Intracellular co-localization of SPLUNC1 protein with nanobacteria in nasopharyngeal carcinoma epithelia HNE1 cells depended on the bactericidal permeability increasing protein domain
Intracellular co-localization of SPLUNC1 protein with nanobacteria in nasopharyngeal carcinoma epithelia HNE1 cells depended on the bactericidal permeability increasing protein domain
复制标题
DOI：
10.1016/j.molimm.2005.10.021
复制
发表时间：
2006-04-01
影响因子：
3.6
通讯作者：
Zhao, J
中科院分区：
文献类型：
--
作者：
Zhou, HD;Li, GY;Zhao, J
上皮细胞表面构成天然的免疫载体，抵御环境威胁。这些屏障充满了结合，运输，切割或降解细菌细胞及其内毒素副产物的液体。唾液和气道表面衬液（ASL）是重要的体液成分。短腭、肺和鼻上皮克隆1（SPLUNC 1）是一种潜在的宿主防御蛋白，其从粘膜下腺体分泌到唾液和鼻灌洗液中。但其抗菌谱和抗菌机制尚不清楚。通过绿色荧光蛋白（GFP）介导的鼻咽癌（NPC）HNE 1细胞系的亚细胞定位实验，我们确定了细胞内GFP标记的SPLUNC 1蛋白在杀菌通透性增加蛋白（BPI）结构域被删除后，与大小约为50-400 nm的微小微生物结合，GFP标记的截短SPLUNC 1蛋白失去了与微小微生物结合的功能。我们证实，这些微生物是纳米细菌（NB）与负染色使用透射电子显微镜（TEM）和免疫荧光分析使用NB特异性抗体。从培养的鼻咽癌上皮细胞HNE 1上清中分离培养NB。我们发现，NB不吸收Hoechst染色，即使当我们延长染色时间到35分钟。然而，随着时间的延长，较大尺寸的NB（大于300 nm）染色阳性。从鼻咽癌活检标本中，我们还检测到了NB，它可以导致感染宿主细胞中的线粒体肿胀。我们推测SPLUNC 1和NB共定位是由于GFP标记的SPLUNC 1蛋白与革兰氏阴性NB的脂多糖（LPS）结合，这在鼻咽上皮的宿主防御中起重要作用。这项研究为SPLUNC 1参与宿主上呼吸道防御系统的机制提供了新的线索。(c)2005 Elsevier Ltd.保留所有权利。
Epithelial surfaces constitute natural immunobarriers against environmental threats. These barriers are brimming with fluids that bind, transport, cleave or degrade bacterial cells and their endotoxic by-products. Saliva and the airway surface-lining fluid (ASL) comprise the important fluid constituents. Short palate, lung and nasal epithelium clone 1 (SPLUNC1) is a potential host defensive protein that is secreted from the submucosal gland to the saliva and nasal lavage fluid. However, its antimicrobial spectrum and antimicrobial mechanism is not clear. Through green fluorescence protein (GFP) mediated subcellular localization experiments in nasopharyngeal carcinoma (NPC) HNE1 cell line, we determined that the intracellular GFP-tagged SPLUNC1 protein binds to a miniscule microorganisms, approximately 50-400 nm in size, after the bactericidal permeability increasing protein (BPI) domain was deleted, GFP-tagged truncated SPLUNC1 protein lost its function of binding to the miniscule microorganisms. We verified that these microorganisms are nanobacteria (NB) with a negative staining using transmitted electronic microscope (TEM) and immunofluorescent analysis using an NB-specific antibody. We isolated and cultured the NB from the cultured nasopharyngeal carcinoma epithelia HNE1 cell supernatant. We found that the NB did not absorb the Hoechst stain, even when we extended the staining time to 35 min. However, with the time extension the larger sized NB (larger than 300 nm) did stain positively. From the biopsy specimen of NPC, we also detected the NB, which can lead to the swelling of mitochondria in the infected host cells. We hypothesize that SPLUNC1 and NB co-localization is due to the GFP-tagged SPLUNC1 protein binding to the lipopolysaccharide (LPS) of the Gram-negative NB, which can play an important role in the host defense of nasopharyngeal epithelium. This research sheds new light on the mechanism of SPLUNC1 involvement in the host upper respiratory tract defense system. (c) 2005 Elsevier Ltd. All rights reserved.