Advanced oxidation protein products induce microglia-mediated neuroinflammation via MAPKs-NF-κB signaling pathway and pyroptosis after secondary spinal cord injury

Abstract

Background: Inflammatory response mediated by oxidative stress is considered as an important pathogenesis of spinal cord injury (SCI). Advanced oxidation protein products (AOPPs) are novel markers of oxidative stress and their role in inflammatory response after SCI remained unclear. This study aimed to investigate the role of AOPPs in SCI pathogenesis and explore the possible underlying mechanisms. Methods: A C5 hemi-contusion injury was induced in Sprague-Dawley rats to confirm the involvement of AOPPs after SCI. For in vivo study, apocynin, the NADPH oxidase inhibitor was used to study the neuroprotective effects after SCI. For in vitro study, the BV2 microglia cell lines were pretreated with or without the inhibitor or transfected with or without small interference RNA (siRNA) and then stimulated with AOPPs. A combination of molecular and histological methods was used to clarify the mechanism and explore the signaling pathway both in vivo and in vitro. One-way analysis of variance (ANOVA) was conducted with Bonferroni post hoc tests to examine the differences between groups. Results: The levels of AOPPs in plasma and cerebrospinal fluid as well as the contents in the spinal cord showed significant increase after SCI. Meanwhile, apocynin ameliorated tissue damage in the spinal cord after SCI, improving the functional recovery. Immunofluorescence staining and western blot analysis showed activation of microglia after SCI, which was in turn inhibited by apocynin. Pretreated BV2 cells with AOPPs triggered excessive generation of reactive oxygen species (ROS) by activating NADPH oxidase. Increased ROS induced p38 MAPK and JNK phosphorylation, subsequently triggering nuclear translocation of NF-κB p65 to express pro-inflammatory cytokines. Also, treatment of BV2 cells with AOPPs induced NLRP3 inflammasome activation and cleavage of Gasdermin-d (GSDMD), causing pyroptosis. This was confirmed by cleavage of caspase-1, production of downstream mature interleukin (IL)-1β and IL-18 as well as rupture of rapid cell membrane.

ROS Generation in Microglia: Understanding Oxidative Stress and Inflammation in Neurodegenerative Disease

“…Attenuating fluoride-induced oxidative stress in BV-2 microglia with the antioxidant, melatonin, results in reduced JNK activation [ 154 ]. Moreover, BV-2 microglia treated with advanced oxidation protein products, a hallmark of oxidative stress, displayed increased NOX4 expression and elevated ROS, along with p38 MAPK and JNK activation [ 155 ]. ROS from NOX4 also contribute to activation of the NLRP3 inflammasome, a driver of both amyloid-β [ 156 ] and tau [ 23 ] pathology.…”

“…ROS from NOX4 also contribute to activation of the NLRP3 inflammasome, a driver of both amyloid-β [ 156 ] and tau [ 23 ] pathology. Knockdown of NOX4 by RNAi reduced NLRP3 activation, suggesting a potential target to reduce NLRP3-stimulated pathology in neurodegenerative disease [ 155 ]. In a model of spinal cord injury, apocynin, a NOX-inhibitor (see above), reduced activation of p38 MAPK and JNK and expression of TNFα and IL1β [ 155 ].…”

“…Knockdown of NOX4 by RNAi reduced NLRP3 activation, suggesting a potential target to reduce NLRP3-stimulated pathology in neurodegenerative disease [ 155 ]. In a model of spinal cord injury, apocynin, a NOX-inhibitor (see above), reduced activation of p38 MAPK and JNK and expression of TNFα and IL1β [ 155 ]. Whilst the ROS generated here are believed to originate from NOX4 expression, the specificity of apocynin for NOX4 has been questioned [ 157 ].…”

Autophagy inhibitor treatment alleviates cerebral inflammatory responses in a mouse model of Japanese encephalitis

“…The histopathological results of this study showed that the brain tissue of mice in the JEV group and the JEV + Rapa group had obvious vascular in ammation and late glial cell proliferation, while the mice in the JEV + Wort group and the JEV + CQ group had slight vascular in ammatory response and obvious glial cell proliferation, other groups of mice showed normal. JEV infection leads to excessive microglia activation and the subsequent release of numerous pro-in ammatory cytokines, resulting in an in ammatory storm [27][28]. In order to attempt to further con rm that autophagy inhibitors can alleviate the vascular in ammatory responses of brain tissues in JEV-challenged mice, we observed that IL-6, IL-1β, and TNF-α were signi cantly downregulated in the brain tissues of JEV-challenged mice treated with autophagy inhibitors compared with those of the JEV and JEV + Rapa groups.…”

“…JE is an in ammatory disease of the central nervous system. JEV infection can cause excessive activation of microglia in the brain and, in turn, release of a large number of pro-in ammatory cytokines such as IL-6, TNF-α, and RANTES, which promote the migration and penetration of numerous white blood cells into the brain, resulting in an in ammatory storm that causes severe damage to the brain tissues [27][28].…”