Oxidative stress induced by NOX2 contributes to neuropathic pain via plasma membrane translocation of PKC\u03b5 in rat dorsal root ganglion neurons

Jing Xu,Yi Yan,Junfei Wang,Daying Zhang,Mengye Zhu,Tao Liu,Shinan Wu,Changyu Jiang,Jianmei Wang

doi:10.1186/s12974-021-02155-6

Abstract

BackgroundNicotinamide adenine dinucleotide phosphate oxidase 2 (NOX2)-induced oxidative stress, including the production of reactive oxygen species (ROS) and hydrogen peroxide, plays a pivotal role in neuropathic pain. Although the activation and plasma membrane translocation of protein kinase C (PKC) isoforms in dorsal root ganglion (DRG) neurons have been implicated in multiple pain models, the interactions between NOX2-induced oxidative stress and PKC remain unknown.MethodsA spared nerve injury (SNI) model was established in adult male rats. Pharmacologic intervention and AAV-shRNA were applied locally to DRGs. Pain behavior was evaluated by Von Frey tests. Western blotting and immunohistochemistry were performed to examine the underlying mechanisms. The excitability of DRG neurons was recorded by whole-cell patch clamping.ResultsSNI induced persistent NOX2 upregulation in DRGs for up to 2 weeks and increased the excitability of DRG neurons, and these effects were suppressed by local application of gp91-tat (a NOX2-blocking peptide) or NOX2-shRNA to DRGs. Of note, the SNI-induced upregulated expression of PKCε but not PKC was decreased by gp91-tat in DRGs. Mechanical allodynia and DRG excitability were increased by ψεRACK (a PKCε activator) and reduced by εV1-2 (a PKCε-specific inhibitor). Importantly, εV1-2 failed to inhibit SNI-induced NOX2 upregulation. Moreover, the SNI-induced increase in PKCε protein expression in both the plasma membrane and cytosol in DRGs was attenuated by gp91-tat pretreatment, and the enhanced translocation of PKCε was recapitulated by H2O2 administration. SNI-induced upregulation of PKCε was blunted by phenyl-N-tert-butylnitrone (PBN, an ROS scavenger) and the hydrogen peroxide catalyst catalase. Furthermore, εV1-2 attenuated the mechanical allodynia induced by H2O2ConclusionsNOX2-induced oxidative stress promotes the sensitization of DRGs and persistent pain by increasing the plasma membrane translocation of PKCε.

Highlights

Nicotinamide adenine dinucleotide phosphate oxidase 2 (NOX2)-induced oxidative stress, including the production of reactive oxygen species (ROS) and hydrogen peroxide, plays a pivotal role in neuropathic pain
We investigated whether oxidative stress induced by NOX2 in dorsal root ganglion (DRG) leads to mechanical allodynia in spared nerve injury (SNI) models of neuropathic pain and the underlying mechanisms
SNI surgery induces persistent upregulation of NOX2 in DRGs The 50% paw withdrawal threshold (PWT) in the ipsilateral hind paw was significantly decreased at day 3 (p < 0.05) and persisted until day 14 (p < 0.01) after SNI surgery (Fig. 1a, compared with the sham-operation group, Mann–Whitney U test)

Summary

Introduction

Nicotinamide adenine dinucleotide phosphate oxidase 2 (NOX2)-induced oxidative stress, including the production of reactive oxygen species (ROS) and hydrogen peroxide, plays a pivotal role in neuropathic pain. A proposal that O2− was intimately involved in the inflammatory response was raised as early as the 1970s [6] It implicates that reactive oxygen species (ROS) is one of mediators of neuroinflammation [7] as·O2−, H2O2, and·OH occurs at the site of inflammation [8]. Kallenborn-Gerhardt et al showed that nerve injury activated NOX2-mediated oxidative stress in dorsal root ganglia (DRGs), leading to neuropathic pain behavior, which were reduced in Nox2-deficient mice [12]. This is suggested that NOX2 plays a critical role in neuropathic pain.

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