Cellular Detachment and Deformation induce Cytokine Gene Expression in Human Bronchial Epithelial Cells

神经弹性蛋白酶（NE）是一种能使人支气管上皮细胞（BECs）变形和脱落并诱导白细胞介素8（IL-8）基因表达的蛋白酶。然而，NE诱导的IL-8基因表达的机制尚不清楚，特别是关于它们如何与细胞脱离。为了阐明这些机制，通过北方分析检测机械损伤或药理学刺激后细胞脱离和变形对IL-8基因表达的影响。当来自人支气管上皮细胞系的BET-1A细胞与NE（100 nM）、胰蛋白酶（0.5mg/ml）、EGTA（7 mM）或EDTA（0.7mM）孵育以诱导变形和脱离时，IL-8 mRNA转录物水平上调，如使用细胞刮刀从培养板机械脱离的情况所示。这些IL-8基因的表达被抑制预处理与5 mM紫杉醇，微管稳定剂。秋水仙碱或长春碱，微管破坏剂，诱导IL-8基因的表达，这也被抑制紫杉醇治疗。这些数据表明，结构的变化，包括变形的细胞骨架，特别是微管，可能有助于IL-8基因在人类BEC的表达。由于蛋白酶引起的BEC的分离和细胞变形在各种炎症性气道疾病中经常被观察到，我们的研究结果提供了证据表明，分离或变形的BEC可能会增强气道炎症发病机制中炎症介质的产生。

Neutrophil elastase (NE) is known to be one of the most potent proteases capable of deforming and detaching human bronchial epithelial cells (BECs) and inducing interleukin-8 (IL-8) gene expression. However, mechanisms of NE-induced IL-8 gene expression are unclear, especially with respect to how they relate to cellular detachment. In order to elucidate these mechanisms, effects of cell detachment and deformation following mechanical injury or pharmacological stimuli on IL-8 gene expression were examined by Northern analyzes. When BET-1A cells from a human bronchial epithelial cell line were incubated with NE (100nM), trypsin (0.5mg/ml), EGTA (7mM) or EDTA (0.7mM) to induce deformation and detachment, IL-8 mRNA transcript levels were upregulated as demonstrated in a case of mechanical detachment from the culture plate using a cell scraper. These IL-8 gene expression was inhibited by pretreatment with 5mM taxol, a microtubule stabilizing agent. Colchicine or vinblastine, microtubule disrupting agents, induced IL-8 gene expression, which was also inhibited by taxol treatment. These data suggest that structural changes, including deformation of the cytoskeleton, especially microtubules, may contribute to IL-8 gene expression in human BECs. Since detachment and cellular deformation of BECs caused by proteases have been frequently observed in a variety of inflammatory airway diseases, our findings provide evidence that detached or deformed BECs potentially enhance production of inflammatory mediators in the pathogenesis of airway inflammation.