5-Aza-2’-deoxycytidine enhances lipopolysaccharide-induced inflammatory cytokine expression in human dental pulp cells by regulating TRAF6 methylation

Abstract

ABSTRACT Dental pulp inflammation is a common bacterially driven inflammation characterized by the local accumulation of inflammatory mediators in human dental pulp. DNA methylation is a crucial epigenetic modification that that plays a fundamental role in gene transcription, and its role in inflammation-related diseases has recently attracted attention. However, its role in dental pulp inflammation is poorly understood. This study is aimed to elucidate the role of DNA methylation in lipopolysaccharide (LPS)-induced inflammatory reaction in human dental pulp cells (hDPCs). hDPCs were pretreated with DNA methylation inhibitor 5-aza-2ʹ-deoxycytidine (5-Aza-CdR) and a cytokine antibody array was used to detect LPS-induced cytokine expression. The results indicated that 5-Aza-CdR significantly increased the expression of several pro-inflammatory cytokines in LPS-treated cells, including IL-6, IL-8, GM-CSF, MCP-2 and RANTES. The increased expression levels of IL-6 and IL-8 were further verified by qRT-PCR and ELISA. Furthermore, pretreatment with 5-Aza-CdR resulted in upregulation of p-IKKα/β, p-IκBα, p-p65 and p-ERK in the NK-κB and MAPK pathways. In addition, the 5mC level of the TRAF6 promoter was significantly decreased following 5-Aza-CdR pretreatment in the LPS-stimulated hDPCs. The findings indicate that 5-Aza-CdR significantly enhances the expression of proinflammatory cytokines and activates the NF-κB and MAPK signaling pathways by eliciting a decline in the 5mc level in the TRAF6 promoter in hDPCs, suggesting that DNA methylation may play an important role in dental pulp inflammation. This study highlights the important role of DNA methylation in the immunity defense of dental pulp infection. Graphical Abstract