Exosomes derived from human umbilical cord mesenchymal stem cells alleviate otitis media in C57BL/6 mice via the miR-149-3p/NOX2 axis.

Otitis media (OM) is a highly prevalent disease leading to hearing loss in children. Current clinical treatment for OM primarily relies on antibiotics; however, the overuse of antibiotics exacerbates bacterial resistance, posing a global public health challenge. Therefore, novel therapeutic strategies for OM are urgently needed. Human umbilical cord mesenchymal stem cell-derived exosomes (hUCMSCs-Exos) have demonstrated significant therapeutic potential in various diseases, but their efficacy in OM remains unclear. In this study, we induced C57BL/6 mice with lipopolysaccharide (LPS) to establish an OM mouse model. OM mice were treated with exosomes derived from human umbilical cord mesenchymal stem cells (hUCMSCs-Exos), through two administration methods, tympanic membrane (TM) and tail vein (TV) injection. Our results showed that hUCMSCs-Exos significantly alleviated OM in mice, as evidenced by improved auditory function, reduced middle ear mucosal thickening, decreased purulent exudation, downregulated expression of inflammatory cytokines, and alleviated oxidative stress. Multi-omics analysis revealed significant alterations in NADPH Oxidase 2 (NOX2) expression and its upstream regulator miR-149-3p within the HIF-1 signaling pathway. qRT-PCR and Western blot confirmed that hUCMSCs-Exos upregulated miR-149-3p and suppressed NOX2 expression, thereby reducing reactive oxygen species (ROS) generation, and consequently alleviating otitis media inflammation. Furthermore, in vivo gain- and loss-of-function experiments demonstrated that miR-149-3p is a key mediator in hUCMSCs-Exos-regulated NOX2 expression and downstream inflammatory responses. Comparative analysis of administration routes revealed that local TM injection yielded superior therapeutic outcomes compared to systemic TV delivery. In summary, our findings indicate that hUCMSCs-Exos may alleviate LPS-induced OM in mice by delivering miR-149-3p to target and inhibit NOX2 expression and its downstream inflammatory cascade, providing a theoretical basis for hUCMSCs-Exos as a novel non-antibiotic therapeutic strategy for OM.