TRAF2 protects against cerebral ischemia-induced brain injury by suppressing necroptosis

Jie Li,Jia Jia,Meijun He,Yanmei Song,Jian Cheng,Zichuang Wang,Shoujiang You,Shanwen Wei,Jingyu Zhang,Yusuo Zhang

doi:10.1038/s41419-019-1558-5

Abstract

Necroptosis contributes to ischemia-induced brain injury. Tumor necrosis factor (TNF) receptor associated factor 2 (TRAF2) has been reported to suppress necroptotic cell death under several pathological conditions. In this study, we investigated the role of TRAF2 in experimental stroke using a mouse middle cerebral artery occlusion (MCAO) model and in vitro cellular models. TRAF2 expression in the ischemic brain was assessed with western blot and real-time RT-PCR. Gene knockdown of TRAF2 by lentivirus was utilized to investigate the role of TRAF2 in stroke outcomes. The expression of TRAF2 was significantly induced in the ischemic brain at 24 h after reperfusion, and neurons and microglia were two of the cellular sources of TRAF2 induction. Striatal knockdown of TRAF2 increased infarction size, cell death, microglial activation and the expression of pro-inflammatory markers at 24 h after reperfusion. TRAF2 expression and necroptosis were induced in mouse primary microglia treated with conditioned medium collected from neurons subject to oxygen and glucose deprivation (OGD) and in TNFα-treated mouse hippocampal neuronal HT-22 cells in the presence of the pan-caspase inhibitor Z-VAD. In addition, TRAF2 knockdown exacerbated microglial cell death and neuronal cell death under these conditions. Moreover, pre-treatment with a specific necroptosis inhibitor necrostatin-1 (nec-1) suppressed the cell death exacerbated by TRAF2 knockdown in the brain following MCAO, indicating that TRAF2 impacted ischemic brain damage through necroptosis mechanism. Taken together, our results demonstrate that TRAF2 is a novel regulator of cerebral ischemic injury.

Highlights

Stroke is a leading cause of mortality and disability worldwide[1], and ischemic stroke accounts for >80% of total stroke
TRAF2 expression was mostly co-localized with the neuronal marker NeuN and the microglial marker Iba[1] in the ischemic cortex and striatum at 24 h after reperfusion (Fig. 1f, g, h, i), and TRAF2+NeuN+ cells accounted for 56.0% and 71.2% of TRAF+ cells in the ipsilateral cortex and striatum (Fig. 1f, g), respectively
tumor necrosis factor α (TNFα), iNOS and CD32 were increased in the ischemic striatum at 24 h after reperfusion in the mouse middle cerebral artery occlusion (MCAO) model (Fig. 2h–j), and striatal TRAF2 knockdown augmented ischemia-induced expression of these pro-inflammatory markers (Fig. 2h–j). These results suggest that TRAF2 induction following MCAO/reperfusion likely plays a protective role in the brain following cerebral ischemia by inhibiting ischemia-induced cell death and neuroinflammation

Summary

Introduction

Stroke is a leading cause of mortality and disability worldwide[1], and ischemic stroke accounts for >80% of total stroke. As ischemia damages the brain tissue by deprivation of oxygen and metabolic substrates, danger signals are released from cells under ischemic stress. The activate pattern recognition receptors on microglia and induce an inflammatory response by expressing proinflammatory mediators, such as tumor necrosis factor α (TNFα), resulting in inflammation-induced necrotic neuronal cell death[2]. Neuronal necrosis and loss of interaction between neurons and microglia further promote the inflammatory signaling[2], exacerbate the ischemic brain injury[3]. Necroptosis is a form of programmed necrosis, and mounting evidence has shown that necroptosis is of great pathophysiological relevance in ischemic brain injury[4,5,6,7,8,9,10]. Inhibition of necroptosis significantly reduces infarct volumes[4,5,7], attenuates inflammatory response[8], improves locomotive ability[9] and cognitive function[8,9] following cerebral ischemia

Methods

Results

Conclusion