Pizzichini E, Pizzichini M M, Efthimiadis A, Evans S, Morris M M, Squillace D, Gleich G J, Dolovich J, Hargreave F E
Department of Medicine, St. Joseph's Hospital, Hamilton, Ontario, Canada.
Am J Respir Crit Care Med. 1996 Aug;154(2 Pt 1):308-17. doi: 10.1164/ajrccm.154.2.8756799.
Methods to examine sputum for indices of airway inflammation are evolving. We have examined the repeatability and the validity of an improved method to measure sputum cells and fluid-phase eosinophil cationic protein (ECP), major basic protein (MBP), eosinophil-derived neurotoxin (EDN), albumin, fibrinogen, tryptase, and interleukin-5 (IL-5). Sputum was induced with hypertonic saline twice within 6 d in 10 healthy subjects, 19 stable asthmatics, and 10 smokers with nonobstructive bronchitis. The method included the processing of freshly expectorated sputum separated from saliva, treatment with a fixed proportion of dithiothreitol 0.1% followed by Dulbecco's phosphate-buffered saline, making cytospins, and collecting the supernatant. The reproducibility of measurements, calculated by the intraclass correlation coefficient, was high for all indices measured with the exception of total cell counts and proportion of lymphocytes. Asthmatics, in comparison with healthy subjects and smokers with bronchitis, had a higher proportion of sputum eosinophils (median percent 5.2 versus 0.5 and 0.3), metachromatic cells (0.3 versus 0.07 and 0.08), ECP (1,040 micrograms/L versus 288 and 352), MBP (1,176 micrograms/L versus 304 and 160), and EDN (1,512 micrograms/L versus 448 and 272). Asthmatics differed from healthy subjects, but not from smokers with bronchitis, in the proportion of neutrophils (46.9% versus 24.1%), albumin (704 versus 288 micrograms/mL), and fibrinogen (2,080 versus 440 ng/mL). Smokers with bronchitis showed a trend for a higher neutrophil count and levels of albumin and fibrinogen than healthy subjects. The proportion of sputum eosinophils correlated positively with ECP, MBP, EDN, albumin and fibrinogen levels, and metachromatic cell counts correlated with tryptase. In asthmatics, IL-5 correlated with eosinophil counts. There was a significant negative correlation between sputum indices and expiratory flows and methacholine PC20. Thus, the methods of measuring cell and fluid phase markers in induced sputum used in this study are reproducible and valid. They can therefore be used to reliably measure these indices of airway inflammation.
用于检查痰液以获取气道炎症指标的方法正在不断发展。我们已经检验了一种用于测量痰液细胞以及液相嗜酸性粒细胞阳离子蛋白（ECP）、主要碱性蛋白（MBP）、嗜酸性粒细胞衍生神经毒素（EDN）、白蛋白、纤维蛋白原、类胰蛋白酶和白细胞介素-5（IL-5）的改良方法的可重复性和有效性。在10名健康受试者、19名稳定期哮喘患者和10名患有非阻塞性支气管炎的吸烟者中，于6天内用高渗盐水诱导痰液两次。该方法包括处理从唾液中分离出的新鲜咳出的痰液，先用固定比例的0.1%二硫苏糖醇处理，然后用杜氏磷酸盐缓冲盐水处理，制作细胞涂片，并收集上清液。通过组内相关系数计算得出的测量重复性，除总细胞计数和淋巴细胞比例外，所有测量指标的重复性都很高。与健康受试者和患有支气管炎的吸烟者相比，哮喘患者痰液中的嗜酸性粒细胞比例更高（中位数百分比分别为5.2%、0.5%和0.3%）、异染细胞比例更高（0.3%、0.07%和0.08%）、ECP水平更高（1040微克/升、288微克/升和352微克/升）、MBP水平更高（1176微克/升、304微克/升和160微克/升）以及EDN水平更高（1512微克/升、448微克/升和272微克/升）。哮喘患者与健康受试者相比，但与患有支气管炎的吸烟者相比，中性粒细胞比例不同（46.9%对24.1%）、白蛋白水平不同（704微克/毫升对288微克/毫升）以及纤维蛋白原水平不同（2080纳克/毫升对440纳克/毫升）。患有支气管炎的吸烟者中性粒细胞计数以及白蛋白和纤维蛋白原水平有高于健康受试者的趋势。痰液嗜酸性粒细胞比例与ECP、MBP、EDN、白蛋白和纤维蛋白原水平呈正相关，异染细胞计数与类胰蛋白酶相关。在哮喘患者中，IL-5与嗜酸性粒细胞计数相关。痰液指标与呼气流量和乙酰甲胆碱PC20之间存在显著负相关。因此，本研究中用于测量诱导痰液中细胞和液相标志物的方法具有可重复性和有效性。所以它们可用于可靠地测量这些气道炎症指标。
