Controlled Cortical Impact Research Articles

The Effect of Riboflavin on Neurological Rehabilitation after Traumatic Brain Injury in Children.

Traumatic brain injury (TBI), which is the brain impairment and lesion caused by the external force injuring the head and the underlying brain, can cause pediatric death, disability, neurological disorders, and even lifelong disability. This study was to explore the effect of riboflavin (RF) on neurological rehabilitation and functional recovery after TBI. The rat models of TBI were constructed by treating rats with controlled cortical impact (CCI). By treating TBI rats with RF, we investigated whether the administration of RF would affect the sensorimotor function and cognitive ability recovery through adhesive removal test, modified neurological severity score (mNSS), corner test, wire-grip test and the Morris water maze. The effects of RF on lesion volume and water content were investigated using hematoxylin and eosin (H&E) staining and wet-dry method. The Nissl staining and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) staining were used to demonstrate the effect of RF on neural apoptosis. Inflammation-related cytokines of interleukin (IL)-6, IL-1β, tumor necrosis factor (TNF)-α, and transforming growth factor (TGF)-β1 were measured by enzyme-linked immunosorbent assay (ELISA) to evaluate the effect of RF on neuroinflammation. The impact of RF on oxidative stress was assessed by measuring malondialdehyde (MDA) content and superoxide dismutase (SOD) activity, and the platelet endothelial cell adhesion molecule-1 (CD31) staining for observing vessel density, the reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) for measuring vascular endothelial growth factor (VEGF) mRNA expression and western blot for VEGF protein expression were used for evaluated angiogenesis. The administration of RF could facilitate the recovery of neurological function by promoting the recovery of sensorimotor function and cognitive ability (p < 0.05). Furthermore, RF could reduce the lesion volume and water content after TBI and ameliorate neural apoptosis, neuroinflammation, and oxidative stress (p < 0.05). Finally, RF increased vessel density (p < 0.01) and VEGF levels (p < 0.01) in brain tissues after TBI, promoting angiogenesis. RF benefits neurological rehabilitation after TBI by promoting neurological function recovery, ameliorating the pathogenesis after TBI, and facilitating brain vascular remodeling. These findings provide a novel mechanism for RF treating pediatric TBI.

Pediatric Traumatic Brain Injury: Models, Therapeutics, and Outcomes.

Pediatric traumatic brain injury (TBI) is a significant healthcare issue, but potential treatments are absent despite robust investigation in several clinical trials. Factors attributed to clinical TBI, such as heterogeneity of injury and single-dose pharmacological treatments as well as timing of administration, may be reasons for the negative studies. Preclinical models of TBI can reduce some of the impediments by highlighting differences in injury depending on injury severity and location and by conducting dose response studies, thus providing better therapeutic targets and pharmacological profiles for clinical use. In this chapter, there were sufficient reports to make comparisons between the models in terms of pathophysiology, behavioral dysfunction, and the efficacy of therapeutic interventions. The models used to date include controlled cortical impact (CCI), weight drop, fluid percussion, and abusive head trauma. Several therapeutics were identified after CCI injury but none in the other models, which underscores the need for studies evaluating the therapies reported after CCI injury as well as novel potential approaches.

Nicorandil attenuates cognitive impairment after traumatic brain injury via inhibiting oxidative stress and inflammation: Involvement of BDNF and NGF.

Cognitive impairment is a prevalent adverse consequence of traumatic brain injury (TBI). The neuroprotective effects of nicorandil (N-(2-hydroxyethyl)-nicotinamide nitrate) has been previously documented, yet its protective effects against cognitive dysfunction post-TBI remain unclear. Hence, the present study was aimed to evaluate whethernicorandil attenuates cognitive dysfunction in TBI rats and the underlying mechanism behind this process. The TBI model was established with a controlled cortical impact (CCI). The effects of nicorandil on cognitive dysfunction of rats with TBI were examined through Novel object recognition (NOR) test, Y-maze test, and Morris water maze (MWM) task. After behavioral tests, hippocampal tissue was collected for Quantitative real-time PCR, Western blot analysis, and Enzyme-linked immunosorbent assay (ELISA) assay. We observed that nicorandil administration effectively ameliorates learning and memory impairment in TBI rats.Alongside, nicorandil treatment attenuated oxidative stress in the hippocampus of TBI rats, characterized by the decreased reactive oxygen species generation, malondialdehyde, and protein carbonyls levels, and concurrent promotion of antioxidant-related factors (including superoxide dismutase, glutathione peroxidase, and catalase) activities. Additionally, nicorandil treatment attenuated the inflammatory response in the hippocampus of TBI rat, as evidenced by the upregulated levels of interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α), as well as the downregulated level of IL-10. Mechanistically, nicorandil treatment significantly enhanced the mRNA and protein levels of neurotrophic factors, brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) in the hippocampus of TBI rats. These findings suggest that nicorandil mitigates cognitive impairment after TBI by suppressing oxidative stress and inflammation, potentially through enhancing BDNF and NGF levels.

Protective Effects of Hinokitiol on Neuronal Ferroptosis by Activating the Keap1/Nrf2/HO-1 Pathway in Traumatic Brain Injury.

In this study, we investigated the effects of hinokitiol, a small-molecule natural compound, against neuronal ferroptosis after traumatic brain injury (TBI). A controlled cortical impact (CCI) mouse model and excess glutamate-treated HT-22 cells were used to study the effects of hinokitiol on TBI. Hinokitiol mitigated TBI brain tissue lesions and significantly improved neurological function. Neuron loss and iron deposition were ameliorated after hinokitiol administration. Hinokitiol alleviated excessive glutamate-induced intracellular reactive oxygen species (ROS), lipid peroxidation, and Fe2+ accumulation in HT-22. Mechanistically, hinokitiol upregulated heme oxygenase-1 (HO-1) expression, promoted nuclear factor-erythroid factor 2-related factor 2 (Nrf2) nuclear translocation, and inhibited the activation of microglia and astrocyte after TBI. These results suggest that hinokitiol has neuroprotective effects on rescuing cells from TBI-induced neuronal ferroptosis. In summary, hinokitiol is a potential therapeutic candidate for TBI by activating the Nrf2/Keap1/HO-1 signaling pathway.

Vepoloxamer improves functional recovery in rat after traumatic brain injury: A dose-response and therapeutic window study

Traumatic brain injury (TBI) is a major cause of death and disability worldwide. There are no effective therapies available for TBI patients. Vepoloxamer is an amphiphilic polyethylene-polypropylene-polyethylene tri-block copolymer that seals membranes and restores plasma membrane integrity in damaged cells. We previously demonstrated that treatment of TBI rats with Vepoloxamer improves functional recovery. However, additional studies are needed to potentially translate Vepoloxamer treatment from preclinical studies into clinical applications. We thus conducted a study to investigate dose-response and therapeutic window of Vepoloxamer on functional recovery of adult rats after TBI. To identify the most effective dose of Vepoloxamer, male Wistar adult rats with controlled cortical impact (CCI) injury were randomly treated with 0 (vehicle), 100, 300, or 600 mg/kg of Vepoloxamer, administered intravenously (IV) at 2 h after TBI. We then performed a therapeutic window study in which the rats were treated IV with the most effective single dose of Vepoloxamer at different time points of 2 h, 4 h, 1 day, or 3 days after TBI. A battery of cognitive and neurological tests was performed. Animals were killed 35 days after TBI for histopathological analysis. Dose-response experiments showed that Vepoloxamer at all three tested doses (100, 300, 600 mg/kg) administered 2 h post injury significantly improved cognitive functional recovery, whereas Vepoloxamer at doses of 300 and 600 mg/kg, but not the 100 mg/kg dose, significantly reduced lesion volume compared to saline treatment. However, Vepoloxamer at 300 mg/kg showed significantly improved neurological and cognitive outcomes than treatment with a dose of 600 mg/kg.In addition, our data demonstrated that the dose of 300 mg/kg of Vepoloxamer administered at 2 h, 4 h, 1 day, or 3 days post injury significantly improved neurological function compared with vehicle, whereas Vepoloxamer administered at 2 h or 4 h post injury significantly improved cognitive function compared with the 1-day and 3-day treatments, with the most robust effect administered at 2 h post injury. The present study demonstrated that Vepoloxamer improves functional recovery in a dose-and time-dependent manner, with therapeutic efficacy compared with vehicle evident even when the treatment is initiated 3 days post TBI in the rat.

Deferoxamine Induces Autophagy Following Traumatic Brain Injury via TREM2 on Microglia.

Previous studies have indicated that iron disorder, inflammation, and autophagy play an important role in traumatic brain injury (TBI). The triggering receptor expressed on myeloid cells 2 (TREM2), an immunoglobulin superfamily transmembrane receptor, is involved in inflammation. However, the role of TREM2 in modulating the microglia response in TBI has been rarely investigated. The present study aimed to investigate if the iron chelator deferoxamine (DFO) could ameliorate TBI through autophagy mediated by the TREM2. TBI was developed by the controlled cortical impact (CCI) mouse model and stretching of individual primary cortical microglia taken from the tissue of the rat brain. DFO was intraperitoneally used for intervention. Western blotting assay, qRT-PCR, TUNEL staining, immunofluorescence staining, confocal microscopy analysis, transmission electron microscopy, H&E staining, brain water content measurement, and the neurobehavioral assessments were performed. TREM2 expression was up-regulated in cortex of TBI mice model and in microglia stretching model, which was attenuated by DFO. After the mice were subjected to CCI, DFO treatment significantly up-regulated the protein levels of autophagy compared with the TBI group at 3days and caused an increase of autophagic vacuoles. Treatment with DFO reduced TBI-induced cell apoptosis, cerebral edema, neuroinflammation, and motor function impairment in mice, at least partly via the mTOR signaling pathway that facilitates the TREM2 activity. The results indicated that the maintenance of iron homeostasis by DFO plays neuroprotection by modulating the inflammatory response to TBI through TREM2-mediated autophagy. This study suggested that TREM2-mediated autophagy might be a potential target for therapeutic intervention in TBI.

Notoginsenoside R1 attenuates brain injury in rats with traumatic brain injury: Possible mediation of apoptosis via ERK1/2 signaling pathway.

Traumatic brain injury (TBI) occurs worldwide and is associated with high mortality and disability rate. Apoptosis induced by TBI is one of the important causes of secondary injury after TBI. Notoginsenoside R1 (NGR1) is the main phytoestrogen extracted from Panax notoginseng. Many studies have shown that NGR1 has potent neuroprotective, anti-inflammatory, and anti-apoptotic properties and is effective in ischemia-reperfusion injury. Therefore, we investigated the potential neuroprotective effects of NGR1 after TBI and explored its molecular mechanism of action. A rat model of TBI was established using the controlled cortical impact (CCI) method. The expression levels of Bcl-2, Bax, caspase 3, and ERK1/2-related molecules in the downstream pathway were also detected by western blotting. The expression levels of pro-inflammatory cytokines were detected by real-time quantitative PCR. Nissl staining was used to clarify the morphological changes around the injury foci in rats after TBI. Fluoro-Jade B (FJB) and terminal deoxynucleotidyl transferase (TdT) dUTP Nick-End Labeling (TUNEL) fluorescence staining were used to detect the apoptosis of neural cells in each group of rats. The results showed that NGR1 administration reduced neurological deficits after TBI, as well as brain edema and brain tissue apoptosis. It also significantly inhibited the expression of pro-inflammatory cytokines. Furthermore, NGR1 decreased the expression levels of extracellular signal-regulated kinase (ERK) and p-RSK1, which are phosphorylated after trauma. This study suggests that NGR1 can improve neuronal apoptosis in brain injury by inhibiting the ERK signaling pathway. NGR1 is a potential novel neuroprotective agent for the treatment of secondary brain injury after TBI.

Elovanoids and Precursor Free Fatty Acid 34:6 Confer Robust Neuroprotection after Experimental Traumatic Brain Injury

One-third of the close to 70 million people suffering from traumatic brain injury (TBI) have an increased risk for developing chronic cognitive impairments, including Alzheimer’s-like Dementia (AD). We studied neuroprotection by Elovanoid-34:6 (ELV-N34) and its precursor free-fatty- acid-34:6 (FFA34:6) following TBI in male Sprague-Dawley rats using fluid percussion injury (FPI) or controlled cortical impact (CCI) models. At 1 and 24 hours post-surgery, rats were intranasally administered saline, ELV-N34, or FFA34:6 (only in the CCI model). Rats underwent composite neurological testing to assess sensorimotor integration. On day 14, FPI animals were sacrificed, and MRI and diffuse tensor imaging (DTI) were performed to assess lesion size and white matter connectivity. On day 3, CCI animals were used for multi-omic analysis using 10x Chromium, Illumina Sequencing, and Seurat v4.0.1. Total neurological scores for animals administered ELV-N34 and FFA34:6 were lower than controls, and scores for FFA34:6 treated rats were lower than those of ELV34:6 treated rats (p&lt;0.05). MRI and DTI showed smaller lesions and preserved white matter connectivity in the ELV34:6 group. Single cell analysis with ELV-N34 treatment revealed downregulation of Spp1, Fos, and Gfap genes and upregulation of Trem2 and Pld5 genes. FFA34:6 treatment revealed downregulation of inflammatory genes Spp1, Ccl2, and Vim. These changing gene cluster patterns are implicated in neurodegenerative diseases. We demonstrated that ELV-N34 and FFA34:6 convey behavioral, structural, and genetic neuroprotective bioactivity in experimental TBI.

Characterization of Vasogenic and Cytotoxic Brain Edema Formation After Experimental Traumatic Brain Injury by Free Water Diffusion Magnetic Resonance Imaging.

Brain edema formation is a key factor for secondary tissue damage after traumatic brain injury (TBI), however, the type of brain edema and the temporal profile of edema formation are still unclear. We performed free water imaging, a bi-tensor model based diffusion MRI analysis, to characterize vasogenic brain edema (VBE) and cytotoxic edema (CBE) formation up to 7 days after experimental TBI. Male C57/Bl6 mice were subjected to controlled cortical impact (CCI) or sham surgery and investigated by MRI 4h, 1, 2, 3, 5, and 7 days thereafter (n = 8/group). We determined mean diffusivity (MD) and free water (FW) in contusion, pericontusional area, ipsi- and contralateral brain tissue. Free (i.e., non-restricted) water was interpreted as VBE, restricted water as CBE. To verify the results, VBE formation was investigated by in-vivo 2-Photon Microscopy (2-PM) 48h after surgery. We found that MD and FW values decreased for 48h within the contusion, indicating the occurrence of CBE. In pericontusional tissue, MD and FW indices were increased at all time points, suggesting the formation of VBE. This was consistent with our results obtained by 2-PM. Taken together, CBE formation occurs for 48h after trauma and is restricted to the contusion, while VBE forms in pericontusional tissue up to 7 days after TBI. Our results indicate that free water magnetic resonance imaging may represent a promising tool to investigate vasogenic and cytotoxic brain edema in the laboratory and in patients.

Acute brain injury increases pulmonary capillary permeability via sympathetic activation-mediated high fluid shear stress and destruction of the endothelial glycocalyx layer

Neurogenic pulmonary edema secondary to acute brain injury (ABI) is a common and fatal disease condition. However, the pathophysiology of brain-lung interactions is incompletely understood. This study aims to investigate whether sympathetic activation-mediated high fluid shear stress after ABI would damage pulmonary endothelial glycocalyx thus leading to increased pulmonary capillary permeability. The tricuspid annular plane systolic excursion (TAPSE) was detected in a rat model of controlled cortical impact (CCI) and CCI + transection of the cervical sympathetic trunk (TCST). Changes in pulmonary capillary permeability were assessed by analyzing the Evans blue, measuring the dry/wet weight ratio of the lungs and altering protein levels in the bronchoalveolar lavage fluid (BALF). The parallel-plate flow chamber system was used to simulate the fluid shear stress in vitro. Western blotting and immunofluorescence staining were used to determine the expression levels of hyaluronan-binding protein (CEMIP), syndecan-1 and tight junction proteins (TJPs, including claudin-5 and occludin). TCST could restrain cardiac overdrive and sympathetic activation in a rat model of CCI. Compared to the CCI group, the CCI + TCST group showed a reduction of CEMPI (which degrades hyaluronic acid), along with an increase of syndecan-1 and TJPs. CCI + TCST group presented decreasing pulmonary capillary permeability. In vitro, high shear stress (HSS) increased the expression of CEMIP and reduced syndecan-1 and TJPs, which was coordinated with the results in vivo. Our findings show that sympathetic activation-mediated high fluid shear stress after ABI would damage pulmonary endothelial glycocalyx thus leading to increased pulmonary capillary permeability.

Microglial histone deacetylase 2 is dispensable for functional and histological outcomes in a mouse model of traumatic brain injury

The Class-I histone deacetylases (HDACs) mediate microglial inflammation and neurological dysfunction after traumatic brain injury (TBI). However, whether the individual Class-I HDACs play an indispensable role in TBI pathogenesis remains elusive. HDAC2 has been shown to upregulate pro-inflammatory genes in myeloid cells under brain injuries such as intracerebral hemorrhage, thereby worsening outcomes. Thus, we hypothesized that HDAC2 drives microglia toward a pro-inflammatory neurotoxic phenotype in a murine model of controlled cortical impact (CCI). Our results revealed that HDAC2 expression was highly induced in CD16/CD32+ pro-inflammatory microglia 3 and 7d after TBI. Surprisingly, microglia-targeted HDAC2 knockout (HDAC2 miKO) mice failed to demonstrate a beneficial phenotype after CCI/TBI compared to their wild-type (WT) littermates. HDAC2 miKO mice exhibited comparable levels of grey and white matter injury, efferocytosis, and sensorimotor and cognitive deficits after CCI/TBI as WT mice. RNA sequencing of isolated microglia 3d after CCI/TBI indicated the elevation of a panel of pro-inflammatory cytokines/chemokines in HDAC2 miKO mice over WT mice, and flow cytometry showed further elevated brain infiltration of neutrophils and B cells in HDAC2 miKO mice. Together, this study does not support a detrimental role for HDAC2 in microglial responses after TBI and calls for investigation into alternative mechanisms.

Enhancing cognitive function in chronic TBI: The Role of α7 nicotinic acetylcholine receptor modulation

Traumatic brain injury (TBI) results in several pathological changes within the hippocampus that result in adverse effects on learning and memory. Therapeutic strategies to enhance learning and memory after TBI are still in the early stages of clinical development. One strategy is to target the α7 nicotinic acetylcholine receptor (nAChR), which is highly expressed in the hippocampus and contributes to the formation of long-term memory. In our previous study, we found that AVL-3288, a positive allosteric modulator of the α7 nAChR, improved cognitive recovery in rats after moderate fluid-percussion injury (FPI). However, whether AVL-3288 improved cognitive recovery specifically through the α7 nAChR was not definitively determined. In this study we utilized Chrna7 knockout mice and compared their recovery to wild-type mice treated with AVL-3288 after TBI. We hypothesized that AVL-3288 treatment would improve learning and memory in wild-type mice, but not Chrna7−/− mice after TBI. Adult male C57BL/6 wild-type and Chrna7−/− mice received sham surgery or moderate controlled cortical impact (CCI) and recovered for 3 months. Mice were then treated with vehicle or AVL-3288 at 30 min prior to contextual fear conditioning. At 3 months after CCI, expression of α7 nAChR, choline acetyltransferase (ChAT), high-affinity choline transporter (ChT), and vesicular acetylcholine transporter (VAChT) were found to be significantly decreased in the hippocampus. Treatment of wild-type mice at 3 months after CCI with AVL-3288 significantly improved cue and contextual fear conditioning, whereas no beneficial effects were observed in Chrna7−/− mice. Parietal cortex and hippocampal atrophy were not improved with AVL-3288 treatment in either wild-type or Chrna7−/− mice. Our results indicate that AVL-3288 improves cognition during the chronic recovery phase of TBI through modulation of the α7 nAChR.

Effects of Intrathecal Ketamine on Cerebrospinal Fluid Levels of Brain-Derived Neurotrophic Factor and Mechanical Allodynia in a Rat Model of Mild Traumatic Brain Injury.

BACKGROUND Ketamine, a compelling candidate for neuropathic pain management, has attracted interest for its potential to elevate brain-derived neurotrophic factor (BDNF) levels. We aimed to assess the effects of intrathecally administered ketamine on the cerebrospinal fluid (CSF) levels of BDNF(c-BDNF) and allodynia in a rat model of traumatic brain injury (TBI). MATERIAL AND METHODS Forty-five rats were divided into 3 groups: sham operation (Group S), untreated TBI (Group T), and ketamine-treated TBI (Group K), with 15 rats in each group. Rats were anesthetized, and their skulls were secured in a stereotactic frame before undergoing craniotomy. A controlled cortical impact (CCI) was induced, followed by injection of ketamine (3.41 µg/g) into the CSF in Group K. In Group T, no drug was injected after CCI delivery. On postoperative days (POD) 1, 7, and 14, the 50% mechanical withdrawal threshold (50% MWT) and c-BDNF levels were assessed. RESULTS Groups T and K exhibited a significantly lower 50% MWT than Group S on POD 1(6.6 [5.7, 8.7] g, 10.0 [6.8, 11.6] g, and 18.7 [11.6, 18.7] g, respectively; P<0.001). The c-BDNF levels in Group K were significantly higher than those in Groups S and T on POD 1 (18.9 [16.1, 23.0] pg/ml, 7.3 [6.0, 8.8] pg/ml, and 11.0 [10.6, 12.3] pg/ml, respectively; P=0.006). CONCLUSIONS Intrathecal ketamine administration did not exhibit anti-allodynic effects following mild TBI. c-BDNF level is a promising potential indicator for predicting the expression of allodynia after mild TBI.

Emerging role of miR‐223‐3p in regulating sex dimorphic neuroinflammation and cognitive function in mice following controlled cortical impact

AbstractBackgroundNeuroinflammation is a key feature of many neurodegenerative diseases, however its regulation is not well understood. MicroRNAs (miRNAs) are small, non‐coding RNAs that involve in the regulation of many biological and pathological pathways including inflammation. MiR‐223‐3p is macrophage/microglia enriched miRNA that involves in inflammation and immunity. However, the role of miR‐223‐3p in neurodegeneration is unknown.MethodMiR‐223‐3p knockout (KO) and genetic background matched wildtype (WT), female and male young (12‐14‐week) or aged (18‐20‐month) mice were used in the studies. CD11b+ cells were isolated from brain tissues of KO and WT, female and male naïve mice and RT‐qPCR was used to quantify inflammatory markers. Immunofluorescent studies using anti‐IBA1 was used to determine the number of activated microglia in brain sections of aged KO and WT female mice following a CCI. To evaluate the impact of miR‐223‐3p on cognitive and behavior, young (12‐14‐week) or old (18‐20‐month) mice were subjected to CCI followed by a radio arm water maze (RAWM) test (young mice) or Y‐maze test (aged mice) 2 weeks after controlled cortical impact (CCI).ResultOur preliminary studies reveal that deficiency of miR‐223‐3p resulted in a female‐biased elevation of inflammatory transcripts including such as NLRP3, TRAF6, TNFα, NFκB, and ITGAM in brain CD11b+ cells. In addition, immunofluorescent analysis demonstrated that a greater number of IBA1 staining microglia in KO than that of WT mice brains. Furthermore, deficiency of miR‐223‐3p seemed to associate with a deficit in cognitive function in aged female mice following CCI. We also examined the level of CCAAT‐enhancer‐binding protein beta (CEBPβ), an important transcription factor regulating inflammation and a validated target of miR‐223‐3p. Our preliminary data showed that CEBPβ protein was increased in the brain of miR‐223‐3p KO mice brain tissue relative to that of WT.ConclusionOur preliminary data suggest that miR‐223‐3p may regulate a sexually dimorphic neuroinflammation and cognitive function, likely via regulating inflammation transcription factor CEBPβ.

Hyperbaric Oxygen Therapy Alleviates Memory and Motor Impairments Following Traumatic Brain Injury via the Modulation of Mitochondrial-Dysfunction-Induced Neuronal Apoptosis in Rats.

Traumatic brain injury (TBI) is a leading cause of morbidity and mortality in young adults, characterized by primary and secondary injury. Primary injury is the immediate mechanical damage, while secondary injury results from delayed neuronal death, often linked to mitochondrial damage accumulation. Hyperbaric oxygen therapy (HBOT) has been proposed as a potential treatment for modulating secondary post-traumatic neuronal death. However, the specific molecular mechanism by which HBOT modulates secondary brain damage through mitochondrial protection remains unclear. Spatial learning, reference memory, and motor performance were measured in rats before and after Controlled Cortical Impact (CCI) injury. The HBOT (2.5 ATA) was performed 4 h following the CCI and twice daily (12 h intervals) for four consecutive days. Mitochondrial functions were assessed via high-resolution respirometry on day 5 following CCI. Moreover, IHC was performed at the end of the experiment to evaluate cortical apoptosis, neuronal survival, and glial activation. The current result indicates that HBOT exhibits a multi-level neuroprotective effect. Thus, we found that HBOT prevents cortical neuronal loss, reduces the apoptosis marker (cleaved-Caspase3), and modulates glial cell proliferation. Furthermore, HBO treatment prevents the reduction in mitochondrial respiration, including non-phosphorylation state, oxidative phosphorylation, and electron transfer capacity. Additionally, a superior motor and spatial learning performance level was observed in the CCI group treated with HBO compared to the CCI group. In conclusion, our findings demonstrate that HBOT during the critical period following the TBI improves cognitive and motor damage via regulating glial proliferation apoptosis and protecting mitochondrial function, consequently preventing cortex neuronal loss.

Oxyberberine an oxoderivative of berberine confers neuroprotective effects in controlled-cortical impact mouse model of traumatic brain injury

Background Traumatic brain injury (TBI) is known as a silent epidemic that causes many deaths and disabilities worldwide. We examined the response of oxyberberine (OBB) in lipopolysaccharide-stimulated BV2 microglial cells and a controlled-cortical impact (CCI) mouse model of TBI. Methods We synthesized OBB from berberine, and also prepared OBB-nanocrystals (OBB-NC). Male C57BL/6 mice were used for CCI surgery, and post-CCI neurobehavioral deficits were assessed from 1 h after injury through 21 days post-injury (dpi). Results OBB treatment reduced the lipopolysaccharide-triggered elevated levels of reactive oxygen species, nitric oxide, and nuclear factor kappa B (NF-κB) in BV2 microglial cells, indicating a neuroprotective potential. CCI-operated mice exhibited significant neurological deficits on 1, 3, and 5 dpi in neurological severity scoring and rotarod assay. OBB (25 and 50 mg/kg/day) and OBB-NC (3 mg/kg/day) ameliorated these neurological aberrations. Mice subjected to CCI surgery also displayed anxiogenic- and depression-like behaviours, and cognitive impairments in forced-swimming test and elevated-zero maze, and novel object recognition task, respectively. Administration of OBB reduced these long-term neuropsychiatric complications, and also levels of toll-like receptor 4 (TLR4), high-motility group protein 1 (HMGB1), NF-κB, tumour necrosis factor-alpha and interleukin 6 cytokines in the ipsilateral cortex of mice. Conclusion We suggest that the administration of OBB offers neuroprotective effects via inhibition of HMGB1-mediated TLR4/NFκB pathway.

Experimental traumatic brain injury increases epichaperome formation

Under normal conditions, heat shock proteins work in unison through dynamic protein interactions collectively referred to as the “chaperome.” Recent work revealed that during cellular stress, the functional interactions of the chaperome are modified to form the “epichaperome,” which results in improper protein folding, degradation, aggregation, and transport. This study is the first to investigate this novel mechanism of protein dishomeostasis in traumatic brain injury (TBI). Male and female adult, Sprague-Dawley rats received a lateral controlled cortical impact (CCI) and the ipsilateral hippocampus was collected 24 h 1, 2, and 4 weeks after injury. The epichaperome complex was visualized by measuring HSP90, HSC70 and HOP expression in native-PAGE and normalized to monomeric protein expression. A two-way ANOVA examined the effect of injury and sex at each time-point. Native HSP90, HSC70 and HOP protein expression showed a significant effect of injury effect across all time-points. Additionally, HSC70 and HOP showed significant sex effects at 24 h and 4 weeks. Altogether, controlled cortical impact significantly increased formation of the epichaperome across all proteins measured. Further investigation of this pathological mechanism can lead to a greater understanding of the link between TBI and increased risk of neurodegenerative disease and targeting the epichaperome for therapeutics.

Endothelial deletion of EPH receptor A4 alters single-cell profile and Tie2/Akap12 signaling to preserve blood–brain barrier integrity

Neurobiological consequences of traumatic brain injury (TBI) result from a complex interplay of secondary injury responses and sequela that mediates chronic disability. Endothelial cells are important regulators of the cerebrovascular response to TBI. Our work demonstrates that genetic deletion of endothelial cell (EC)-specific EPH receptor A4 (EphA4) using conditional EphA4f/f/Tie2-Cre and EphA4f/f/VE-Cadherin-CreERT2 knockout (KO) mice promotes blood-brain barrier (BBB) integrity and tissue protection, which correlates with improved motor function and cerebral blood flow recovery following controlled cortical impact (CCI) injury. scRNAseq of capillary-derived KO ECs showed increased differential gene expression of BBB-related junctional and actin cytoskeletal regulators, namely, A-kinase anchor protein 12, Akap12, whose presence at Tie2 clustering domains is enhanced in KO microvessels. Transcript and protein analysis of CCI-injured whole cortical tissue or cortical-derived ECs suggests that EphA4 limits the expression of Cldn5, Akt, and Akap12 and promotes Ang2. Blocking Tie2 using sTie2-Fc attenuated protection and reversed Akap12 mRNA and protein levels cortical-derived ECs. Direct stimulation of Tie2 using Vasculotide, angiopoietin-1 memetic peptide, phenocopied the neuroprotection. Finally, we report a noteworthy rise in soluble Ang2 in the sera of individuals with acute TBI, highlighting its promising role as a vascular biomarker for early detection of BBB disruption. These findings describe a contribution of the axon guidance molecule, EphA4, in mediating TBI microvascular dysfunction through negative regulation of Tie2/Akap12 signaling.

Neural-Cardiac Inflammasome Axis after Traumatic Brain Injury.

Traumatic brain injury (TBI) affects not only the brain but also peripheral organs like the heart and the lungs, which influences long-term outcomes. A heightened systemic inflammatory response is often induced after TBI, but the underlying pathomechanisms that contribute to co-morbidities remain poorly understood. Here, we investigated whether extracellular vehicles (EVs) containing inflammasome proteins are released after severe controlled cortical impact (CCI) in C57BL/6 mice and cause activation of inflammasomes in the heart that result in tissue damage. The atrium of injured mice at 3 days after TBI showed a significant increase in the levels of the inflammasome proteins AIM2, ASC, caspases-1, -8 and -11, whereas IL-1β was increased in the ventricles. Additionally, the injured cortex showed a significant increase in IL-1β, ASC, caspases-1, -8 and -11 and pyrin at 3 days after injury when compared to the sham. Serum-derived extracellular vesicles (EVs) from injured patients were characterized with nanoparticle tracking analysis and Ella Simple Plex and showed elevated levels of the inflammasome proteins caspase-1, ASC and IL-18. Mass spectrometry of serum-derived EVs from mice after TBI revealed a variety of complement- and cardiovascular-related signaling proteins. Moreover, adoptive transfer of serum-derived EVs from TBI patients resulted in inflammasome activation in cardiac cells in culture. Thus, TBI elicits inflammasome activation, primarily in the atrium, that is mediated, in part, by EVs that contain inflammasome- and complement-related signaling proteins that are released into serum and contribute to peripheral organ systemic inflammation, which increases inflammasome activation in the heart.

MNSCs overexpressing Rimkla transplantation facilitates cognitive recovery in a mouse model of traumatic brain injury

N-acetyl aspartyl-glutamate (NAAG) is easily inactivated for the hydrolysis of NAAG peptidase on the surface of glial cells, thereby losing its endogenous neuroprotective effect after traumatic brain injury. In this study, lentiviral vectors were used to over express/knock out NAAG synthetase II (Rimkla) in mouse embryonic neural stem cells (mNSCs) invitro and these mNSCs were transplanted at the lesion site in a mouse model of controlled cortical impact (CCI). Invivo experiments showed that transplantation of mNSCs overexpressing Rimkla regulated glutamate-glutamine cycling between adjacent astrocytes and neurons in the subacute phase of CCI, thereby enhancing support for neuronal metabolism and promoting neuronal synaptic repair in the hippocampal CA3 region. Taken together, these findings demonstrate that transplantation of neural stem cells overexpressing Rimkla can effectively increase the NAAG concentration in local brain regions, which opens up new ideas for the maintenance of NAAG neuroprotective effects after TBI.