Abstract:

Objective

To investigate the expression of miR-146a in normal human dental pulp cells （hDPCs） and hDPCs stimulated by lipopolysaccharide （LPS）， and explore the possible role of miR-146a in pulpitis.

Methods

The expression levels of miR-146a in normal human dental pulp cells and hDPCs stimulated by LPS were detected by real-time PCR. Moreover， miR-146a mimics and miR-146a inhibitor， as well as their counterpart negative controls were transfected into cultured hDPCs by Lipofectamine 2000 for 24 h， followed by 1 μg/ml LPS stimulation for 4 h. MRNA of IL-6 and IL-8 were tested by real-time quantitative PCR， and IL-6 and IL-8 levels in the culture supernatant were detected by ELISA. Moreover， IRAK1 and TRAF6 which previously identified as actual target of miR-146a were examined by Western blot. Comparisons between groups were performed with t test or oneway ANOVA analysis.

Results

It was shown that miR-146a was significantly up-regulated in LPS treated hDPCs compared with normal hDPCs （12 h：t=8.488， P=0.014； 24 h：t=39.661， P＜0.01）.Up-regulation of miR-146a could decrease secretion of IL-6 and IL-8 both in mRNA and supernatant levels （IL-6 PCR： P＜0.001， IL-6 ELISA： P＜0.001； IL-8 PCR： P＜0.001， IL-8 ELISA：P=0.011）. The expression of IRAK1 and TRAF6 of hDPCs transfected with miR-146a mimics was obviously downregulated compared to those transfected with mimics control （IRAK1：P=0.002； TRAF6：P＜0.001）.

Conclusions

MiR-146a is up-regulated in hDPCs induced by LPS. Up-regulation of miR-146a could not only down-regulate IRAK1 and TRAF6 expression， but also decrease IL-6 and IL-8 Ievels. This indicates miR-146a may be involved in the regulation of pulpitis.

Key words: MiR-146a, Dental pulp cell, Lipopolysaccharide, Pro-inflammatory cytokines