Research Papers: Gerotarget (Focus on Aging):

Autophagy through 4EBP1 and AMPK regulates oxidative stress-induced premature senescence in auditory cells

Nana Akagi Tsuchihashi _, Ken Hayashi, Katsuaki Dan, Fumiyuki Goto, Yasuyuki Nomura, Masato Fujioka, Sho Kanzaki, Shizuo Komune and Kaoru Ogawa

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Oncotarget. 2015; 6:3644-3655. https://doi.org/10.18632/oncotarget.2874

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Nana Akagi Tsuchihashi1,2, Ken Hayashi1,3, Katsuaki Dan4, Fumiyuki Goto1, Yasuyuki Nomura5, Masato Fujioka1, Sho Kanzaki1, Shizuo Komune2, Kaoru Ogawa1

1Department of Otorhinolaryngology, Head and Neck Surgery, Keio University, School of Medicine, Tokyo 160–8582, Japan

2Department of Otorhinolaryngology, Head and Neck Surgery, Kyushu University, School of Medicine, Fukuoka 812–0054, Japan

3Department of Otorhinolaryngology, Kamio Memorial Hospital, Tokyo 101–0063, Japan

4Collaborative Research Resources, Core Instrumentation Facility, Keio University, Tokyo 160–8582, Japan

5Department of Otorhinolaryngology-Head and Neck Surgery, Nihon University, School of Medicine, Tokyo 173–8610, Japan

Correspondence to:

Kaoru Ogawa, e-mail: [email protected]

Keywords: premature senescence, autophagy, AMPK, oxidative stress, auditory cell

Received: November 17, 2014     Accepted: December 08, 2014     Published: December 30, 2014


The aim of this study was to determine whether autophagy and AMPK contribute to premature senescence in auditory cells. Incubating HEI-OC1 auditory cells with 5 mM H2O2 for 1 h induced senescence, as demonstrated by senescence-associated β-galactosidase (SA-β-gal) staining. H2O2 treatment significantly delayed population-doubling time, leaving cell viability unchanged. Furthermore, the proportion of SA-β-gal-positive cells significantly increased. Autophagy-related protein expression increased, with Atg7 and LC3-II peaking 6 h and Lamp2 peaking 24 h after H2O2 treatment. The expression of these proteins decreased 48 h after treatment. Transmission electron microscopy revealed lipofuscin and aggregates within autolysosomes, which accumulated markedly in the cytoplasm of HEI-OC1 cells 48 h after treatment. Akt and P70S6 phosphorylation markedly decreased after H2O2 treatment, but 4EBP1 phosphorylation significantly increased 48 h after treatment. After RNAi-mediated knockdown (KD) of Atg7 and AMPK, H2O2-treated cells displayed dense SA-β-gal staining. Also, premature senescence was significantly induced. These suggest that a negative feedback loop may exist between autophagy and AMPK signaling pathways in HEI-OC1 cells. In our model, oxidative stress-induced premature senescence occurred due to impaired autophagy function through 4EBP1 phosphorylation. Our results also indicate that AMPK may regulate premature senescence in auditory cells in an autophagy-dependent and independent manner.

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