Associations between Huwe1 and autophagy in rat cerebral neuron oxygen‑glucose deprivation and reperfusion injury

He,Guo‑Qian; Chen,Yan; Liao,Hui‑Juan; Xu,Wen‑Ming; Zhang,Wei; He,Guo‑Lin

doi:10.3892/mmr.2020.11611

Associations between Huwe1 and autophagy in rat cerebral neuron oxygen‑glucose deprivation and reperfusion injury

Authors:
- Guo‑Qian He
- Yan Chen
- Hui‑Juan Liao
- Wen‑Ming Xu
- Wei Zhang
- Guo‑Lin He
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Affiliations: Key Laboratory of Birth Defects and Related Diseases of Women and Children, Ministry of Education, Chengdu, Sichuan 610041, P.R. China, Department of Medical Oncology, Sichuan Cancer Hospital and Institute, Sichuan Cancer Center, Cancer Hospital Affiliated to School of Medicine, Chengdu, Sichuan 610041, P.R. China
Published online on: October 19, 2020 https://doi.org/10.3892/mmr.2020.11611
Pages: 5083-5094
Copyright: © He et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Autophagy and the ubiquitin proteasome system (UPS) are two major protein degradation pathways involved in brain ischemia. Autophagy can compensate for UPS impairment‑induced cellular dysfunction. HECT, UBA and WWE domain containing E3 ubiquitin protein ligase 1 (Huwe1), an E3 ubiquitin ligase, serves critical roles in nervous system plasticity, regeneration and disease. However, the role of Huwe1 in autophagy in brain ischemia/reperfusion (I/R) injury remains unknown. The aim of the present study was to investigate the crosstalk between autophagy and the UPS in brain ischemia. The present study established an oxygen‑glucose deprivation and reperfusion (OGD/R) model in rat primary cortex neurons in vitro. Lentiviral interference was used to silence the expression of Huwe1. An autophagy promoter (rapamycin), an autophagy inhibitor (wortmannin) and a JNK pathway inhibitor (SP600125) were also used in the current study. Cellular autophagy‑related proteins, including Beclin‑1, autophagy related (ATG) 7, ATG5, ATG3 and microtubule associated protein 1 light chain 3 α, and apoptosis‑related proteins, such as P53, cleaved caspase 3, Bax and Bcl2, were detected via western blotting and immunocytochemistry. Neuronal apoptosis was evaluated using a TUNEL assay. The results demonstrated that silencing Huwe1 increased the expression levels of autophagy‑related proteins at 24 h after OGD/R. Treatment with a JNK inhibitor or cotreatment with Huwe1 shRNA significantly increased autophagy. Rapamycin increased apoptosis under OGD/R conditions. However, treatment with Huwe1 shRNA decreased the number of TUNEL‑positive cells at 24 h after OGD/R. Cotreatment with Huwe1 shRNA and wortmannin alleviated neuronal apoptosis under OGD/R conditions compared with cotreatment with DMSO. Collectively, the present results suggested that silencing Huwe1 was accompanied by a compensatory induction of autophagy under OGD/R conditions. Furthermore, the JNK pathway may be a key mediator of the interaction between Huwe1 and autophagy in response to UPS impairment.

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