Characterisation of the range of neutrophil stimulating mediators in cystic fibrosis sputum
Characterisation of the range of neutrophil stimulating mediators in cystic fibrosis sputum
复制标题
DOI：
10.1136/thx.2007.089359
复制
发表时间：
2008-07-01
期刊：
影响因子：
10
通讯作者：
Jose, P. J.
中科院分区：
文献类型：
--
作者：
Mackerness, K. J.;Jenkins, G. R.;Jose, P. J.
背景：大多数囊性纤维化（CF）患者因慢性感染和破坏性中性粒细胞炎症导致呼吸衰竭而死亡。目的：通过表征 CF 气道中的中性粒细胞刺激介质来确定潜在的治疗靶点。方法：在磷酸盐缓冲盐水中提取自发咳出的 CF 痰，用于分析中性粒细胞趋化性、细胞内钙动员和细胞形状变化。通过离子交换、C-18 反相和尺寸排阻色谱法纯化介体。结果：CF 痰池含有相当大的中性粒细胞刺激活性，但白细胞介素 (IL)8/CXCL8 的中和对中性粒细胞趋化性几乎没有抑制作用（10149 (2023) 迁移细胞与 62 mg 的 8661 (2597) 细胞相比 痰/ml； NS）或形状变化（19 mg 痰/ml 时前向散射百分比增加 46 (8) vs 38 (5)；p < 0.05）反应。此外，即使在中性粒细胞对 IL8 脱敏后，CF 痰池也会诱导细胞内钙离子升高。色谱法鉴定出 IL8、其他 CXC 趋化因子、白三烯 (LT) B-4 和两种甲酰基肽对中性粒细胞形状变化诱导活性的贡献。还有证据表明血小板激活因子 (PAF) 和 C5a 也有贡献。使用未经色谱分析的个体痰样本，单独的抗IL8确实对中性粒细胞趋化性具有抑制作用（中位抑制率为41％；p = 0.0002）。然而，即使在本实验中，CF 痰对中性粒细胞也存在明显重要的、非 IL8 介导的影响，抗 IL8 加 CXCR2、LTB4、甲酰基肽、PAF 和 C5a 受体拮抗剂的抑制剂混合物可抑制中位数 97% 的趋化性 (p= 0.0002)。 结论：许多趋化剂有助于中性粒细胞 尽管这些介质的相对贡献在不同患者中有所不同，但刺激 CF 痰液的活性。选择性阻断单一介质可能不足以控制 CF 气道中的中性粒细胞募集和激活。
Background: Most patients with cystic fibrosis (CF) die of respiratory failure due to chronic infection and destructive neutrophilic inflammation. Objective: To identify potential therapeutic targets by characterising the neutrophil stimulating mediators in the CF airway.Methods: Spontaneously expectorated CF sputum was extracted in phosphate buffered saline for assays of neutrophil chemotaxis, intracellular calcium mobilisation and cell shape change. Mediators were purified by ion exchange, C-18 reversed phase and size exclusion chromatography.Results: A pool of CF sputum contained considerable neutrophil stimulating activity but neutralisation of interleukin (IL)8/CXCL8 had little inhibitory effect on neutrophil chemotactic (10149 (2023) migrating cells vs 8661 (2597) at 62 mg sputum/ml; NS) or shape change (% forward scatter increase 46 (8) vs 38 (5) at 19 mg sputum/ml; p < 0.05) responses. Furthermore, the CF sputum pool induced an elevation in intracellular calcium ions even after desensitisation of the neutrophils to IL8. Chromatography identified contributions to the neutrophil shape change inducing activity from IL8, other CXC chemokines, leukotriene (LT) B-4 and two formyl peptides. There was also suggestive evidence for contributions from platelet activating factor (PAF) and C5a. Using non-chromatographed individual sputum samples, anti-IL8 alone did have an inhibitory effect on neutrophil chemotaxis (median inhibition 41%; p= 0.0002). However, even in this experiment, there were clearly significantly important, non-IL8 mediated, effects of CF sputum on neutrophils, and an inhibitor cocktail of anti-IL8 plus CXCR2, LTB4, formyl peptide, PAF and C5a receptor antagonists inhibited chemotaxis by a median of 97% (p= 0.0002).Conclusion: Many chemoattractants contribute to the neutrophil stimulating activity in CF sputum although the relative contribution of these mediators differs in different patients. Selective blockade of single mediators may not be sufficient to control neutrophil recruitment and activation in the CF airway.