Expression Profiling and Functional Characterization of MicroRNAs in Apical Periodontitis

Abstract

Introduction: MicroRNAs (miRNAs) are evolutionarily conserved small noncoding RNAs that may orchestrate the pathogenesis of apical periodontitis (AP). This study aimed to identify differentially expressed miRNAs and investigate their target gene pathways in AP.

Methods: Total RNA was extracted from 10 human AP and 2 healthy apical tissues (controls) and subjected to miRNA sequencing for the identification of differentially expressed miRNAs (>1.5-fold changes). The function of the most up-regulated miRNA was further studied in vitro. miR-10a-5p mimics and inhibitors were introduced to human stem cells from the apical papilla and K-562 cells challenged with lipopolysaccharide, and expressions of predicted target genes were examined via quantitative reverse-transcription polymerase chain reaction and RNA sequencing.

Results: A total of 852 miRNAs were identified, of which 12 were significantly up-regulated (1.54- to 8.44-fold) and 94 were significantly down-regulated (0.14- to 0.67-fold) in AP. Predicted target genes of these miRNAs are involved in inflammation, pain, and related pathways. miR-10a-5p showed the highest expression levels in AP. Overexpression of miR-10a-5p in LPS-challenged stem cells from the apical papilla resulted in down-regulation of messenger RNA levels of TNFA and up-regulation of interleukin IL10. RNA sequencing of K-562 cells treated with miR-10a-5p mimics and inhibitors identified miR-10a-5p target genes associated with multiple pathways, including macrophage-mediated inflammation and coagulation pathways.

Conclusions: Over 100 miRNAs were differentially expressed in AP and appeared to be involved with modulation of genes in inflammatory and immune pathways. MiR-10a-5p was the most significantly up-regulated miRNA in AP and may play a critical role in suppressing inflammation and promoting healing.

Keywords: Apical periodontitis; RNA sequencing; inflammation; miR-10a-5p; microRNA.