miRNA-21, -29 Family and -155 Expression in Gingival Crevicular Fluid during Orthodontic Tooth Movement

MicroRNAs (miRNA) are short non-coding RNAs that act as post-transcription regulators of gene expression that may serve as potential biomarkers for understanding and modulating orthodontic tooth movement (OTM). miRNAs have been quantified in various bodily fluids including gingival crevicular fluid (GCF) and have been shown to be stable under harsh conditions, thus making them attractive candidates as biomarkers. The ultimate goal of understanding miRNAs is to be able to utilize them diagnostically or therapeutically to reduce orthodontic treatment time and enhance results. Particularly, miRNA-21, -29 family and -155 play critical roles in osteoclastogenesis, osteoblastogenesis, angiogenesis, extracellular matrix homeostasis and the inflammatory process, which are essential for OTM. The aim of the study is to evaluate the expression profiles of secretory miRNA-21, -29a, b, c and -155 in GCF during the course of OTM. Furthermore, we investigated potential correlations between the miRNA expression profile and the rate of tooth movement. A total of 15 subjects who were receiving orthodontic treatment at the University of Illinois at Chicago’s orthodontic clinic, were recruited for this study. Due to some dropouts, analysis was completed only on 12 healthy subjects between 11 and 21 years of age. The inclusion criteria consisted of subjects requiring maxillary first premolar extractions for orthodontic treatment with maximum anchorage retraction protocols. Subjects also required excellent oral hygiene throughout the GCF collection component of the study. GCF was collected and digital intraoral scans were performed at the following time points: T0: Prior to bonding of orthodontic brackets T1: The day of canine retraction prior to anesthesia, Temporary anchorage device (TAD) placement and activation of closed coil springs T2: 2 weeks post canine retraction initiation T3: 5 weeks post canine retraction initiation T4: 7 weeks post canine retraction initiation Total miRNA was isolated from the GCF samples and quantified using realtime miRNA RT-qPCR. Human let-7d, let-g and let-7i expression were used for normalization of expression for each studied miRNA. The intraoral digital scans were analyzed using Geomagic Control™ 2014 to assess the distance of tooth movement over time. Nonparametric tests including Friedman repeated measures test, Wilcoxon signed rank test with Bonferroni correction and Spearman’s rho correlation tests were used to evaluate the data (P=0.05). Over the 7-week canine retraction period, the teeth moved at a rate of 0.2 mm/week. Significant change of miRNA-155 expression in GCF was detected during tooth movement from T0 – T2 (p-value=0.006). Mild correlations were found between tooth movement and miRNA-21 at 2- and 5-weeks (0.579, p-value=0.010 and 0.484, p-value=0.036) and miRNA-29b at 2 weeks (0.458, p-value=0.037). These findings reveal that miRNA-155 expression significantly changed at an early time point during OTM and there was a correlation between OTM and miRNA-21 and miRNA-29b expression. Therefore, these miRNAs may be useful for modulating of the rate of OTM.