Eucalyptol-Limonene-Pinene Attenuates Mucus Hypersecretion and Inflammation via Multitarget Mechanisms in Chronic...

Chronic rhinosinusitis with nasal polyps (CRSwNP) is a common chronic inflammatory disease of the mucosa, which significantly impairs patients' quality of life. Eucalyptol-Limonene-Pinene (ELP) is a plant-derived mucoactive agent used in respiratory diseases, including CRSwNP; however, its underlying mechanisms remain unclear. This study aimed to evaluate ELP's effect on CRSwNP and explore its mechanisms.

Predicted targets and pathways from network pharmacology were validated in vitro using nasal epithelial cells (NECs) and single-cell suspensions from CRSwNP patients, and in vivo using Aspergillus oryzae protease/ovalbumin-induced mouse CRSwNP model. Quantitative real-time polymerase chain reaction, enzyme-linked immunosorbent assay, Western blot, Luminex, hematoxylin and eosin staining, Alcian blue-periodic acid-Schiff staining, immunofluorescence, and immunohistochemistry were used to quantify inflammatory mediators and mucociliary changes.

Network pharmacology revealed that epidermal growth factor receptor (EGFR), hypoxia-inducible factor 1-alpha, interleukin (IL)-2 are core targets and enriched in inflammation pathways. In vitro, ELP inhibited the overexpression of mucin 5AC induced by amphiregulin, hypoxia, IL-2 and IL-13 in NECs. ELP decreased expression of type 2 inflammatory mediators, arachidonate 15-lipoxygenase, C-C motif chemokine ligand 26, periostin and epithelial-derived alarmins thymic stromal lymphopoietin, IL-33 in NECs, reduced the concentrations of cytokines/chemokines and countered lipopolysaccharide-induced inflammation in single-cell suspensions. In vivo, ELP reduced epithelial thickness, inflammatory infiltration, goblet cell hyperplasia and cilia damage.

ELP reduced mucus hypersecretion and inflammatory mediators in vitro and in vivo in CRSwNP by specifically inhibiting EGFR-, hypoxia-, IL-2- and IL-13-mediated pathways, suppressing inflammatory responses, and alleviating ciliary damage, suggesting its potential therapeutic value.