Rebuli Meghan E, Speen Adam M, Clapp Phillip W, Jaspers Ilona
Curriculum in Toxicology, School of Medicine, University of North Carolina, Chapel Hill, North Carolina.
Curriculum in Toxicology, School of Medicine, University of North Carolina, Chapel Hill, North Carolina;
Am J Physiol Lung Cell Mol Physiol. 2017 Feb 1;312(2):L288-L296. doi: 10.1152/ajplung.00476.2016. Epub 2016 Dec 23.
Reliable methods for sampling the nasal mucosa provide clinical researchers with key information regarding respiratory biomarkers of exposure and disease. For quick and noninvasive sampling of the nasal mucosa, nasal lavage (NL) collection has been widely used as a clinical tool; however, limitations including volume variability, sample dilution, and storage prevent NL collection from being used in nonlaboratory settings and analysis of low abundance biomarkers. In this study, we optimize and validate a novel methodology using absorbent Leukosorb paper cut to fit the nasal passage to extract epithelial lining fluid (ELF) from the nasal mucosa. The ELF sampling method limits the dilution of soluble mediators, allowing quantification of both high- and low-abundance soluble biomarkers such as IL-1β, IL-8, IL-6, interferon gamma-induced protein 10 (IP-10), and neutrophil elastase. Additionally, we demonstrate that this method can successfully detect the presence of respiratory pathogens such as influenza virus and markers of antibiotic-resistant bacteria in the nasal mucosa. Efficacy of ELF collection by this method is not diminished in consecutive-day sampling, and percent recovery of both recombinant IL-8 and soluble mediators are not changed despite freezing or room temperature storage for 24 h. Our results indicate that ELF collection using Leukosorb paper sampling of ELF provides a sensitive, easy-to-use, and reproducible methodology to collect concentrated amounts of soluble biomarkers from the nasal mucosa. Moreover, the methodology described herein improves upon the standard NL collection method and provides researchers with a novel tool to assess changes in nasal mucosal host defense status.
可靠的鼻黏膜采样方法为临床研究人员提供了有关暴露和疾病的呼吸生物标志物的关键信息。为了快速、无创地采集鼻黏膜样本,鼻腔灌洗(NL)采集已被广泛用作一种临床工具;然而,包括体积变异性、样本稀释和储存等局限性使得NL采集无法用于非实验室环境以及低丰度生物标志物的分析。在本研究中,我们优化并验证了一种新方法,即使用裁剪成适合鼻腔通道大小的吸收性白细胞吸附纸从鼻黏膜中提取上皮衬液(ELF)。ELF采样方法限制了可溶性介质的稀释,能够对高丰度和低丰度的可溶性生物标志物进行定量,如白细胞介素-1β(IL-1β)、白细胞介素-8(IL-8)、白细胞介素-6(IL-6)、干扰素γ诱导蛋白10(IP-10)和中性粒细胞弹性蛋白酶。此外,我们证明该方法能够成功检测鼻黏膜中流感病毒等呼吸道病原体以及抗生素耐药菌的标志物的存在。连续多天采样时,该方法采集ELF的效果不会降低,重组IL-8和可溶性介质的回收率在冷冻或室温储存24小时后均未改变。我们的结果表明,使用白细胞吸附纸采样ELF能够提供一种灵敏、易用且可重复的方法,从鼻黏膜中采集浓缩的可溶性生物标志物。此外,本文所述方法改进了标准的NL采集方法,为研究人员提供了一种评估鼻黏膜宿主防御状态变化的新工具。