Ramanathan Murray, Lee Won-Kyung, Dubin Marc G, Lin Sandra, Spannhake Ernst W, Lane Andrew P
Department of Otolaryngology-Head and Neck Surgery, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
Am J Rhinol. 2007 Jan-Feb;21(1):110-6. doi: 10.2500/ajr.2007.21.2997.
Innate immune recognition of pathogens by sinonasal epithelial cells may play an important role in the pathogenesis of chronic rhinosinusitis (CRS). Previous studies have indicated that toll-like receptor (TLR) mRNA is present in sinonasal mucosa, and levels of TLR9 expression are decreased in recalcitrant CRS with nasal polyps (CRSwNP). However, the cellular source and function of TLR9 in the sinonasal epithelium is not known. In this study, primary epithelial cell cultures were analyzed from control subjects and CRSwNP patients to determine the presence and function of TLR9 protein.
Primary epithelial cell cultures were established from 5 controls and 10 CRSwNP patients undergoing sinus surgery. Flow cytometry was used to confirm purity of epithelial cells and to assess expression of TLR9 protein. Epithelial cells were stimulated with TLR9 agonist, and mRNA was analyzed by real-time PCR for expression of human beta-defensin (HBD) 2 and interleukin (IL)-8.
Flow cytometry showed TLR9 protein in 100% of epithelial cells from controls and CRSwNP patients. The level of expression was 50% lower in CRS patients than in controls. Stimulation of epithelial cells with TLR9 agonist produced a 1.5- to 9-fold increase in HBD-2 and IL-8 mRNA expression.
Functional TLR9 protein is expressed by normal and diseased sinonasal epithelial cells. The level of TLR9 expression is decreased in CRSwNP patients, consistent with the previous finding of decreased TLR9 mRNA in whole sinonasal tissue. These findings suggest that impaired innate immune responses to pathogens via TLR9 on sinonasal epithelial cells may represent a critical mechanism in chronic inflammatory sinus disease.
鼻窦上皮细胞对病原体的天然免疫识别可能在慢性鼻窦炎（CRS）的发病机制中起重要作用。先前的研究表明，鼻窦黏膜中存在Toll样受体（TLR）mRNA，并且在伴有鼻息肉的顽固性CRS（CRSwNP）中TLR9表达水平降低。然而，鼻窦上皮中TLR9的细胞来源和功能尚不清楚。在本研究中，对来自对照受试者和CRSwNP患者的原代上皮细胞培养物进行分析，以确定TLR9蛋白的存在和功能。
从5名对照者和10名接受鼻窦手术的CRSwNP患者中建立原代上皮细胞培养物。采用流式细胞术确认上皮细胞的纯度并评估TLR9蛋白的表达。用TLR9激动剂刺激上皮细胞，并通过实时PCR分析mRNA，以检测人β-防御素（HBD）2和白细胞介素（IL）-8的表达。
流式细胞术显示，对照者和CRSwNP患者的100%上皮细胞中存在TLR9蛋白。CRS患者的表达水平比对照者低50%。用TLR9激动剂刺激上皮细胞后，HBD-2和IL-8 mRNA表达增加了1.5至9倍。
正常和患病的鼻窦上皮细胞均表达功能性TLR9蛋白。CRSwNP患者中TLR9表达水平降低，这与先前在整个鼻窦组织中TLR9 mRNA降低的发现一致。这些发现表明，鼻窦上皮细胞上通过TLR9对病原体的天然免疫反应受损可能是慢性炎症性鼻窦疾病的关键机制。
