Orexin-A exerts neuroprotective effect in experimental intracerebral hemorrhage by suppressing autophagy via OXR1-mediated ERK/mTOR signaling pathway

Dexin Zhang; Ying Cui; Manman Zhao; Xuecheng Zheng; Chunyan Li; Jingbo Wei; Kaijie Wang; Jianzhong Cui

doi:10.3389/fncel.2022.1045034

Orexin-A exerts neuroprotective effect in experimental intracerebral hemorrhage by suppressing autophagy via OXR1-mediated ERK/mTOR signaling pathway

Front Cell Neurosci. 2022 Dec 22:16:1045034. doi: 10.3389/fncel.2022.1045034. eCollection 2022.

Authors

Dexin Zhang¹, Ying Cui², Manman Zhao³, Xuecheng Zheng¹, Chunyan Li⁴, Jingbo Wei³, Kaijie Wang⁵, Jianzhong Cui^{1

5}

Affiliations

¹ Department of Surgery, Hebei Medical University, Shijiazhuang, China.
² Department of Neurology, Tangshan Gongren Hospital, Tangshan, China.
³ Department of Histology and Embryology, North China University of Science and Technology, Tangshan, China.
⁴ Department of Neurology, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan, China.
⁵ Department of Neurosurgery, Tangshan Gongren Hospital, Tangshan, China.

Abstract

Background: Orexin-A (OXA) is a polypeptide produced in the hypothalamus, which binds to specific receptors and exerts multiple physiological effects. Autophagy plays a vital role in early brain injury (EBI) after intracerebral hemorrhage (ICH). However, the relationship between OXA and autophagy after ICH has not been confirmed.

Methods: In this study, the protective role of OXA was investigated in a model of hemin-induced injury in PC12 cells and blood-injection ICH model in rats, and its potential molecular mechanism was clarified. Neurobehavioral tests, brain water content, and pathologic morphology were assessed after ICH. Cell survival rate was determined using Cell Counting Kit-8 (CCK-8), while apoptosis was detected using flow cytometry. The autophagy protein LC3 that was originally identified as microtubule-associated protein 1 light 3 was evaluated by immunohistochemistry. The ultrastructural changes of cells following ICH were observed by transmission electron microscopy. Western blotting was performed to determine the expression levels of LC3, p62/SQSTM1 (p62), phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK_1/2), total extracellular signal-regulated kinase 1/2 (t-ERK_1/2), mammalian target of rapamycin (mTOR), and phosphorylated mammalian target of rapamycin (p-mTOR).

Results: OXA treatment significantly improved neurofunctional outcomes, reduced brain edema, and alleviated neuronal apoptosis. OXA administration upregulated p-mTOR and p62, while it downregulated p-ERK_1/2 and LC3; this effect was reversed by the orexin receptor 1 (OXR1) antagonist SB-334867.

Conclusions: This study demonstrates that OXA suppresses autophagy via the OXR1-mediated ERK/mTOR signaling pathway to exert neuroprotective effects, and it might provide a novel therapeutic approach in patients suffering from ICH.

Keywords: autophagy; intracerebral hemorrhage; neuroprotection; orexin A; orexin receptor.