Objective Chronic pain associated with inflammation is a common clinical problem, and the underlying mechanisms have only begun to be unraveled.Hydrogen sulfide (H2S), an endogenous gaseotransmitter/modulator, is becoming appreciated that it may be involved in a wide variety of processes including inflammation and nociception.However, the molecular and epigenetic mechanisms for expression of H2S synthesizing enzyme cystathioninebeta-synthetase (CBS) in peripheral nociceptive processing remain unknown.The present study is to explore the role ofCBS gene promoter methylation following peripheral inflammation.Methods Complete Freunds adjuvant (CFA) was injected into plantar surface (100 ml each) of hindpaw of adult male Sprague Dawley rats (6-8 week old).Real-time polymerase chain reaction (qPCR) and Western blot analysis were employed to measure expression of mRNA and protein in lumbar dorsal root ganglion (DRGs, L4-L6) from control and CFA-treated rats, respectively.Methylation-specific PCR (MSP) and bisulfite sequencing (BSP) were carried out for determining methylation status of the cbs gene promoter.Results Peripheral inflammation significantly reduced hind paw withdrawal threshold and remarkably upregulated expression of CBS at both protein and mRNA levels in DRGs when compared with control rats.The ratio of methylated and unmethylated CpG island for MSP primers (+325--425; +657~+767) was markedly reduced in inflammatory rats when compared with control group.DNA sequencing of the bisulfite-treated cbs promoter demonstrated that the fragment (F210bp, +496 bp~+705 bp) of cbs promoter region, which contains 11 CpG sites was significantly demethylated in DRGs 3 days after CFA injection.Furthermore, expression of MBD4 and Gadd45α was highly enhanced in CFA-treated rats while expression of DNMT3a and 3b was not significantly altered.Conclusion Our data suggest that peripheral inflammation leads to an active DNA demethylation in cbs gene promoter region, thus contributing to the enhanced CBS expression.Together with our previous report that H2S producing enzyme CBS is sufficient and necessary for development of inflammatory hyperalgesia, it is concluded that epigenetic regulation is involved in inflammatory pain and that this and future indepth investigation may shield light on epigenetic mechanisms of chronic pain and provide evidence for treatment options for the same.