Fraunhofer Institute for Toxicology and Experimental Medicine, Hannover, Germany.
Member of Fraunhofer International Consortium for Anti-Infective Research (iCAIR), Member of Fraunhofer CIMD, Hannover, Germany.
Respir Res. 2024 Jan 4;25(1):15. doi: 10.1186/s12931-023-02609-w.
Aging is associated with an increased incidence and mortality of Pseudomonas aeruginosa-induced pneumonias. This might be partly due to age-dependent increases in inflammatory mediators, referred to as inflamm-aging and a decline in immune functions, known as immunosenescence. Still, the impact of dysregulated immune responses on lung infection during aging is poorly understood. Here, we aimed to mimic inflamm-aging using ex vivo precision-cut lung slices (PCLS) and neutrophils - as important effector cells of innate immunity - from young and old mice and investigated the influence of aging on inflammation upon infection with P. aeruginosa bacteria.
Murine PCLS were infected with the P. aeruginosa standard lab strain PAO1 and a clinical P. aeruginosa isolate D61. After infection, whole-transcriptome analysis of the tissue as well as cytokine expression in supernatants and tissue lysates were performed. Responses of isolated neutrophils towards the bacteria were investigated by quantifying neutrophil extracellular trap (NET) formation, cytokine secretion, and analyzing expression of surface activation markers using flow cytometry.
Inflamm-aging was observed by transcriptome analysis, showing an enrichment of biological processes related to inflammation, innate immune response, and chemotaxis in uninfected PCLS of old compared with young mice. Upon P. aeruginosa infection, the age-dependent pro-inflammatory response was even further promoted as shown by increased production of cytokines and chemokines such as IL-1β, IL-6, CXCL1, TNF-α, and IL-17A. In neutrophil cultures, aging did not influence NET formation or cytokine secretion during P. aeruginosa infection. However, expression of receptors associated with inflammatory responses such as complement, adhesion, phagocytosis, and degranulation was lower in neutrophils stimulated with bacteria from old mice as compared to young animals.
By using PCLS and neutrophils from young and old mice as immunocompetent ex vivo test systems, we could mimic dysregulated immune responses upon aging on levels of gene expression, cytokine production, and receptor expression. The results furthermore reflect the exacerbation of inflammation upon P. aeruginosa lung infection as a result of inflamm-aging in old age.
衰老与铜绿假单胞菌引起的肺炎发病率和死亡率增加有关。这可能部分归因于炎症介质的年龄依赖性增加，称为炎症衰老，以及免疫功能的下降，称为免疫衰老。尽管如此，免疫反应失调对衰老过程中肺部感染的影响仍知之甚少。在这里，我们旨在使用来自年轻和老年小鼠的体外精密切割肺切片（PCLS）和中性粒细胞 - 作为先天免疫的重要效应细胞 - 来模拟炎症衰老，并研究衰老对铜绿假单胞菌感染时炎症的影响。
用铜绿假单胞菌标准实验室菌株 PAO1 和临床铜绿假单胞菌分离株 D61 感染小鼠 PCLS。感染后，对组织的全转录组分析以及上清液和组织裂解物中的细胞因子表达进行分析。通过定量中性粒细胞细胞外陷阱（NET）形成、细胞因子分泌以及使用流式细胞术分析表面激活标记物的表达，研究分离的中性粒细胞对细菌的反应。
通过转录组分析观察到炎症衰老，与年轻小鼠相比，老年小鼠未感染的 PCLS 中与炎症、先天免疫反应和趋化作用相关的生物学过程富集。在铜绿假单胞菌感染后，年龄依赖性促炎反应进一步增强，表现为细胞因子和趋化因子（如 IL-1β、IL-6、CXCL1、TNF-α 和 IL-17A）的产生增加。在中性粒细胞培养物中，衰老不会影响铜绿假单胞菌感染期间 NET 的形成或细胞因子的分泌。然而，与用来自年轻动物的细菌刺激的中性粒细胞相比，来自老年动物的细菌刺激的中性粒细胞中与炎症反应相关的受体（如补体、粘附、吞噬和脱颗粒）的表达较低。
通过使用来自年轻和老年小鼠的 PCLS 和中性粒细胞作为免疫功能健全的体外测试系统，我们可以模拟衰老时基因表达、细胞因子产生和受体表达的失调免疫反应。结果进一步反映了炎症衰老导致的老年铜绿假单胞菌肺部感染的炎症加剧。
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