NPD1 inhibits excessive autophagy by targeting RNF146 and wnt/β-catenin pathway in cerebral ischemia-reperfusion injury

Abstract

Objective: Cerebral ischemia-reperfusion (I/R) injury is a common pathological feature in ischemic stroke. Autophagy plays a key role in I/R-induced neuronal death. Neuroprotectin D1 (NPD1) is a docosahexaenoic acid derivative with neuroprotective and anti-inflammatory properties. The purpose of this study was to investigate the mediatory role of NPD1 on I/R-induced injury and to elucidate the underlying mechanisms involved in this process.

Methods: An I/R injury model was established in PC12 cells induced by oxygen and glucose deprivation/reoxygenation (OGD/R). NPD1 at increasing doses (5, 10, 20, 50, 100 nM) were added to cells one hour before OGD/R. To investigate the effect of ring finger protein 146 (RFP146) deficiency in I/R injury, PC12 cells were transiently transfected with small interfering RNF146 before further experiment.

Results: Compared to the controls, OGD/R-challenged cells exhibited significantly decreased cell viability, induced oxidative stress, and excessive autophagic cell death following OGD/R. Pretreatment with NPD1 protected cells against ischemic injury as evidenced by enhanced cell survival, decreased oxidative stress markers, and a lower level of autophagy compared to drug-free group. OGD/R also increased the level of RFP146 and inhibited the expression of β-catenin in PC12 cells. NPD1 treatment promoted the production of RNF146 and β-catenin in cells following OGD/R experiment. Moreover, RNF146 deficiency significantly inhibited β-catenin expression and reversed the protective effects of NPD1 in OGD/R-induced cells.

Conclusion: NPD1 alleviated excessive autophagy via regulating RNF146 and Wnt/β-catenin signaling, suggesting the potential therapeutic use of NPD1 for the protection against cerebral I/R injury.

Keywords:

• Ischemia-reperfusion (I/R)
• microtubule-associated protein-1 light chain 3 (LC3-I)
• neuroprotectin D1 (NPD1)
• oxygen and glucose deprivation/reoxygenation (OGD/R)
• ring finger protein 146 (RFP146)
• Wnt/β-catenin

Disclosure statement

No potential conflict of interest was reported by the author(s).

Author contributions

QM and HLZ conceived and designed the experiments, YNX、QH and XL analyzed and interpreted the results of the experiments, WSZ and HBL performed the experiments.

Data availability

All data generated or analyzed during this study are included in this published article.

Additional information

Funding

This work was supported by the Science and Technology Plan Project of Guizhou Province [Grant No. LH[2017]7188].