Regulating effect of Circ_ATRNL1 on the promotion of SOX9 expression to promote chondrogenic differentiation of hAMSCs mediated by MiR-145-5p.

Abstract

Circ_ATRNL1 is significantly highly expressed in cartilage tissues of patients with osteoarthritis. This study explored the role and mechanism of circ_ATRNL1 in cartilage differentiation of human adipose-derived mesenchymal stem cells (hAMSCs). hAMSCs were isolated and identified by flow cytometry. The degree of chondrocyte and adipogenic differentiation was assessed using Alcian blue staining and Oil Red O staining, respectively. The expressions of cartilage- and adipogenic-related genes, circ_ATRNL1, and SOX9 were detected by reverse transcription quantitative polymerase chain reaction. The correlation between SOX9 and circ_ATRNL1 was analyzed using Pearson test. Bioinformatics and luciferase analysis were used to detect the overlapped target miRNAs of circ_ATRNL1 and SOX9. The role of circ_ATRNL1/miRNA/SOX9 was examined using functional rescue assays. hAMSCs were identified as CD90-, CD105-, and CD44-positive. The degree of cartilage differentiation of hAMSCs was significantly enhanced after 2 weeks. Cartilage-related genes, circ_ATRNL1 and SOX9, were significantly upregulated, and positively correlated with each other. Circ_ATRNL1 overexpression enhanced hAMSC proliferation and differentiation into chondrogenesis, and promoted the expressions of COL2, Aggrecan, and SOX9. Overexpression of circ_ATRNL1 inhibited the adipogenic differentiation of hAMSCs and the expressions of adipogenic-related genes. miR-145-5p was a target miRNA for circ_ATRNL1 and SOX9. miR-145-5p mimic inhibited hAMSC differentiation toward cartilage, and inhibited the expression of cartilage-related factors. miR-145-5p mimic effectively reversed the regulating effect of circ_ATRNL1 on hAMSCs. Circ_ATRNL1 regulates the promotion of SOX9 expression to promote chondrogenic differentiation of hAMSCs mediated by miR-145-5p.