Sinomenine Regulates the TRIM32/IRF1/TRAF6 Axis to Inhibit Pyroptosis in Atopic Dermatitis.

Atopic dermatitis (AD) is a persistent skin disorder involving inflammation and marked by immune dysregulation. Sinomenine, a plant-derived alkaloid with known anti-inflammatory properties, remains underexplored regarding its role in pyroptosis associated with AD. An in vitro AD-like model was established using HaCaT cells stimulated with IFN-γ (10 ng/mL) and TNF-α (10 ng/mL). Sinomenine pretreatment was evaluated for its ability to attenuate inflammation, pyroptosis, and cell damage using ELISA, MTT, LDH assays, flow cytometry, and Western blot. The underlying mechanism was explored via ChIP-qPCR, luciferase assays, and protein interaction studies including co-immunoprecipitation and immunofluorescence. IFN-γ/TNF-α triggered robust pyroptosis in HaCaT cells, characterized by elevated IL-18, IL-6, IL-8, IL-1β, and increased NLRP3 expression levels, cleaved Caspase-1, and GSDMD-N. Sinomenine pretreatment significantly reversed these effects, improving cell viability and reducing inflammatory cytokine production and pyroptosis markers. Mechanistically, sinomenine downregulated TRAF6 expression, a known activator of the NLRP3 inflammasome, by inhibiting its transcriptional regulator IRF1. IRF1 directly bound the TRAF6 promoter and promoted its transcription. Furthermore, sinomenine enhanced TRIM32 expression, which promoted the ubiquitination and proteasomal degradation of IRF1, thus interrupting the IRF1/TRAF6/NLRP3 axis. Sinomenine protects HaCaT cells from IFN-γ/TNF-α-induced pyroptosis by promoting TRIM32-mediated degradation of IRF1, leading to downregulation of TRAF6 and subsequent attenuation of the NLRP3 inflammasome activation. These findings highlight the therapeutic potential of sinomenine for inflammatory skin diseases like AD.