Sinomenine alleviates chronic inflammatory pain through DNA methyltransferase 1-mediated DNA methylation of the P2X2 purinergic receptor: an epigenetic mechanism of analgesia.

BACKGROUND

Chronic inflammatory pain lacks effective and non-addictive therapeutic options. Sinomenine (SIN), a non-opioid alkaloid derived from the traditional Chinese medicine Sinomenium acutum, has demonstrated analgesic potential in clinical practice, yet its precise mechanism remains incompletely understood.

PURPOSE: This study aimed to investigate the analgesic mechanism of SIN and to identify a novel epigenetic pathway involved.

METHODS

The effects of SIN were evaluated in complete Freund's adjuvant-induced chronic inflammatory pain models in rats and lipopolysaccharide stimulated BV2 microglial cells using behavioral tests, enzyme-linked immunosorbent assay, Western blot, reverse transcription quantitative PCR, immunohistochemistry, hematoxylin and eosin staining, calcium imaging, and amplicon bisulfite sequencing. Pharmacological interventions with a P2X2 purinergic receptor (P2X2R) agonist BZATP and antagonist A317491, as well as small interfering RNA mediated knockdown of DNA methyltransferase 1 (DNMT1) in BV2 cells, were employed to dissect the pathway.

RESULTS

SIN dose-dependently alleviated pain hypersensitivity and inflammation, accompanied by downregulation of P2X2R expression. SIN inhibited P2X2R-dependentoveractivation of downstream extracellular signal-regulated kinase 1/2 signaling. Amplicon bisulfite sequencing revealed that SIN promoted methylation of the P2X2R gene promoter. Importantly, DNMT1 knockdown in BV2 cells completely abolished the suppressive effect of SIN on P2X2R expression.

CONCLUSION

This study provides the first evidence that SIN suppresses P2X2R transcription via upregulation of DNMT1, leading to P2X2R promoter hypermethylation-a previously unrecognized epigenetic mechanism underlying the analgesic action of this natural product. This DNMT1-dependent pathway offers a novel therapeutic target for analgesic strategies focusing on epigenetic regulation.