中文摘要：

目的研究聚肌胞苷酸[Poly（I：C）]、脂多糖（LPS）及肽聚糖（PGN）对人气道黏膜屏障及炎性介质表达的影响,进而了解气道黏膜对不同Toll样受体配体的天然免疫应答反应。方法采用Transwell系统培养人16HBEs气道上皮细胞建立人气道黏膜体外模型,分别给予Poly（I：C）、LPS及PGN顶侧刺激,对照组仅给予MEM＋GlutaMax-Ⅰ培养基培养。通过测定跨膜电阻抗（TER）判断16HBEs细胞间的小分子通透性,测定基底侧培养液中FITC-右旋糖酐浓度判断大分子细胞通透性,用ELISA法检测干预24h后培养细胞顶侧及基底侧上清液中IL-8、粒-巨噬细胞集落刺激因子（GM-CSF）及TNF-α的蛋白含量。结果 10μg/ml Poly（I：C）导致人气道上皮细胞间TER显著降低,FITC-右旋糖酐通透性增加（P〈0.01）,10μg/ml LPS及100μg/ml PGN刺激对气道上皮细胞TER及FITC-右旋糖酐通透性无明显影响。IL-8和TNF-α表达在Poly（I：C）、LPS及PGN组细胞顶侧和基底侧均较对照组显著增加（P〈0.05）。GM-CSF表达在3组细胞顶侧均较对照组增加（P〈0.05）,仅Poly（I：C）组基底侧表达较对照组增加（P〈0.05）,LPS和PGN组基底侧与对照组比较无明显变化。Poly（I：C）组、LPS组和PGN组细胞顶侧IL-8和GM-CSF浓度增高程度高于基底侧,形成浓度梯度。结论 Poly（I：C）可破坏气道黏膜屏障完整性,导致小分子和大分子通透性增加,同时刺激细胞定向向顶侧分泌炎性介质,LPS及PGN对气道黏膜屏障无影响,但能诱导细胞向顶侧定向分泌炎性介质,提示病毒及细菌感染所致气道炎症与Toll样受体通路介导的气道黏膜天然免疫应答有关。

英文摘要：

Objective To study the effect of polyinosinic：polycytidylic acid（poly（I：C））,lipolysaccharide（LPS）,peptidoglycan（PGN） on the barrier function and inflammatory mediators release of bronchial epithelium,and explore the impact of different specific Toll-like receptor（TLR 2,3,4） ligands on airway mucosal innate immune responses.Methods Polarized human bronchial epithelial cells（16HBEs） were cultured using transwell system to establish human bronchial epithelium model in vitro.The stimulation of Poly（I：C）,LPS and PGN,respectively,was given on 16HBEs apically.The permeabilities of micromolecule and macromolecule between cells were measured by detecting trans-epithelial electrical resistance（TER） and basolateral medium FITCdextran concentrations after 24h stimulation.IL-8,GM-CSF and TNF-α protein content of apical and basal supernatants released by 16HBEs were measured by ELISA after 24h stimulation.Results 10μg/ml Poly（I：C） resulted in decrease of TER and reduction of increase of FITC-dextran permeability significantly（P＆lt;0.001）,while 10μg/ml LPS and 100μg/ml PGN showed no obvious effects on TER and FITC-dextran permeability.Poly（I：C）,LPS and PGN induced an increase in apical and basolateral IL-8,TNF-α release and apical GM-CSF release of the cells.There was no change in basal expression of GM-CSF after being stimulated by LPS and PGN.The apical increase in IL-8 and GM-CSF was higher in Poly（I：C）,LPS and PGN groups than their basal increase,thus forming the concentration gradient between two sides of the polarized cells.Conclusions Poly（I：C） can damage the barrier integrity of the bronchial epithelium,leading to an increase in micromolecular and macromolecular permeability,and induce the secretion of inflammatory mediators towards the top side at the same time.LPS and PGN can only induce the secretion of inflammatory mediators towards the top side mildly,without affecting the barrier function.It is suggested that airway mucosal innate immune resp