Capsaicin suppresses microglial inflammation through TRPV1-SSR4 interaction.

BACKGROUND: Transient receptor potential vanilloid 1 (TRPV1) is increasingly recognized as an important regulator of microglial response during neuroinflammation, a key pathological feature of various neurodegenerative diseases such as Alzheimer's disease. Capsaicin, the primary pungent component in chili peppers, executes its anti-inflammatory properties as a TRPV1 agonist. Although the mechanism of TRPV1 channel activation and signal transduction by capsaicin has been extensively studied, the protein-protein interaction-based mechanisms by which TRPV1 modulates microglial inflammatory responses in the context of neurodegeneration remains unclear.

METHODS: We used a lipopolysaccharide-induced acute neuroinflammation model to examine the effects of TRPV1 activation and pharmacological blockade of TRPV1 channel gating on microglial inflammatory responses. Proximity labeling-based proteomics, bioluminescence resonance energy transfer, and NanoLuc Binary Technology were used to identify and characterize TRPV1-interacting proteins. Signal sequence receptor subunit 4 (SSR4) knockdown and forced interaction between TRPV1 and SSR4 were performed to assess the functional contribution of the TRPV1-SSR4 interaction. Structural modeling, BRET-based domain mapping, and colocalization analyses were further used to examine the molecular and subcellular features of SSR4 recruitment by TRPV1.

RESULTS: In the neuroinflammatory model, the TRPV1 agonist capsaicin attenuated pro-inflammatory cytokine production, neuronal apoptosis, microgliosis, and microglial soma enlargement in a TRPV1-dependent manner. Notably, pharmacological blockade of TRPV1 channel gating by capsazepine did not abolish the anti-inflammatory action of capsaicin, suggesting an alternative mechanism of action beyond the canonical ligand-gated Ca2⁺ influx. Screening of TRPV1 prey proteins and validation strategy identified SSR4 as a capsaicin-responsive, TRPV1-associated protein. SSR4 knockdown in microglia significantly diminished the suppression of inflammatory responses caused by TRPV1 activation. Conversely, forced interaction between TRPV1 and SSR4 partially mimicked the anti-inflammatory effects of TRPV1 activation. Further structural and cellular analyses suggested that the transmembrane region of TRPV1 supports SSR4 recruitment.

CONCLUSIONS: Our findings suggest that capsaicin suppresses microglial inflammation mediated by the TRPV1-SSR4 interaction, providing new insights into the chemical biology basis of TRPV1 interoceptive signaling and potential applications of natural TRPV1 agonists for modifying therapy of neuroinflammation-associated neurodegeneration.