NOX2-Dependent Phagocyte Signaling Mediates Silica-Induced Murine Lung Injury via Macrophage-Neutrophil Interactions.

In phagocytes, reactive oxygen species produced by NADPH oxidase 2 (NOX2) play a crucial role in the digestion of phagocytosed material as well as in immune response signaling. We recently reported the role of NOX2 in regulating inflammatory signaling in macrophages following silica nanoparticles (SiNPs) phagocytosis in vitro. However, it remains unclear whether pharmacological inhibition of NOX2 signaling modulates the SiNPs-induced lung injury. Here, we show that local administration of a NOX2-specific inhibitory peptide effectively suppressed neutrophilic lung injury caused by airway exposure to SiNPs. Formation of neutrophil extracellular traps (NETs) by neutrophils recruited to the alveoli was evident in SiNPs-induced lung injury tissues, as assessed by histological evidence of activated neutrophil elastase and myeloperoxidase as well as an increased DNA-histone concentration in bronchoalveolar lavage. Intratracheal administration of gp91ds-tat, a synthetic inhibitory peptide with specific activity against NOX2, significantly reduced the production of neutrophil chemoattractants in SiNPs-induced lung injury. Furthermore, NOX2 inhibition also suppressed NET formation upon silica-induced lung injury in vivo. We also confirmed that an NOX2 inhibitory peptide prevented NETs formation in neutrophils stimulated with silica nanoparticles in vitro. We also observed a potential role of silica-induced NETs in macrophage priming in vitro. Collectively, our findings suggest that silica-induced lung injury is exacerbated through a positive feedback loop, in which macrophages and neutrophils amplify inflammation via the production of chemoattractants and formation of NETs, respectively. The airway administration of NOX2 inhibitors effectively disrupts this feedback loop formed by two types of phagocytes, thereby mitigating silica-induced lung injury. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10753-025-02389-z.