Research Paper Volume 12, Issue 2 pp 1159—1170

Circular RNA HIPK3 downregulation mediates hydrogen peroxide-induced cytotoxicity in human osteoblasts

Jinqian Liang 1, *, , Ying-chao Shen 2, *, , Xiang-yang Zhang 3, *, , Chong Chen 1, , Hong Zhao 1, , Jianhua Hu 1, ,

  • 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
* Co-first authors

received: September 27, 2019 ; accepted: December 25, 2019 ; published: January 18, 2020 ;

https://doi.org/10.18632/aging.102674
How to Cite

Copyright © 2020 Liang et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY 3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Abstract

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.

Abbreviations

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].