The Effect of microRNA Targeting IL-17RA in the Regulation of RANKL and OPG Expressions in Stem Cells from Human Exfoliated Deciduous Teeth

Abstract

AbstractmicroRNA is a small RNA molecule able to regulate gene expressions at post transcription level, either via mRNA degradation or translational repression. This study is designed to determine the potential microRNA targeting IL-17RA and its effects towards OPG and RANKL expressions in SHED. Complex filtration process via in silico study, using the most recent algorithmically programmes (DIANA-micro T CDS, mirWalk v2.0, and TargetScan v7.1) was done to predict potential microRNA. The concentration of 25, 50, and 100 nM microRNA targeting GAPDH were optimized to determine the most efficient downregulation activity. The result showed that 50 nM microRNA mimic transfected for 48 h resulted in the lowest level of GAPDH mRNA expression measured by quantitative real time PCR. Following microRNA optimization, SHED were grown for 7 days in complete α-mem supplemented with osteoinducing reagents and treated with 50 ng/mL IL-17A to enhance osteogenic differentiation. Treated cells were then transfected with 50 nM of predicted microRNA (hsa-miR-4524a-3p and hsa-miR-6761-5p) for 48 h. The expressions of IL-17RA, OPG and RANKL were measured by qPCR and normalized with β-actin. The microRNA mimic hsa-miR-4524a-3p downregulated IL-17RA expression more than the microRNA mimic hsa-miR-6761-5p (p < 0.01). Additionally, both OPG and RANKL expressions were downregulated by both mimics, although only OPG expression was significantly decreased. These findings highlight the importance of microRNA targeting IL-17RA and its effects on regulating the expressions of OPG and RANKL in SHED, implying a role in the bone metabolism process.KeywordsBoneInterleukin-17AmicroRNAOsteogenic differentiationOsteoprotegerinRANKLSHED