Macrophage-derived exosomal miR-146b-5p exacerbates ILC2-mediated inflammation in allergic asthma by regulating the...

Type 2 innate lymphoid cells (ILC2) are key drivers of the type 2 inflammatory response. This study aims to investigate the mechanism by which macrophage-derived extracellular vesicles regulate ILC2 cells during the pathogenesis of allergic asthma, thereby providing novel evidence and directions for the prevention, diagnosis, and treatment of this condition.

An ovalbumin-induced allergic asthma mouse model was established. Macrophage-derived exosomes were isolated from bronchoalveolar lavage fluid (BALF) via ultracentrifugation, followed by miRNA sequencing analysis. Using Deseq2 software and RT-qPCR to screen and validate key miRNAs and their enriched pathways; In vivo experiments were conducted by tail vein injection of miRNA-146b-5p mimics into asthmatic mice. Subsequent assessments included Haematoxylin Eosin (H&E) staining, Masson's trichrome staining, TUNEL fluorescence, pulmonary function testing, RT-qPCR, Western blot analysis, and ELISA. These evaluated the effects of miRNA-146b-5p mimics on pulmonary inflammation, lung function, ILC2 cell proliferation, and the inflammatory cytokines produced (IL-4, IL-5, IL-13) and the TRAF-6/NF-κB pathway; In vitro experiments: Co-culturing macrophage exosomes and miRNA-146b-5p mimics with ILC2 cells, followed by ELISA detection of ILC2-associated inflammatory cytokines and Western blot analysis of relevant inflammatory pathways.

Compared with normal mice, 99 miRNAs were upregulated and 92 miRNAs were downregulated in macrophage exosomes from the alveolar lavage fluid of allergic asthma mice. Differentially expressed miRNAs were significantly enriched in pathways such as NF-κB. By intersecting the top 50 differentially expressed miRNAs with those targeting candidate pathways, yielding three candidate miRNAs: mmu-miR-181b-5p, mmu-miR-146b-5p, and mmu-miR-222-3p. Sequencing and RT-qPCR validation revealed that miR-146b-5p was significantly downregulated in the model group, with upregulation of TRAF-6 gene expression and NF-κB pathway activity. Intravenous tail vein injection of miRNA-146b-5p mimics into model group mice alleviated pulmonary inflammatory infiltration, improved lung function, and reduced levels of ILC2-associated inflammatory cytokines (IL-4, IL-5, IL-13, IL-25, IL-33, TSLP); Co-culture of macrophage exosomes with ILC2 cells demonstrated that exosomes from asthma mice promoted ILC2 cell proliferation. Co-incubation with miRNA-146b-5p mimics inhibited ILC2 cell proliferation and reduced NF-κB pathway expression.

Alveolar macrophage-derived extracellular vesicles containing miR-146b-5p promote ILC2 cell activation and inflammatory cytokine release by targeting the TRAF6/NF-κB signaling pathway, thereby exacerbating the pathological progression of allergic asthma. Exogenous supplementation with miR-146b-5p mimics effectively suppresses activation of the TRAF6/NF-κB pathway, thereby alleviating ILC2-mediated inflammatory responses. This provides novel potential targets and theoretical rationale for the prevention and treatment of allergic asthma.