Mercury-containing preparations attenuate neutrophil extracellular trap formation in mice and humans through inhibiting the ERK1/2 pathway

Ethnopharmacological relevance: Neutrophil extracellular trap (NET) formation plays a crucial role in wound healing disorders, including chronic skin ulcers and diabetic foot ulcers (DFUs). Over the years, traditional Chinese topical medications, such as Cinnabar (composed of HgS and soluble mercury salt) and hydrargyria oxydum rubrum (containing HgO and soluble mercury salt), have been utilized for treating these ailments. Nevertheless, the fundamental processes remain mostly ambiguous.

Aim of the study: This study sought to investigate the potential effects of topical mercury-containing preparations on the process of NET formation.

Materials and methods: Neutrophils isolated from healthy individuals and mouse models of type 1 and type 2 diabetes were cultured with phorbol 12-myristate 13-acetate (PMA), both with and without the mercury-containing preparations (MCP). The formation of NETs was monitored using confocal and scanning electron microscopes. Immunofluorescence and fluorescent probes were employed to assess the levels of citrulline histone H₃ (Cit-H₃) and intracellular Reactive Oxygen Species (ROS), respectively. The impact of MCP extracts on cytokine expression, peptidylarginine deiminase 4 (PAD4), and myeloperoxidase (MPO) was measured through Luminex and ELISA assays. Phagocytosis of human neutrophils was analyzed using Flow Cytometry. Finally, the phosphorylation levels of ERK were detected by western blotting.

Results: Treatment with MCP led to a reduction in PAD4, Cit-H₃, and MPO expressions in neutrophils, consequently inhibiting PMA-induced NET formation. MCP treatment also dampened ERK1/2 activation in neutrophils. Furthermore, MCP exhibited inhibitory effects on the secretion of the cytokine IL-8 and ROS production while enhancing neutrophil phagocytosis.

Conclusion: Our findings suggest that MCP can mitigate the release of NETs, likely by suppressing the ERK1/2 signaling pathway.

Diabetic foot ulcer; ERK1/2 signaling pathway; Inflammation; Mercury-containing preparations; Neutrophil extracellular traps.