miR-192-5p Inhibits Osteo-/odontogenic Differentiation in Dental Pulp Stem Cells by Targeting COL5A1.

Abstract

Objective: To investigate the biological function of miR-192-5p in osteogenic and odontogenic differentiation of dental pulp stem cells (DPSCs).

Methods: Alkaline phosphatase (ALP) activity assay, Alizarin Red S (ARS) staining and western blot analysis were applied to investigate the osteogenic differentiation potential of DPSCs in vitro. The osteogenic capacity was estimated by subcutaneous transplantation in nude mice in vivo. Carboxyfluorescein diacetate, succinimidyl ester (CFSE) assay was used to examine cell proliferation. Bioinformatics analyses, dual-luciferase reporter assays and real-time reverse transcriptase-polymerase chain reactions (RT-PCR) were applied to explore the regulatory mechanism of miR-192-5p.

Results: miR-192-5p decreased the ALP activity, nodule mineralisation and expression of BSP and OCN in DPSCs, and inhibited cell proliferation. Conversely, the miR-192-5p inhibitor motivated the osteogenic and odontogenic differentiation of DPSCs both in vitro and in vivo, along with promoting cell proliferation. COL5A1 was recognised as the target gene of miR-192- 5p through bioinformatics analysis. Furthermore, dual luciferase reporter assays and RT-PCR experiments confirmed this interaction. Subsequent research verified that COL5A1 knockdown negatively affects the osteo-/odontogenic differentiation of DPSCs, with the PI3K/AKT signalling pathway involved in this process. COL5A1 knockdown promoted the proliferation of DPSCs.

Conclusion: miR-192-5p suppressed osteo/odontogenic differentiation by targeting COL5A1 in DPSCs and negatively regulated their proliferation. Conversely, COL5A1 knockdown promoted proliferation.