机制尚待阐明。本研究的目的是确定AZM在体外模型和哮喘患者中对气道上皮屏障的影响。原代人支气管上皮细胞（HBEC）在气液界面（ALI）培养，并用铜绿假单胞菌的脂多糖进行刺激。在不同阶段添加AZM，并使用跨上皮电阻（TEER）和对异硫氰酸荧光素标记葡聚糖（FITC-葡聚糖）的通透性评估屏障完整性。使用酶联免疫吸附测定法（ELISA）测量基质金属蛋白酶-9（MMP-9）水平。在体外正常上皮屏障形成过程中，AZM增强了屏障完整性（TEER/FITC-葡聚糖），增加了厚度，抑制了粘蛋白产生以及MMP-9释放。MMP-9水平与TEER呈负相关。AZM还增强了屏障的维持能力，并促进了脂多糖刺激后的修复。为了验证我们的研究结果，招募了10名中重度哮喘患者，给予每日一次250mg AZM，持续6周。在AZM治疗前后采集的支气管活检样本未显示上皮屏障厚度增加或粘蛋白产生减少的证据。同样，支气管灌洗样本也未显示MMP-9水平降低。总体而言，我们的数据表明，AZM可在体外显著改善正常支气管上皮屏障的形成，模拟损伤后的再上皮化过程。AZM还抑制了与屏障完整性相关的MMP-9释放，提示了一种可能的机制。然而，在接受AZM治疗的哮喘患者的活检样本中未观察到这些效应，可能是由于样本量较小。
