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PINK1 Protects Against Gentamicin-Induced Sensory Hair Cell Damage: Possible Relation to Induction of Autophagy and Inhibition of p53 Signal Pathway.

ABSTRACT: Phosphatase and tensin homolog (PTEN)-induced putative kinase 1 (PINK1) is a gatekeeper of mitochondrial quality control. The present study was aimed to examine whether PINK1 possesses a protective function against gentamicin (GM)-induced sensory hair cell (HC) damage in vitro. The formation of parkin particles (a marker revealing the activation of PINK1 pathway which is a substrate of PINK1 and could signal depolarized mitochondria for clearance) and autophagy were determined by immunofluorescence staining. The expressions of PINK1, LC3B, cleaved-caspase 3 and p53 were measured by Western blotting. The levels of reactive oxygen species (ROS) and apoptosis were respectively evaluated by DCFH-DA staining, Annexin V Apoptosis Detection Kit and TUNEL staining. Cell viability was tested by a CCK8 kit. We found that treatment of 400 ?M GM elicited the formation of ROS, which, in turn, led to PINK1 degradation, parkin recruitment, autophagy formation, an increase of p53 and cleaved-caspase 3 in HEI-OC1 cells and murine HCs. In contrast, co-treatment with ROS scavenger N-acetyl-L-cysteine (NAC) inhibited parkin recruitment, alleviated autophagy and p53 pathway-related damaged-cell elimination. Moreover, PINK1 interference contributed to a decrease of autophagy but an increase of p53 level in HEI-OC1 cells in response to GM stimulus. Findings from this work indicate that PINK1 alleviates the GM-elicited ototoxicity via induction of autophagy and resistance the increase of p53 in HCs.

SUBMITTER: Yang Q

PROVIDER: S-EPMC6240688 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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PINK1 Protects Against Gentamicin-Induced Sensory Hair Cell Damage: Possible Relation to Induction of Autophagy and Inhibition of p53 Signal Pathway.

Yang Qianqian Q Zhou Yiwei Y Yin Haiyan H Li Hongrui H Zhou Meijuan M Sun Gaoying G Cao Zhixin Z Man Rongjun R Wang Haibo H Li Jianfeng J

Frontiers in molecular neuroscience 20181112

Phosphatase and tensin homolog (PTEN)-induced putative kinase 1 (PINK1) is a gatekeeper of mitochondrial quality control. The present study was aimed to examine whether PINK1 possesses a protective function against gentamicin (GM)-induced sensory hair cell (HC) damage <i>in vitro</i>. The formation of parkin particles (a marker revealing the activation of PINK1 pathway which is a substrate of PINK1 and could signal depolarized mitochondria for clearance) and autophagy were determined by immunofl ...[more]

PMID: 30483050

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Project description:BackgroundMutations in lipoxygenase homology domain 1 (LOXHD1) cause autosomal recessive inheritance, leading to high-frequency and intermediate-frequency hearing losses in patients. To date, studies on the localization of LOXHD1 gene expression are limited. In this study, we aimed to observe the expressions of Loxhd1b in zebrafish, C57BL/6 murine cochlea, and HEI-OC1 cells.MethodsThe expression of Loxhd1b in the auditory system of zebrafish was explored by in situ hybridization experiments of zebrafish embryos. The expression of Loxhd1b in cochlear and HEI-OC1 cells of C57BL/6 mice was analyzed by immunofluorescence staining. Confocal microscopic in vivo imaging was used to detect the number and morphological characteristics of lateral line neuromasts and inner ear hair cells in zebrafish that knocked down Loxhd1b gene. The effect of knockdown Loxhd1b gene on the development of zebrafish otolith and semicircular canal was observed using microscopic. Transcriptome sequencing was used to identify downstream molecules and associated signaling pathways and validated by western blotting, immunostaining, and rescue experiments.ResultsResults of the in situ hybridization with zebrafish embryos at different time points showed that Loxhd1b was expressed in zebrafish at the inner ear and olfactory pores, while the immunostaining showed that Loxhd1 was expressed in both C57BL/6 mouse cochlea and HEI-OC1 cells. Loxhd1b knockdown causes a decrease in the number of spinal and lateral line neuromasts in the inner ear of zebrafish, accompanied by weakened hearing function, and also leads to developmental defects of otoliths and ear follicles. The results of transcriptomics analysis revealed the downstream molecule brain-derived neurotrophic factor (BDNF) and verified that Loxhd1b and BDNF regulate the formation of zebrafish hair cells by synergistic regulation of BDNF/TrkB/ERK pathway based on western blotting, immunostaining, and rescue experiments.ConclusionThis was the first time that the BDNF/TrkB/ERK pathway was identified to play a critical role in the molecular regulation of the development of zebrafish hair cells and the auditory development by Loxhd1b.

Dataset Information

PINK1 Protects Against Gentamicin-Induced Sensory Hair Cell Damage: Possible Relation to Induction of Autophagy and Inhibition of p53 Signal Pathway.

Publications

PINK1 Protects Against Gentamicin-Induced Sensory Hair Cell Damage: Possible Relation to Induction of Autophagy and Inhibition of p53 Signal Pathway.

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