Microglial activation and failure of the antioxidant defense mechanisms are major hallmarks in different brain injuries, particularly traumatic brain injury (TBI). Cofilin is a cytoskeleton-associated protein involved in actin binding and severing. In our previous studies, we identified the putative role of cofilin in mediating microglial activation and apoptosis in ischemic and hemorrhagic conditions. Others have highlighted the involvement of cofilin in ROS production and the resultant neuronal death; however, more studies are needed to delineate the role of cofilin in oxidative stress conditions. The present study aims to investigate the cellular and molecular effects of cofilin in TBI using both in vitro and in vivo models as well as the first-in-class small-molecule cofilin inhibitor (CI). An in vitro H₂O₂-induced oxidative stress model was used in two different types of cells, human neuroblastoma (SH-SY5Y) and microglia (HMC3), along with an in vivo controlled cortical impact model of TBI. Our results show that treatment with H₂O₂ increases the expression of cofilin and slingshot-1 (SSH-1), an upstream regulator of cofilin, in microglial cells, which was significantly reduced in the CI-treated group. Cofilin inhibition significantly attenuated H₂O₂-induced microglial activation by reducing the release of proinflammatory mediators. Furthermore, we demonstrate that CI protects against H₂O₂-induced ROS accumulation and neuronal cytotoxicity, activates the AKT signaling pathway by increasing its phosphorylation, and modulates mitochondrial-related apoptogenic factors. The expression of NF-E2-related factor 2 (Nrf2) and its associated antioxidant enzymes were also increased in CI-treated SY-SY5Y. In the mice model of TBI, CI significantly activated the Nrf2 and reduced the expression of oxidative/nitrosative stress markers at the protein and gene levels. Together, our data suggest that cofilin inhibition provides a neuroprotective effect in in vitro and in vivo TBI mice models by inhibiting oxidative stress and inflammatory responses, the pivotal mechanisms involved in TBI-induced brain damage.

Neuroinflammation after intracerebral hemorrhage (ICH) is a crucial factor that determines the extent of the injury. Cofilin is a cytoskeleton-associated protein that drives neuroinflammation and microglia activation. A novel cofilin inhibitor (CI) synthesized and developed in our lab has turned out to be a potential therapeutic agent for targeting cofilin-mediated neuroinflammation in an in vitro model of ICH and traumatic brain injury. The current study aims to examine the therapeutic potential of CI in a mouse collagenase model of ICH and examine the neurobehavioral outcomes and its mechanism of action. Male mice were subjected to intrastriatal collagenase injection to induce ICH, and sham mice received needle insertion. Various concentrations (25, 50, and 100 mg/kg) of CI were administered to different cohorts of the animals as a single intravenous injection 3 h following ICH and intraperitoneally every 12 h for 3 days. The animals were tested for neurobehavioral parameters for up to 7 days and sacrificed to collect brains for hematoma volume measurement, Western blotting, and immunohistochemistry. Blood was collected for cofilin, TNF-α, and IL-1β assessments. The results indicated that 50 mg/kg CI improved neurological outcomes, reversed post-stroke cognitive impairment, accelerated hematoma resolution, mitigated cofilin rods/aggregates, and reduced microglial and astrocyte activation in mice with ICH. Microglia morphological analysis demonstrated that CI restored the homeostasis ramification pattern of microglia in mice treated with CI. CI suppressed endoplasmic reticulum stress-related neuroinflammation by inhibiting inflammasomes and cell death signaling pathways. We also showed that CI prevented synaptic loss by reviving the pre- and post-synaptic markers. Our results unveil a novel therapeutic approach to treating ICH and open a window for using CI in clinical practice.

Background: Intracerebral hemorrhage (ICH) is a severe type of stroke with high mortality. Persistent hyperglycemia following ICH is linked to deteriorated neurological functions and death. However, the exacerbating effect of hyperglycemia on ICH injury at the molecular level is still unclear. Therefore, this study explores the impact of diabetes on ICH injury using a non-obese diabetic (NOD) mouse model of type I diabetes mellitus. Methods: NOD and non-diabetic (non-obese resistant) mice subjected to ICH by intrastriatal injection of collagenase were sacrificed three days following the ICH. Brains were collected for hematoma volume measurement and immunohistochemistry. Neurobehavioral assays were conducted 24 h before ICH and then repeated at 24, 48 and 72 h following ICH. Results: NOD mice showed increased hematoma volume and impairment in neurological function, as revealed by rotarod and grip strength analyses. Immunohistochemical staining showed reduced glial cell activation, as indicated by decreased GFAP and Iba1 staining. Furthermore, the expression of oxidative/nitrosative stress markers represented by 3-nitrotyrosine and inducible nitric oxide synthase was reduced in the diabetic group. Conclusions: Overall, our findings support the notion that hyperglycemia exacerbates ICH injury and worsens neurological function and that the mechanism of injury varies depending on the type of diabetes model used.

BACKGROUND: Trauma is a leading cause of pediatric morbidity and mortality. Children with traumatic brain injury (TBI), especially with extracranial injuries, can develop immune suppression and subsequent infection. Immune checkpoint pathways, for example, programmed death-ligand 1 (PD-L1), may contribute. We hypothesized that an anti–PD-L1 antibody would safely prevent immune suppression in a juvenile rat model of polytraumatic TBI. METHODS: Juvenile rats underwent TBI plus systemic hemorrhage (TBI/H) or sham injury. Rats received daily injections of saline or anti–PD-L1 (10 μg or 100 μg) for 7 days. Systemic immune function was assessed by measuring TNFα production in whole blood and splenocytes after ex vivo stimulation with lipopolysaccharide. Cytokines were measured by ELISA or proteome array. Immunofluorescence was used to quantify microglia, astrocytes, and neurons in postinjury day (PID) 7 brain tissue. Cognitive function was assessed by behavioral testing on PID 1. RESULTS: TBI/H resulted in lower TNFα response in whole blood and spleen ( p =0.02) versus sham-injured rats. TBI/H rats treated with 100 μg anti–PD-L1 had higher TNFα response in whole blood ( p =0.04) and spleen ( p =0.02) versus TBI/H rats treated with saline. Lower dose anti–PD-L1 had no effect on immune function. Unstimulated plasma was examined using a proteome array, and higher levels of inflammatory mediators were noted on PID 7 in injured animals treated with 100 µg anti–PD-L1 versus saline. The 100 μg anti–PD-L1 group had higher perilesional microglia ( p =0.004) and astrocyte ( p =0.03) counts, as compared with TBI/H+saline. Barnes maze results were not different between injured rats treated with 10 μg anti–PD-L1 versus sham injury, but the 100 μg dose resulted in worse performance ( p =0.007). CONCLUSIONS: Anti–PD-L1 treatment prevented posttraumatic immune suppression but only at a dose that resulted in higher numbers of perilesional microglia and no improvement in spatial memory. As such, PD-L1 blockade may not be a good candidate for safely reversing immune suppression after pediatric TBI. ( J Trauma Acute Care Surg. 2026;00:00-00. Copyright © 2026 The Author(s). Published byWolters Kluwer Health, Inc. on behalf of the American Association for the Surgery of Trauma.) LEVEL OF EVIDENCE: Not applicable (basic science/animal research).