Intranasal delivery of mitochondria targeted neuroprotective compounds for traumatic brain injury: screening based on pharmacological and physiological properties.

Targeting drugs to the mitochondrial level shows great promise for acute and chronic treatment of traumatic brain injury (TBI) in both military and civilian sectors. Perhaps the greatest obstacle to the successful delivery of drug therapies is the blood brain barrier (BBB). Intracerebroventricular and intraparenchymal routes may provide effective delivery of small and large molecule therapies for preclinical neuroprotection studies. However, clinically these delivery methods are invasive, and risk inadequate exposure to injured brain regions due to the rapid turnover of cerebral spinal fluid. The direct intranasal drug delivery approach to therapeutics holds great promise for the treatment of central nervous system (CNS) disorders, as this route is non-invasive, bypasses the BBB, enhances the bioavailability, facilitates drug dose reduction, and reduces adverse systemic effects. Using the intranasal method in animal models, researchers have successfully reduced stroke damage, reversed Alzheimer's neurodegeneration, reduced anxiety, improved memory, and delivered neurotrophic factors and neural stem cells to the brain. Based on literature spanning the past several decades, this review aims to highlight the advantages of intranasal administration over conventional routes for TBI, and other CNS disorders. More specifically, we have identified and compiled a list of most relevant mitochondria-targeted neuroprotective compounds for intranasal administration based on their mechanisms of action and pharmacological properties. Further, this review also discusses key considerations when selecting and testing future mitochondria-targeted drugs given intranasally for TBI.

Revert total protein normalization method offers a reliable loading control for mitochondrial samples following TBI.

Owing to evidence that mitochondrial dysfunction plays a dominant role in the traumatic brain injury (TBI) pathophysiology, the Western blot (WB) based immunoblotting method is widely employed to identify changes in the mitochondrial protein expressions after neurotrauma. In WB method, the housekeeping proteins (HKPs) expression is routinely used as an internal control for sample normalization. However, the traditionally employed HKPs can be susceptible to complex cascades of TBI pathogenesis, leading to their inconsistent expression. Remarkably, our data illustrated here that mitochondrial HKPs, including Voltage-dependent anion channels (VDAC), Complex-IV, Cytochrome C and Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) yielded altered expressions following penetrating TBI (PTBI) as compared to Sham. Therefore, our goal was to identify more precise normalization procedure in WB. Adult male Sprague Dawley rats (N = 6 rats/group) were used to perform PTBI, and the novel REVERT Total Protein (RTP) method was used to quantify mitochondrial protein load consistency between samples at 6 h and 24 h post-injury. Notably, the RTP method displayed superior protein normalization compared to HKPs method with higher sensitivity at both time-points between experimental groups. Our data favors application of RTP based normalization to accurately quantify protein expression where inconsistent HKPs may be evident in neuroscience research.

Multifaceted Benefit of Whole Blood Versus Lactated Ringer's Resuscitation After Traumatic Brain Injury and Hemorrhagic Shock in Mice.

BACKGROUND: Despite increasing use in hemorrhagic shock (HS), whole blood (WB) resuscitation for polytrauma with traumatic brain injury (TBI) is largely unexplored. Current TBI guidelines recommend crystalloid for prehospital resuscitation. Although WB outperforms lactated Ringer's (LR) in increasing mean arterial pressure (MAP) in TBI + HS models, effects on brain tissue oxygenation (PbtO2), and optimal MAP remain undefined. METHODS: C57BL/6 mice (n = 72) underwent controlled cortical impact followed by HS (MAP = 25-27 mmHg). Ipsilateral hippocampal PbtO2 (n = 40) was measured by microelectrode. Mice were assigned to four groups (n = 18/group) for "prehospital" resuscitation (90 min) with LR or autologous WB, and target MAPs of 60 or 70 mmHg (LR60, WB60, LR70, WB70). Additional LR (10 ml/kg) was bolused every 5 min for MAP below target. RESULTS: LR requirements in WB60 (7.2 ± 5.0 mL/kg) and WB70 (28.3 ± 9.6 mL/kg) were markedly lower than in LR60 (132.8 ± 5.8 mL/kg) or LR70 (152.2 ± 4.8 mL/kg; all p < 0.001). WB70 MAP (72.5 ± 2.9 mmHg) was higher than LR70 (59.8 ± 4.0 mmHg, p < 0.001). WB60 MAP (68.7 ± 4.6 mmHg) was higher than LR60 (53.5 ± 3.2 mmHg, p < 0.001). PbtO2 was higher in WB60 (43.8 ± 11.6 mmHg) vs either LR60 (25.9 ± 13.0 mmHg, p = 0.04) or LR70 (24.1 ± 8.1 mmHg, p = 0.001). PbtO2 in WB70 (40.7 ± 8.8 mmHg) was higher than in LR70 (p = 0.007). Despite higher MAP in WB70 vs WB60 (p = .002), PbtO2 was similar. CONCLUSION: WB resuscitation after TBI + HS results in robust improvements in brain oxygenation while minimizing fluid volume when compared to standard LR resuscitation. WB resuscitation may allow for a lower prehospital MAP without compromising brain oxygenation when compared to LR resuscitation. Further studies evaluating the effects of these physiologic benefits on outcome after TBI with HS are warranted, to eventually inform clinical trials.

Mitochondria-Targeted Antioxidant Therapeutics for Traumatic Brain Injury.

Traumatic brain injury (TBI) is a major global health problem that affects both civilian and military populations worldwide. Post-injury acute, sub-acute, and chronic progression of secondary injury processes may contribute further to other neurodegenerative diseases. However, there are no approved therapeutic options available that can attenuate TBI-related progressive pathophysiology. Recent advances in preclinical research have identified that mitochondria-centric redox imbalance, bioenergetics failure and calcium dysregulation play a crucial role in secondary injury progression after TBI. Mitochondrial antioxidants play an important role in regulating redox homeostasis. Based on the proven efficacy of preclinical and clinical compounds and targeting numerous pathways to trigger innate antioxidant defense, we may be able to alleviate TBI pathology progression by primarily focusing on preserving post-injury mitochondrial and cerebral function. In this review, we will discuss novel mitochondria-targeted antioxidant compounds, which offer a high capability of successful clinical translation for TBI management in the near future.

The game changer: UCH-L1 and GFAP-based blood test as the first marketed in vitro diagnostic test for mild traumatic brain injury.

INTRODUCTION: Major organ-based in vitro diagnostic (IVD) tests like ALT/AST for the liver and cardiac troponins for the heart are established, but an approved IVD blood test for the brain has been missing, highlighting a gap in medical diagnostics. AREAS COVERED: In response to this need, Abbott Diagnostics secured FDA clearance in 2021 for the i-STAT Alinity™, a point-of-care plasma blood test for mild traumatic brain injury (TBI). BioMerieux VIDAS, also approved in Europe, utilizes two brain-derived protein biomarkers: neuronal ubiquitin C-terminal hydrolase-L1 (UCH-L1) and glial fibrillary acidic protein (GFAP). These biomarkers, which are typically present in minimal amounts in healthy individuals, are instrumental in diagnosing mild TBI with potential brain lesions. The study explores how UCH-L1 and GFAP levels increase significantly in the bloodstream following traumatic brain injury, aiding in early and accurate diagnosis. EXPERT OPINION: The introduction of the i-STAT Alinity™ and the Biomerieux VIDAS TBI blood tests mark a groundbreaking development in TBI diagnosis. It paves the way for the integration of TBI biomarker tools into clinical practice and therapeutic trials, enhancing the precision medicine approach by generating valuable data. This advancement is a critical step in addressing the long-standing gap in brain-related diagnostics and promises to revolutionize the management and treatment of mild TBI.

Intravenous Administration of Anti-CD47 Antibody Augments Hematoma Clearance, Mitigates Acute Neuropathology, and Improves Cognitive Function in a Rat Model of Penetrating Traumatic Brain Injury.

Traumatic brain injury (TBI)-induced intracerebral hematoma is a major driver of secondary injury pathology such as neuroinflammation, cerebral edema, neurotoxicity, and blood-brain barrier dysfunction, which contribute to neuronal loss, motor deficits, and cognitive impairment. Cluster of differentiation 47 (CD47) is an antiphagocytic cell surface protein inhibiting hematoma clearance. This study was designed to evaluate the safety and efficacy of blockade of CD47 via intravenous (i.v.) administration of anti-CD47 antibodies following penetrating ballistic-like brain injury (PBBI) with significant traumatic intracerebral hemorrhage (tICH). The pharmacokinetic (PK) profile of the anti-CD47 antibody elicited that antibody concentration decayed over 7 days post-administration. Blood tests and necropsy analysis indicated no severe adverse events following treatment. Cerebral hemoglobin levels were significantly increased after injury, however, anti-CD47 antibody administration at 0.1 mg/kg resulted in a significant reduction in cerebral hemoglobin levels at 72 h post-administration, indicating augmentation of hematoma clearance. Immunohistochemistry assessment of glial fibrillary acidic protein (GFAP) and ionized calcium-binding adaptor molecule 1 (IBA1) demonstrated a significant reduction of GFAP levels in the lesion core and peri-lesional area. Based on these analyses, the optimal dose was identified as 0.1 mg/kg. Lesion volume showed a reduction following treatment. Rotarod testing revealed significant motor deficits in all injured groups but no significant therapeutic benefits. Spatial learning performance revealed significant deficits in all injured groups, which were significantly improved by the last testing day. Anti-CD47 antibody treated rats showed significantly improved attention deficits, but not retention scores. These results provide preliminary evidence that blockade of CD47 using i.v. administration of anti-CD47 antibodies may serve as a potential therapeutic for TBI with ICH.

Health Disparities in Young Adults: A Direct Comparison of Distress and Unmet Needs Across Cancer Centers.

PURPOSE: Information on concerns that young adults (YAs) with cancer face when receiving care outside of specialized treatment centers is needed to increase equitable care to YAs at greater risk of marginalization by the health care system. The current study compared distress and unmet needs at the time of clinic visit between YAs receiving care from three different cancer clinics: (1) a National Cancer Institute-designated center, (2) a community-based clinic, and (3) a county hospital outpatient clinic. METHODS: The Adolescent and Young Adult Psycho-Oncology Screening Tool (AYA-POST) was administered to measure distress and cancer-related concerns of YAs in active treatment. A one-way analysis of variance (ANOVA) compared distress scores by treatment site. A Fisher's exact test compared the number of participants endorsing each item on the Needs Assessment Checklist from each site. A simple linear regression determined the association between distress and number of items endorsed on the Needs Assessment Checklist. RESULTS: Ninety-seven participants completed the AYA-POST, endorsing, on average, 11 concerns. Fisher's exact test showed significant differences between sites in the proportion of participants endorsing eight items: boredom (P < .001), eating/appetite (P < .001), nausea/vomiting (P < .001), financial concern (P = .002), hopelessness/helplessness (P = .03), confidentiality (P = .04), sibling concern (P = .04), and insurance (P = .05). The simple linear regression model was significant (F(1, 94) = 39.772, P < .001, R2 = 0.297), indicating the number of unmet needs accounted for almost 30% of the variance in distress. The one-way ANOVA was not significant (F(2, 93) = 1.34, P = .267). CONCLUSION: Social determinants of health can influence the number and type of unmet needs experienced, affecting distress and other outcomes and underscoring the importance of timely, effective, age-appropriate screening and intervention for distress and unmet needs in YAs with cancer.

PEG hydrogel containing dexamethasone-conjugated hyaluronic acid reduces secondary injury and improves motor function in a rat moderate TBI model.

Traumatic brain injury (TBI) leads to long-term impairments in motor and cognitive function. TBI initiates a secondary injury cascade including a neuro-inflammatory response that is detrimental to tissue repair and limits recovery. Anti-inflammatory corticosteroids such as dexamethasone can reduce the deleterious effects of secondary injury; but challenges associated with dosing, administration route, and side effects have hindered their clinical application. Previously, we developed a hydrolytically degradable hydrogel (PEG-bis-AA/HA-DXM) composed of poly (ethylene) glycol-bis-(acryloyloxy acetate) (PEG-bis-AA) and dexamethasone-conjugated hyaluronic acid (HA-DXM) for local and sustained dexamethasone delivery. In this study, we evaluated the effect of locally applied PEG-bis-AA/HA-DXM hydrogel on secondary injury and motor function recovery after moderate controlled cortical impact (CCI) TBI. Hydrogel treatment significantly improved motor function evaluated by beam walk and rotarod tests compared to untreated rats over 7 days post-injury (DPI). We also observed that the hydrogel treatment reduced lesion volume, inflammatory response, astrogliosis, apoptosis, and increased neuronal survival compared to untreated rats at 7 DPI. These results suggest that PEG-bis-AA/HA-DXM hydrogels can mitigate secondary injury and promote motor functional recovery following moderate TBI.

EFFECTS OF TRANEXAMIC ACID ON NEUROPATHOLOGY, ELECTROENCEPHALOGRAPHY, AND CEREBRAL FIBRIN DEPOSITION IN A RAT MODEL OF POLYTRAUMA WITH CONCOMITANT PENETRATING TRAUMATIC BRAIN INJURY.

ABSTRACT: Several studies have demonstrated the clinical utility of tranexamic acid (TXA) for use in trauma patients presenting with significant hemorrhage. Tranexamic acid is an antifibrinolytic that inhibits plasminogen activation, and plasmin activity has been shown to mitigate blood loss and reduce all-cause mortality in the absence of adverse vascular occlusive events. Recent clinical developments indicate TXA is safe to use in patients with concomitant traumatic brain injury (TBI); however, the prehospital effects are not well understood. Importantly, TXA has been associated with seizure activity. Therefore, this study sought to evaluate the effects of early administration of TXA on neurological recovery and electroencephalogram (EEG) abnormalities following penetrating TBI with concomitant hypoxemia and hemorrhagic shock. We hypothesized that early administration of TXA will provide hemodynamic stabilization and reduce intracerebral hemorrhage, which will result in improved neurological function. To test this hypothesis, Sprague-Dawley rats received a unilateral, frontal penetrating ballistic-like brain injury by inserting a probe into the frontal cortex of the anesthetized rat. Five minutes following brain injury, animals underwent 30 min of respiratory distress and 30 min of hemorrhage. Upon completion of the hemorrhage phase, animals received the initial dose of drug intravenously over 10 min after which the prehospital phase was initiated. During the prehospital phase, animals received autologous shed whole blood as needed to maintain a MAP of 65 mm Hg. After 90 min, "in-hospital" resuscitation was performed by administering the remaining shed whole blood providing 100% oxygen for 15 min. Upon recovery from surgery, animals were administered their second dose of vehicle or TXA intravenously over 8 h. Tranexamic acid induced an early improvement in neurologic deficit, which was statistically significant compared with vehicle at 24, 48, and 72 h at three doses tested. Analysis of cerebral hemoglobin content and intracerebral lesion progression revealed 100 mg/kg provided the optimal effects for improvement of neuropathology and was continued for determination of adverse treatment effects. We observed no exacerbation of cerebral thrombosis, but TXA treatment caused an increased risk of EEG abnormalities. These results suggest that TXA following polytrauma with concomitant brain injury may provide mild neuroprotective effects by preventing lesion progression, but this may be associated with an increased risk of abnormal EEG patterns. This risk may be associated with TXA inhibition of glycine receptors and may warrant additional considerations during the use of TXA in patients with severe TBI.

Comprehensive evaluation of mitochondrial redox profile, calcium dynamics, membrane integrity and apoptosis markers in a preclinical model of severe penetrating traumatic brain injury.

Traumatic Brain Injury (TBI) is caused by the external physical assaults damages the brain. It is a heterogeneous disorder that remains a leading cause of death and disability in the military and civilian population of the United States. Preclinical investigations of mitochondrial responses in TBI have ascertained that mitochondrial dysfunction is an acute indicator of cellular damage and plays a pivotal role in long-term injury progression through cellular excitotoxicity. The current study was designed to provide an in-depth evaluation of mitochondrial endpoints with respect to redox and calcium homeostasis, and cell death responses following penetrating TBI (PTBI). To evaluate these pathological cascades, anesthetized adult male rats (N = 6/group) were subjected to either 10% unilateral PTBI or Sham craniectomy. Animals were euthanized at 24 h post-PTBI, and purified mitochondrial fractions were isolated from the brain injury core and perilesional areas. Overall, increased reactive oxygen and nitrogen species (ROS/RNS) production, and elevated oxidative stress markers such as 4-hydroxynonenal (4-HNE), 3-nitrotyrosine (3-NT), and protein carbonyls (PC) were observed in the PTBI group compared to Sham. Mitochondrial antioxidants such as glutathione, peroxiredoxin (PRX-3), thioredoxin (TRX), nicotinamide adenine dinucleotide phosphate (NADPH), superoxide dismutase (SOD), and catalase (CAT) levels were significantly decreased after PTBI. Likewise, PTBI mitochondria displayed significant loss of Ca2+ homeostasis, early opening of mitochondrial permeability transition pore (mPTP), and increased mitochondrial swelling. Both, outer and inner mitochondrial membrane integrity markers, such as voltage-dependent anion channels (VDAC) and cytochrome c (Cyt C) expression were significantly decreased following PTBI. The apoptotic cell death was evidenced by significantly decreased B-cell lymphoma-2 (Bcl-2) and increased glyceraldehyde 3-phosphate dehydrogenase (GAPDH) expression after PTBI. Collectively, current results highlight the comprehensive picture of mitochondria-centric acute pathophysiological responses following PTBI, which may be utilized as novel prognostic indicators of disease progression and theragnostic indicators for evaluating neuroprotection therapeutics following TBI.

Upregulation of multiple toll-like receptors in ferret brain after blast exposure: Potential targets for treatment.

Although blast-induced traumatic brain injury (bTBI) has been designated as the signature injury of recent combat operations, its precise pathological mechanism(s) has not been identified thus far. Prior preclinical studies on bTBI demonstrated acute neuroinflammatory cascades which are known to be contributing to neurodegeneration. Danger-associated chemical patterns are released from the injured cells, which activate non-specific pattern recognition receptors, such as toll-like receptors (TLRs) leading to increased expression of inflammatory genes and release of cytokines. Upregulation of specific TLRs in the brain has been described as a mechanism of injury in diverse brain injury models unrelated to blast exposure. However, the expression profile of various TLRs in bTBI has not been investigated thus far. Hence, we have evaluated the expression of transcripts for TLR1-TLR10 in the brain of a gyrencephalic animal model of bTBI. We exposed ferrets to tightly coupled repeated blasts and determined the differential expression of TLRs (TLR1-10) by quantitative RT-PCR in multiple brain regions at 4 hr, 24 hr, 7 days and 28 days post-blast injury. The results obtained indicate that multiple TLRs are upregulated in the brain at 4 hr, 24 hr, 7 days and 28 days post-blast. Specifically, upregulation of TLR2, TLR4 and TLR9 was noted in different brain regions, suggesting that multiple TLRs might play a role in the pathophysiology of bTBI and that drugs that can inhibit multiple TLRs might have enhanced efficacy to attenuate brain damage and thereby improve bTBI outcome. Taken together, these results suggest that several TLRs are upregulated in the brain after bTBI and participate in the inflammatory response and thereby provide new insights into the disease pathogenesis. Therefore, inhibition of multiple TLRs, including TLR2, 4 and 9, simultaneously might be a potential therapeutic strategy for the treatment of bTBI.

Amygdala functional connectivity mediates the association between catastrophizing and threat-safety learning in youth with chronic pain.

ABSTRACT: There is a need to identify brain connectivity alterations predictive of transdiagnostic processes that may confer vulnerability for affective symptomology. Here, we tested whether amygdala resting-state functional connectivity (rsFC) mediated the relationship between catastrophizing (negative threat appraisals and predicting poorer functioning) and altered threat-safety discrimination learning (critical to flexibly adapt to new and changing environments) in adolescents with persistent pain. We examined amygdala rsFC in 46 youth with chronic pain and 29 healthy peers (age M = 15.8, SD = 2.9; 64 females) and its relationship with catastrophizing and threat-safety learning. We used a developmentally appropriate threat-safety learning paradigm and performed amygdala seed-based rsFC and whole-brain mediation analyses. Patients exhibited enhanced connectivity between the left amygdala and right supramarginal gyrus (SMG) (cluster-level P-FDR < 0.05), whereas right amygdala rsFC showed no group differences. Only in patients, elevated catastrophizing was associated with facilitated threat-safety learning (CS+>CS-; rp = 0.49, P = 0.001). Furthermore, in patients, elevated catastrophizing was associated with reduced left amygdala connectivity with SMG / parietal operculum, and increased left amygdala connectivity with hippocampus, dorsal striatum, paracingulate, and motor regions (P < 0.001). In addition, blunted left amygdala rsFC with right SMG/parietal operculum mediated the association between catastrophizing and threat-safety learning (P < 0.001). To conclude, rsFC between the left amygdala (a core emotion hub) and inferior parietal lobe (involved in appraisal and integration of bodily signals and attentional reorienting) explains associations between daily-life relevant catastrophizing and threat-safety learning. Findings provide a putative model for understanding pathophysiology involved in core psychological processes that cut across diagnoses, including disabling pain, and are relevant for their etiology.

Rapid Transition to Virtual Assessment and Treatment in an Interdisciplinary Randomized Clinical Trial for Youth With Chronic Pain: Adaptations and Implications for Future Trials.

OBJECTIVES: COVID-19 abruptly halted in-person clinical care and research requiring a shift to virtual assessment and treatment. This unexpected transition of a 2-arm randomized controlled trial (RCT) examining interdisciplinary graded exposure treatment (GET Living) compared with multidisciplinary pain management for youth with chronic pain provided an opportunity to implement the first remotely delivered exposure treatment and remotely delivered biomechanical assessment for pediatric chronic pain. Here we describe these new approaches and provide lessons learned to inform future efforts in digital health care. METHODS: A total of 68 youth (M=14.2 y; 80.9% female) were enrolled in the RCT (n=31 in-person, n=5 hybrid, n=32 virtual, n=9 withdrew). Of those withdrawn, n=3 withdrew due to COVID-19 related reasons. Some RCT elements required slight modification (eg, e-consent, actigraphy deployment, recruitment, and screening), while others were significantly altered (eg, session format and lab-based biomechanical assessment). Data from exit interviews were also examined to assess perspectives on the virtual format transition. RESULTS: Results showed an increased enrollment rate when virtual care was an option (70.7%) compared with in-person (44.3%). Equivalent rates of completion for daily assessment (in-person, 72.8%; virtual, 73.3) were also observed, and participants described enhanced experience when able to complete exercises and exposures in their home environment during session (vs. a rehabilitation gym) allowing for genuine in vivo exposures (eg, household chores, riding bicycles). DISCUSSION: Overall, our data demonstrate acceptability, feasibility, and equivalent patient engagement to virtual treatment. Novel methods implemented in this RCT can inform trial design and measures of clinical endpoints for future digital health interventions.

Mitoquinone Helps Combat the Neurological, Cognitive, and Molecular Consequences of Open Head Traumatic Brain Injury at Chronic Time Point.

Traumatic brain injury (TBI) is a heterogeneous disease in its origin, neuropathology, and prognosis, with no FDA-approved treatments. The pathology of TBI is complicated and not sufficiently understood, which is the reason why more than 30 clinical trials in the past three decades turned out unsuccessful in phase III. The multifaceted pathophysiology of TBI involves a cascade of metabolic and molecular events including inflammation, oxidative stress, excitotoxicity, and mitochondrial dysfunction. In this study, an open head TBI mouse model, induced by controlled cortical impact (CCI), was used to investigate the chronic protective effects of mitoquinone (MitoQ) administration 30 days post-injury. Neurological functions were assessed with the Garcia neuroscore, pole climbing, grip strength, and adhesive removal tests, whereas cognitive and behavioral functions were assessed using the object recognition, Morris water maze, and forced swim tests. As for molecular effects, immunofluorescence staining was conducted to investigate microgliosis, astrocytosis, neuronal cell count, and axonal integrity. The results show that MitoQ enhanced neurological and cognitive functions 30 days post-injury. MitoQ also decreased the activation of astrocytes and microglia, which was accompanied by improved axonal integrity and neuronal cell count in the cortex. Therefore, we conclude that MitoQ has neuroprotective effects in a moderate open head CCI mouse model by decreasing oxidative stress, neuroinflammation, and axonal injury.

Mitoquinone supplementation alleviates oxidative stress and pathologic outcomes following repetitive mild traumatic brain injury at a chronic time point.

Traumatic brain injury (TBI) is a major cause of disability and death. Mild TBI (mTBI) constitutes ~75% of all TBI cases. Repeated exposure to mTBI (rmTBI), leads to the exacerbation of the symptoms compared to single mTBI. To date, there is no FDA-approved drug for TBI or rmTBI. This research aims to investigate possible rmTBI neurotherapy by targeting TBI pathology-related mechanisms. Oxidative stress is partly responsible for TBI/rmTBI neuropathologic outcomes. Thus, targeting oxidative stress may ameliorate TBI/rmTBI consequences. In this study, we hypothesized that mitoquinone (MitoQ), a mitochondria-targeted antioxidant, would ameliorate TBI/rmTBI associated pathologic features by mitigating rmTBI-induced oxidative stress. To model rmTBI, C57BL/6 mice were subjected to three concussive head injuries. MitoQ (5 mg/kg) was administered intraperitoneally to rmTBI+MitoQ mice twice per week over one month. Behavioral and cognitive outcomes were assessed, 30 days following the first head injury, using a battery of behavioral tests. Immunofluorescence was used to assess neuroinflammation and neuronal integrity. Also, qRT-PCR was used to evaluate the expression levels of antioxidant enzymes. Our findings indicated that MitoQ alleviated fine motor function and learning impairments caused by rmTBI. Mechanistically, MitoQ reduced astrocytosis, microgliosis, dendritic and axonal shearing, and increased the expression of antioxidant enzymes. MitoQ administration following rmTBI may represent an efficient approach to ameliorate rmTBI neurological and cellular outcomes with no observable side effects.

Time-Course Evaluation of Brain Regional Mitochondrial Bioenergetics in a Pre-Clinical Model of Severe Penetrating Traumatic Brain Injury.

Mitochondrial dysfunction is a pivotal target for neuroprotection strategies for traumatic brain injury (TBI). However, comprehensive time-course evaluations of mitochondrial dysfunction are lacking in the pre-clinical penetrating TBI (PTBI) model. The current study was designed to characterize temporal responses of mitochondrial dysfunction from 30 min to 2 weeks post-injury after PTBI. Anesthetized adult male rats were subjected to either PTBI or sham craniectomy (n = 6 animals per group × 7 time points). Animals were euthanized at 30 min, 3 h, 6 h, 24 h, 3 days, 7 days, and 14 days post-PTBI, and mitochondria were isolated from the ipsilateral hemisphere of brain regions near the injury core (i.e., frontal cortex [FC] and striatum [ST]) and a more distant region from the injury core (i.e., hippocampus [HIP]). Mitochondrial bioenergetics parameters were measured in real time using the high-throughput procedures of the Seahorse Flux Analyzer (Agilent Technologies, Santa Clara, CA). The post-injury time course of FC + ST showed a biphasic mitochondrial bioenergetics dysfunction response, indicative of reduced adenosine triphosphate synthesis rate and maximal respiratory capacity after PTBI. An initial phase of energy crisis was detected at 30 min (-42%; p < 0.05 vs. sham), which resolved to baseline levels between 3 and 6 h (non-significant vs. sham). This was followed by a second and more robust phase of bioenergetics dysregulation detected at 24 h that remained unresolved out to 14 days post-injury (-55% to -90%; p < 0.05 vs. sham). In contrast, HIP mitochondria showed a delayed onset of mitochondrial dysfunction at 7 days (-74%; p < 0.05 vs. sham) that remained evident out to 14 days (-51%; p < 0.05 vs. sham) post-PTBI. Collectively, PTBI-induced mitochondrial dysfunction responses were time and region specific, evident differentially at the injury core and distant region of PTBI. The current results provide the basis that mitochondrial dysfunction may be targeted differentially based on region specificity post-PTBI. Even more important, these results suggest that therapeutic interventions targeting mitochondrial dysfunction may require extended dosing regimens to achieve clinical efficacy after TBI.

Penetrating Ballistic Brain Injury Produces Acute Alterations in Sleep and Circadian-Related Genes in the Rodent Cortex: A Preliminary Study.

Traumatic brain injury (TBI) affects millions of Americans each year, with extremely high prevalence in the Veteran community, and sleep disturbance is one of the most commonly reported symptoms. Reduction in the quality and amount of sleep can negatively impact recovery and result in a wide range of behavioral and physiological symptoms, such as impaired cognition, mood and anxiety disorders, and cardiovascular effects. Thus, to improve long-term patient outcomes and develop novel treatments, it is essential to understand the molecular mechanisms involved in sleep disturbance following TBI. In this effort, we performed transcriptional profiling in an established rodent model of penetrating ballistic brain injury (PBBI) in conjunction with continuous sleep/wake EEG/EMG recording of the first 24 h after injury. Rats subjected to PBBI showed profound differences in sleep architecture. Injured animals spent significantly more time in slow wave sleep and less time in REM sleep compared to sham control animals. To identify PBBI-related transcriptional differences, we then performed transcriptome-wide gene expression profiling at 24 h post-injury, which identified a vast array of immune- related genes differentially expressed in the injured cortex as well as sleep-related genes. Further, transcriptional changes associated with total time spent in various sleep stages were identified. Such molecular changes may underlie the pathology and symptoms that emerge following TBI, including neurodegeneration, sleep disturbance, and mood disorders.

Prehospital Whole Blood Resuscitation Reduces Fluid Requirement While Maintaining Critical Physiology in a Model of Penetrating Traumatic Brain Injury and Hemorrhage: Implications on Resource-Limited Combat Casualty Care.

ABSTRACT: Prehospital resuscitation using whole blood (WB) is the standard of care for hemorrhagic shock (HS) but there is no consensus recommendation for resuscitation in the presence of traumatic brain injury (TBI) due to a lack of sufficient evidence. In order to evaluate the optimal resuscitation strategies for TBI+HS, Sprague-Dawley rats were randomized into four groups based on resuscitation fluid and prehospital mean arterial pressure (MAP) threshold (n = 9-10/group): Lactated Ringer's (LR)-60 mm Hg (LR60), LR-70 mm Hg (LR70), WB-60 mm Hg (WB60), WB-70 mm Hg (WB70). All groups received a frontal penetrating ballistic-like brain injury followed by a 35-min period of HS. During the prehospital phase, rats received an initial bolus of resuscitation fluid (WB or LR) followed by LR as needed to maintain MAP above the designated threshold for 90 min. During the in-hospital phase, rats received definitive resuscitation with shed WB. Physiological parameters were recorded continuously and cerebral edema was measured at 3 and 24 h postinjury. The WB60 group demonstrated a significantly lower prehospital fluid requirement compared WB70, LR60, and LR70 (P < 0.05). Compared to the respective LR groups, both the WB60 and WB70 groups also demonstrated improved MAP, cerebral perfusion pressure, brain tissue oxygen tension, and cerebral edema. The edema benefits were observed at 3 h, but not 24 h postinjury, and were localized to the injury site. Together, these results provide evidence that prehospital WB resuscitation and lower MAP resuscitation thresholds can reduce the prehospital fluid requirement while still maintaining critical cerebral physiology in a model of HS and concomitant TBI.

Choice of Whole Blood versus Lactated Ringer's Resuscitation Modifies the Relationship between Blood Pressure Target and Functional Outcome after Traumatic Brain Injury plus Hemorrhagic Shock in Mice.

Civilian traumatic brain injury (TBI) guidelines recommend resuscitation of patients with hypotensive TBI with crystalloids. Increasing evidence, however, suggests that whole blood (WB) resuscitation may improve physiological and survival outcomes at lower resuscitation volumes, and potentially at a lower mean arterial blood pressure (MAP), than crystalloid after TBI and hemorrhagic shock (HS). The objective of this study was to assess whether WB resuscitation with two different MAP targets improved behavioral and histological outcomes compared with lactated Ringer's (LR) in a mouse model of TBI+HS. Anesthetized mice (n = 40) underwent controlled cortical impact (CCI) followed by HS (MAP = 25-27 mm Hg; 25 min) and were randomized to five groups for a 90 min resuscitation: LR with MAP target of 70 mm Hg (LR70), LR60, WB70, WB60, and monitored sham. Mice received a 20 mL/kg bolus of LR or autologous WB followed by LR boluses (10 mL/kg) every 5 min for MAP below target. Shed blood was reinfused after 90 min. Morris Water Maze testing was performed on days 14-20 post-injury. Mice were euthanized (21 d) to assess contusion and total brain volumes. Latency to find the hidden platform was greater versus sham for LR60 (p < 0.002) and WB70 (p < 0.007) but not LR70 or WB60. The WB resuscitation did not reduce contusion volume or brain tissue loss. The WB targeting a MAP of 60 mm Hg did not compromise function versus a 70 mm Hg target after CCI+HS, but further reduced fluid requirements (p < 0.03). Using LR, higher achieved MAP was associated with better behavioral performance (rho = -0.67, p = 0.028). Use of WB may allow lower MAP targets without compromising functional outcome, which could facilitate pre-hospital TBI resuscitation.