Circ_0003764 Regulates the Osteogenic Differentiation of Periodontal Ligament Stem Cells

Abstract

<p>Objectives: This study aimed to identify circular RNA (circRNA) involved in the osteogenic differentiation of periodontal ligament stem cells (PDLSCs) and investigate its role in regulating differentiation.<br>Methods: Five candidate circRNAs that may be involved in osteogenesis were predicted by the circBank database and previous studies. The circRNA with the most significant changes was screened by qRT-PCR after PDLSCs were cultured in the osteogenic induction medium or medium supplemented with TNF-α (10ng/ml). Identified circRNA was knocked down or overexpressed, and the effect on the osteogenic differentiation of PDLSCs was explored by western blot, qRT-PCR, alkaline phosphatase (ALP) staining, and Alizarin red staining. CCK-8 assay was done to examine the effect of the circRNA on the proliferation of PDLSCs.<br>Results: Circ_0003764 was the most significantly changed circRNA among the five candidate circRNAs (circ_0026344, circ_ACAP2, circ_0003764, circ_0008259, circ_0060731). When PDLSCs were cultured in the osteogenic induction medium for 3 or 7 days, the expression of circ_0003764 was significantly decreased (p<0.05), whereas it was dramatically increased (p<0.05) in TNF-α-induced PDLSCs. Knockdown of circ_0003764 promoted the expression of the osteogenesis-related genes and proteins (RUNX2, ALP, OCN), enhanced the ALP activity, and elevated the mineralization by PDLSCs, as shown by Alizarin red staining. However, with the overexpression of circ_0003764, the osteogenic differentiation capacity of PDLSCs was significantly reduced (p<0.05). The CCK-8 results indicated that circ_0003764 could inhibit the proliferation of PDLSCs.<br>Conclusions: Circ_0003764 inhibits the osteogenic differentiation and proliferation of PDLSCs, which indicates its use as a diagnostic and therapeutic target in bone regeneration-related diseases.<br></p>