Abstract
Dental pulp inflammation is a widespread public health problem caused by oral bacterial infections and can progress to pulp necrosis and periapical diseases. N6‐methyladenosine (m6A) is a prevalent epitranscriptomic modification in mRNA. Previous studies have demonstrated that m6A methylation plays important roles in cell differentiation, embryonic development and stress responses. However, whether m6A modification affects dental pulp inflammation remains unknown. To address this issue, we investigated the expression of m6A and N6‐adenosine methyltransferase (METTL3, METTL14) as well as demethylases (FTO, ALKBH5) and found that the levels of m6A and METTL3 were up‐regulated in human dental pulp cells (HDPCs) stimulated by lipopolysaccharide (LPS). Furthermore, we knocked down METTL3 and demonstrated that METTL3 depletion decreased the expression of inflammatory cytokines and the phosphorylation of IKKα/β, p65 and IκBα in the NF‐κB signalling pathway as well as p38, ERK and JNK in the MAPK signalling pathway in LPS‐induced HDPCs. The RNA sequencing analysis revealed that the vast number of genes affected by METTL3 depletion was associated with the inflammatory response. Previous research has shown that METTL3‐dependent N6‐adenosine methylation plays an important role in mRNA splicing. In this study, we found that METTL3 knockdown facilitated the expression of MyD88S, a splice variant of MyD88 that inhibits inflammatory cytokine production, suggesting that METTL3 might inhibit the LPS‐induced inflammatory response of HDPCs by regulating alternative splicing of MyD88. These data shed light on new findings in epitranscriptomic regulation of the inflammatory response and open new avenues for research into the molecular mechanisms of dental pulp inflammation.
Highlights
N6-methyladenosine (m6A), methylation at the N6 position of adenosine, has been identified as the most abundant epitranscriptomic modification in eukaryotic mRNA since its discovery in the 1970s [1,2,3]
The total m6A content and methyltransferase-like 3 (METTL3) expression level are up-regulated in LPS-treated human dental pulp cells (HDPCs)
Similar up-regulation of METTL3 protein expression was observed using Western blotting (Fig. 1C and D). These findings suggest that the total m6A content and the METTL3 expression level are up-regulated in LPS-treated HDPCs
Summary
N6-methyladenosine (m6A), methylation at the N6 position of adenosine, has been identified as the most abundant epitranscriptomic modification in eukaryotic mRNA since its discovery in the 1970s [1,2,3]. N6-adenosine methylation is installed by an m6A methyltransferase complex including methyltransferase-like 3 (METTL3), METTL14 and the Dental pulp inflammation is a typical inflammatory disease characterized by local accumulation of inflammatory mediators in dental pulp, and this inflammation can progress to pulp necrosis and periapical diseases [11, 12]. Lipopolysaccharide (LPS) from the cell walls of Gramnegative bacteria can penetrate the pulp and trigger the inflammatory response [14].
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