Differential effects of NOX2 and NOX4 inhibition after rodent spinal cord injury

Abstract

Reactive oxygen species (ROS) are a contributing factor to impaired function and pathology after spinal cord injury (SCI). The NADPH oxidase (NOX) enzyme is a key source of ROS; there are several NOX family members, including NOX2 and NOX4, that may play a role in ROS production after SCI. Previously, we showed that a temporary inhibition of NOX2 by intrathecal administration of gp91ds-tat immediately after injury improved recovery in a mouse SCI model. However, chronic inflammation was not affected by this single acute treatment, and other NOX family members were not assessed. Therefore, we aimed to explore the effect of genetic knockout (KO) of NOX2 or acute inhibition of NOX4 with GKT137831. A moderate SCI contusion injury was performed in 3 month old NOX2 KO and wild-type (WT) mice, who received no treatment or GKT137831/vehicle 30 minutes post-injury. Motor function was assessed using the Basso Mouse Scale (BMS), followed by evaluation of inflammation and oxidative stress markers. NOX2 KO mice, but not GKT137831 treated mice, demonstrated significantly improved BMS scores at 7, 14, and 28 days post injury (DPI) in comparison to WT mice. However, both NOX2 KO and GKT137831 significantly reduced ROS production and oxidative stress markers. Furthermore, a shift in microglial activation toward a more neuroprotective, anti-inflammatory state was observed in KO mice at 7 DPI and a reduction of microglial markers at 28 days. While acute alterations in inflammation were noted with GKT137831 administration, this was not sustained through 28 days. In vitro analysis also showed that while GKT137831 reduced ROS production by microglia, it did not translate to changes in pro-inflammatory marker expression within these cells. These data demonstrate that NOX2 and NOX4 play a role in post-injury ROS, but a single dose of NOX4 inhibitor fails to enhance long-term recovery.

Epigenetic mechanism of miR-26b-5p-enriched MSCs-EVs attenuates spinal cord injury

“…NOX4 plays a role in the induction of oxidative stress after SCI [ 24 , 25 ]. In ChIP assay, KDM6A was found to have significant enrichment at the NOX4 promoter in model tissues and cells, and such enrichment was relatively reduced while the enrichment of H3K27me3 was relatively elevated after EVs treatment ( P < 0.01, Fig.…”

“…The imbalance between ROS and antioxidants participates in the functional and pathological impairment after SCI [ 23 ]. NADPH oxidases are a vital source for ROS, and NOX4 is a member of the NADPH oxidase family and plays a driving role in apoptosis, ROS production, and inflammatory responses post SCI [ 24 , 25 ]. Importantly, our bioinformatic data suggested that KDM6A 3′ untranslated region (3′UTR) is a target of miR-26b-5p and the ChIP assay verified the enrichment of KDM6A and H3K27me3 at the NOX4 promoter, suggesting that the interactions between miR-26b-5p and KDM6A and between KDM6A and NOX4 play a role in SCI pathology.…”

Novel NADPH Oxidase-2 Inhibitors as Potential Anti-Inflammatory and Neuroprotective Agents

“…The primary source of ROS in inflammatory cells is NADPH oxidase-2 (Nox2); therefore, the development of pharmacological approaches to inhibit Nox2 is an active area of research with significant translational potential [ 4 , 5 ]. Numerous rodent models of diseases with an inflammatory component showed beneficial effects following the genetic deletion of Nox2-related proteins, validating Nox2 activity as a potential therapeutic target [ 6 , 7 , 8 , 9 ].…”