Research Paper Volume 12, Issue 2 pp 1159—1170
Circular RNA HIPK3 downregulation mediates hydrogen peroxide-induced cytotoxicity in human osteoblasts
- 1 Department of Orthopedic Surgery, Peking Union Medical College Hospital, Beijing, China
- 2 Department of Orthopaedics, Changshu Hospital of Traditional Chinese Medicine, Changshu, China
- 3 Department of Orthopaedics, Tongren Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
Received: September 27, 2019 Accepted: December 25, 2019 Published: January 18, 2020https://doi.org/10.18632/aging.102674
How to Cite
Hydrogen peroxide (H2O2) induces oxidative injury to human osteoblasts. The expression and potential function of circular RNA HIPK3 (circHIPK3) in H2O2-treated human osteoblasts were tested. We show that H2O2 significantly downregulated circHIPK3 in OB-6 cells and primary human osteoblasts. Furthermore, circHIPK3 levels were decreased in the necrotic femoral head tissues of dexamethasone-treated patients. In OB-6 osteoblastic cells and primary human osteoblasts, forced overexpression of circHIPK3 by a lentiviral construct alleviated H2O2-induced viability reduction, cell death and apoptosis. Contrarily, circHIPK3 silencing by targeted shRNA potentiated H2O2-induced cytotoxicity in OB-6 cells and primary human osteoblasts. Moreover, circHIPK3 downregulation by H2O2 induced miR-124 accumulation in OB-6 cells and primary human osteoblasts. On the contrary, miR-124 inhibition by transfection of the miR-124 inhibitor protected human osteoblasts from H2O2. Importantly, forced overexpression of miR-124 by transfection of the miR-124 mimic induced significant cytotoxicity in OB-6 cells and primary human osteoblasts. H2O2 downregulated miR-124’s targets, cyclin dependent kinase 6 and Rho-Associated Protein Kinase 1, in human osteoblasts. In conclusion circHIPK3 downregulation mediates H2O2-induced cytotoxicity in human osteoblasts.
MTT: 3-[4,5-dimethylthylthiazol-2-yl]-2,5 diphenylte-trazolium bromide; circRNAs: circular RNAs; circHIPK3: circular RNA HIPK3; CDK6: cyclin dependent kinase 6; DMEM: Dulbecco's Modified Eagle Medium; ECL: enhanced chemiluminescence; HIPK3: homeodomain-interacting protein kinase 3; LDH: lactate dehydrogenase; FBS: fetal bovine serum; HG: high glucose; H2O2: hydrogen peroxide; HUVECs: human umbilical vein endothelial cells; ncRNAs: non-coding RNAs; OD: optical density; PARP: ploy ADP ribose polymerase; PVDF: polyvinylidene difluoride; qPCR: quantitative real-time polymerase chain reaction assay; ROS: reactive oxygen species; ROCK1: Rho-Associated Protein Kinase 1; TUNEL: [terminal deoxynucleotidyl transferase(TdT)-mediated dUTP nick end labeling].