CircSPI1_005 ameliorates osteoarthritis by sponging miR-370-3p to regulate the expression of MAP3K9

Background: Osteoarthritis (OA), caused by the destruction of joint cartilage, is the most prevalent form of arthritis, causing pain and stiffness in joints among millions of patients worldwide. Increasing evidence suggests that non-coding RNAs, including circular RNAs, play important roles in the pathogenesis of OA, but the precise signaling pathway is still unclear.

Methods: To study OA, we established a mouse model by the destabilized medial meniscus (DMM) surgery and used IL-1β stimulated human cell line C28/I2 as an in vitro study. To further study the role of circSPI1_005 in regulating cell proliferation and apoptosis, EdU staining and FACS-based (fluorescence-activated cell sorting) apoptosis examination were performed after the manipulation of the expression of circSPI1_005. Also, bioinformatics predictions were conducted to analyze the downstream microRNAs of circSPI1_005 and the protein regulated by circSPI1_005. The luciferase assay and the RNA immunoprecipitation (RIP) assay were used to confirm the binding between circSPI1_005 and the predicted microRNA. To verify the role of circSPI1_005 in regulating OA in vivo, we also over-expressed circSPI1_005 by injecting AAV into previously injured knees to improve the OA symptoms.

Results: In this study, we found that circSPI1_005 was significantly down-regulated in IL-1β treated chondrocyte cell lines and cartilage tissues of the OA mouse model. Overexpression of circSPI1_005 ameliorated OA by increasing proliferation and inhibiting apoptosis, and knockdown of circSPI1_005 in chondrocytes mimicked OA phenotypes. Bioinformatics study showed circSPI1_005 could sponge to miR-370-3p, and mechanistic studies confirmed the functional binding between circSPI1_005 and miR-370-3p. Furthermore, we conducted a TargetScan analysis and found that MAP3K9 (mitogen-activated protein kinase kinase kinase 9) could be the downstream protein effector. The expression level of MAP3K9 was regulated by miR-370-3p and overexpression of MAP3K9 could efficiently ameliorate OA. Also, we over-expressed circSPI1_005 in vivo and found that the cartilage surface in the OA mouse model was improved with overexpression of circSPI1_005.

Conclusions: Collectively, circSPI1_005 could sponge to miR-370-3p to regulate the expression of MAP3K9, ameliorating the progression of osteoarthritis.