Zong Fuliang, Xiao Nan, Wang Yifeng, Su Duo, Zhou Dongsheng, Hu Lingfei, Yang Huiying
State Key Laboratory of Pathogen and Biosecurity, Academy of Military Medical Sciences, Beijing, China.
Department of Clinical Laboratory Medicine, Hebei Medical University, Shijiazhuang, Hebei, China.
Virulence. 2025 Dec;16(1):2490206. doi: 10.1080/21505594.2025.2490206. Epub 2025 Apr 11.
(PA) is a key pathogen in hospital-acquired pneumonia (HAP) and ventilator-associated pneumonia (VAP), challenging clinical medicine. This study aims to elucidate the characteristics of the host's innate immune response following inhalational PA infection. We developed a mouse model by aerosolized intratracheal inoculation with PA and conducted a comprehensive analysis at the protein, cellular, and gene expression levels. Protein analysis revealed a substantial increase in inflammatory proteins in the bronchoalveolar lavage fluid and serum, indicating a robust inflammatory response in the lungs and systemic circulation. Cellular investigations showed an increase in neutrophils, monocytes, and alveolar macrophages during infection, whereas NK cells showed a marked reduction from 5.88% pre-infection to 2.41% at 24 h ( = 0.0102) and 1.55% by 48 h ( = 0.0023). To assess gene expression changes, RNA-sequencing technology was employed to map the temporal shifts in the transcriptional profile of the host lung post-infection. We analysed the expression patterns and dynamic transcriptional characteristics of differentially expressed genes (DEGs), describing the inflammation progression. Importantly, Through the analysis of single-cell RNA sequencing (scRNA-seq) datasets in public repositories, we observed the reduction in conventional natural killer (cNK) cells, rather than tissue-resident natural killer (trNK) cells in the early stages of PA infection. Sequential scRNA-seq analysis resolved NK-subset heterogeneity, revealing that cNK dominance (77.8% of total NK cells) under homeostasis collapsed to 9.2% within 24 h post-infection. Our findings establish cNK attrition as the earliest immune checkpoint failure in PA pneumonia and provide proof-of-concept for cNK-targeted immunotherapies to counteract lethal pulmonary infections. Keywords: , aerosolized intratracheal inoculation, conventional NK cells, innate immunity, RNA-sequencing.
铜绿假单胞菌（PA）是医院获得性肺炎（HAP）和呼吸机相关性肺炎（VAP）的关键病原体，给临床医学带来挑战。本研究旨在阐明吸入性PA感染后宿主固有免疫反应的特征。我们通过气管内雾化接种PA建立了小鼠模型，并在蛋白质、细胞和基因表达水平上进行了全面分析。蛋白质分析显示支气管肺泡灌洗液和血清中的炎症蛋白大幅增加，表明肺部和全身循环中存在强烈的炎症反应。细胞研究表明，感染期间中性粒细胞、单核细胞和肺泡巨噬细胞数量增加，而自然杀伤（NK）细胞则显著减少，从感染前的5.88%降至24小时时的2.41%（P = 0.0102），到48小时时降至1.55%（P = 0.0023）。为了评估基因表达变化，采用RNA测序技术绘制感染后宿主肺转录谱的时间变化。我们分析了差异表达基因（DEG）的表达模式和动态转录特征，描述炎症进展。重要的是，通过分析公共数据库中的单细胞RNA测序（scRNA-seq）数据集，我们观察到在PA感染早期，常规自然杀伤（cNK）细胞减少，而非组织驻留自然杀伤（trNK）细胞减少。连续的scRNA-seq分析解决了NK亚群的异质性，揭示了稳态下cNK占主导地位（占总NK细胞的77.8%）在感染后24小时内降至9.2%。我们的研究结果确定cNK耗损是PA肺炎中最早出现的免疫检查点失败，并为针对cNK的免疫疗法对抗致命性肺部感染提供了概念验证。关键词： ，气管内雾化接种，常规NK细胞，固有免疫，RNA测序
