Fasudil prevents neomycin‑induced hair cell damage by inhibiting autophagy through the miR‑489/NDP52 signaling pathway in HEI‑OC1 cells

Li,Wei; Zhang,Yanqiu; Xu,Jifeng; Chen,Jincan; Gao,Xia

doi:10.3892/etm.2021.10965

Fasudil prevents neomycin‑induced hair cell damage by inhibiting autophagy through the miR‑489/NDP52 signaling pathway in HEI‑OC1 cells

Authors:
- Wei Li
- Yanqiu Zhang
- Jifeng Xu
- Jincan Chen
- Xia Gao
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Affiliations: Department of Otolaryngology Head and Neck Surgery, The Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu 221006, P.R. China, Department of Otolaryngology Head and Neck Surgery, Xuzhou Cancer Hospital, Xuzhou, Jiangsu 221005, P.R. China, Department of Otolaryngology Head and Neck Surgery, The First Clinical Medical College of Xuzhou Medical University, Xuzhou, Jiangsu 221004, P.R. China, Research Institute of Otolaryngology, Gulou Hospital Affiliated to Medical College of Nanjing University, Nanjing, Jiangsu 210008, P.R. China
Published online on: November 12, 2021 https://doi.org/10.3892/etm.2021.10965
Article Number: 43

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Abstract

Hearing loss is a common sensory disorder that is mainly caused by the loss of hair cells (HCs). Drug‑induced deafness, for which there is currently no effective treatment, is mainly caused by the inappropriate use of aminoglycoside antibiotics. Fasudil (Fas), a novel isoquinoline sulfonamide derivative, has exhibited antioxidant abilities in a number of previous studies. The aim of the present study was to investigate the potential effects of Fas against neomycin (Neo)‑induced hair cell damage and elucidate the underlying mechanism. Flow cytometry and western blot analysis were used to detect the effects of Fas on cell apoptosis and to determine the expression levels of autophagy‑related proteins, LC3B and Beclin 1, induced by Neo. Mitochondrial membrane potential and reactive oxygen species (ROS) levels were detected using fluorescent probes. The effect of Fas on Neo‑induced hair cell injury marker, GFP‑LC3B, was also examined using the immunofluorescence technique. Fas was found to inhibit Neo‑induced mitochondrial autophagy and mitochondrial membrane potential decline, in addition to reducing ROS levels and cell apoptosis caused by Neo treatment. However, Fas failed to inhibit the Neo‑induced these above changes in cells with NDP52 overexpression. The putative binding sites of microRNA (miR)‑489 on the 3'‑untranslated region of nuclear dot protein 52 (NDP52) were predicted using the TargetScan 7.0 online tool, and this association was further verified using a dual‑luciferase reporter assay. Moreover, the expression of miR‑489 negatively regulated the expression of NDP52. Fas and miR‑489 mimic inhibited the Neo‑induced mitochondrial autophagy and mitochondrial membrane potential decline, in addition to reducing ROS levels and cell apoptosis. Knockdown of miR‑489 expression using a miR‑489 inhibitor blocked the inhibitory effects of Fas on the mitochondrial membrane potential, cell apoptosis and ROS production. Therefore, Fas may upregulate the expression of miR‑489 to negatively regulate the expression of NDP52 at the post‑transcriptional level, which in turn inhibits the activation of mitophagy and cell injury induced by Neo. Thus, Fas may act as a novel therapeutic option in the clinical treatment of hearing loss in the future.

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