Department of Otolaryngology, Massachusetts Eye and Ear Infirmary, Harvard Medical School, Boston, Mass.
Department of Otolaryngology, Massachusetts Eye and Ear Infirmary, Harvard Medical School, Boston, Mass; Department of Otolaryngology/Head and Neck Surgery, University of Erlangen-Nuremberg, Erlangen, Germany.
J Allergy Clin Immunol. 2019 Apr;143(4):1525-1535.e1. doi: 10.1016/j.jaci.2018.08.046. Epub 2018 Nov 12.
Nasal mucosa-derived exosomes (NMDEs) harbor immunodefensive proteins and are capable of rapid interepithelial protein transfer.
We sought to determine whether mucosal exposure to inhaled pathogens stimulates a defensive swarm of microbiocidal exosomes, which also donate their antimicrobial cargo to adjacent epithelial cells.
We performed an institutional review board-approved study of healthy NMDE secretion after Toll-like receptor (TLR) 4 stimulation by LPS (12.5 μg/mL) in the presence of TLR4 inhibitors. Interepithelial transfer of exosomal nitric oxide (NO) synthase and nitric oxide was measured by using ELISAs and NO activity assays. Exosomal antimicrobial assays were performed with Pseudomonas aeruginosa. Proteomic analyses were performed by using SOMAscan.
In vivo and in vitro LPS exposure induced a 2-fold increase in NMDE secretion along with a 2-fold increase in exosomal inducible nitric oxide synthase expression and function through TLR4 and inhibitor of nuclear factor κB kinase activation. LPS stimulation increased exosomal microbiocidal activity against P aeruginosa by almost 2 orders of magnitude. LPS-stimulated exosomes induced a 4-fold increase in NO production within autologous epithelial cells with protein transfer within 5 minutes of contact. Pathway analysis of the NMDE proteome revealed 44 additional proteins associated with NO signaling and innate immune function.
We provide direct in vivo evidence for a novel exosome-mediated innate immunosurveillance and defense mechanism of the human upper airway. These findings have implications for lower airway innate immunity, delivery of airway therapeutics, and host microbiome regulation.
鼻腔黏膜衍生的外泌体（NMDE）携带免疫防御蛋白，能够快速进行上皮内蛋白转移。
我们旨在确定鼻腔黏膜接触吸入性病原体是否会刺激产生防御性的微生物杀伤性外泌体 swarm，这些外泌体还将其抗菌货物捐赠给相邻的上皮细胞。
我们进行了一项机构审查委员会批准的研究，在 TLR4 抑制剂存在的情况下，用 LPS（12.5μg/mL）刺激 TLR4 后，观察健康 NMDE 的分泌情况。通过 ELISA 和 NO 活性测定来测量外泌体一氧化氮（NO）合酶和 NO 的上皮内转移。使用铜绿假单胞菌进行外泌体抗菌测定。通过 SOMAscan 进行蛋白质组分析。
体内和体外 LPS 暴露诱导 NMDE 分泌增加 2 倍，同时通过 TLR4 和核因子κB 激酶抑制剂激活，诱导外泌体诱导型一氧化氮合酶表达和功能增加 2 倍。LPS 刺激使针对 P aeruginosa 的外泌体杀菌活性增加近 2 个数量级。LPS 刺激的外泌体在接触后 5 分钟内将蛋白转移到自体上皮细胞中，使 NO 产生增加 4 倍。NMDE 蛋白质组的通路分析显示，与 NO 信号和先天免疫功能相关的还有 44 种其他蛋白质。
我们提供了人类上呼吸道新型外泌体介导的先天免疫监视和防御机制的直接体内证据。这些发现对下呼吸道先天免疫、气道治疗药物的输送以及宿主微生物组调节具有重要意义。
