CircRNA AFF4 induced by KDM1A promotes osteogenic differentiation through FNDC5/Irisin pathway

Abstract

Background Circular RNA (circ) AFF4 was documented to regulate osteogenesis but the underlying mechanism remains to be elucidated. The preliminary study showed that circ_AFF4 may promote osteogenesis via FNDC5/Irisin. Furthermore, the online prediction tool indicated the interaction of circ_AFF4, insulin-like growth factor-2 mRNA-binding protein 3 (IGF2BP3), FNDC5 and lysine (K)-specific demethylase 1 A (KDM1A). Therefore, this study aims to elucidate the relationships of KDM1A, circ_AFF4, IGF2BP3 and FNDC5/Irisin during osteogenesis. Methods The alkaline phosphatase (ALP) activities and osteogenic-related factors were determined using ALP and alizarin red S (ARS) staining, real-time quantitative PCR(RT-qPCR) and western blot. Immunoprecipitation (RIP), pull-down assay and fluorescence in situ hybridization (FISH) were used to examine the interactions among circ_AFF4/IGF2BP3/FNDC5. A mouse in vivo model was utilized to further confirm the regulatory effect on bone formation. Results Circ_AFF4 and KDM1A expression levels were increased during osteoinduction of BM-MSCs. Knockdown of circ_AFF4 and KDM1A significantly suppressed BM-MSC osteogenesis. We also proved that KDM1A directly bound to circ_AFF4 and FNDC5 promoter and induced circ_AFF4 and FNDC5 expression. Furthermore, circ_AFF4 enhanced the stability of FNDC5 by generating a circ_AFF4, IGF2BP3 and FNDC5 RNA-protein complex, and thereby induced Irisin and osteogenesis. The in vitro data was confirmed with in vivo model. Conclusion These findings elucidate that KDM1A induces circ_AFF4, which promotes promote osteogenesis via IGF2BP3. This study indicates that circ_AFF4 may potentially represent a critical therapeutic target for the diseases.

Circular RNAs: Promising Targets in Osteoporosis

“…Likewise, in human primary MSCs, several other circRNAs were reported as pro-osteogenic, such as circFOXP1 (hsa_circ_0001320) [ 48 ], circ_0001795 [ 49 ], circ_0005564 [ 25 ], circ_0011269 [ 50 ], circ_0019693 [ 51 ], hsa_circ_0026827 [ 52 ], circ_0062582 [ 53 , 54 ], hsa_circ_0070562 [ 55 ], hsa_circ_0001493 [ 55 ], circRUNX2 (hsa_circ_0076694) [ 56 ], circ_AFF4 [ 57 ], circINO80 [ 58 ], circRFWD2 [ 58 ], hsa_circ_0006393 [ 59 ], circRNA_0016624 [ 60 ], circRNA_0048211 [ 61 ], circRNA_33287 [ 29 ], circRNA124534 [ 62 ], circSIPA1L1 [ 63 , 64 ], circVANGL1 [ 65 ], hsa_circ_0001421 [ 66 ], hsa_circ_0076906 [ 67 ], and hsa_circ_0076690 [ 68 ]. Furthermore, circRUNX2 [ 56 ], hsa_circ_0006393 [ 59 ], and circFOXP1 (hsa_circ_0001320) [ 48 ] levels were shown to be impaired in bone from osteoporotic patients, whereas circ_0001795 [ 49 ] was described to be significantly reduced in the bone-marrow (Fig.…”