Diffuse Traumatic Brain Injury Research Articles

Risk factors of aseptic bone resorption: a study after autologous bone flap reinsertion due to decompressive craniotomy

In patients who have undergone decompressive craniectomy, autologous bone flap reinsertion becomes necessary whenever the cerebral situation has consolidated. However, aseptic necrosis of the bone flap remains a concern. The aim of this study was to report possible perioperative complications in patients undergoing autologous bone flap reinsertion and to identify the risk factors that may predispose the bone flap to necrosis. All patients admitted to the authors' neurosurgical department between September 1994 and June 2011 and who received their own cryoconserved bone flap after decompressive craniectomy were studied. The grade of the bone flap necrosis was classified into 2 types. Type II bone necrosis was characterized by aseptic resorption with circumscribed or complete lysis of tabula interna and externa requiring surgical revision. To define predisposing factors, a multivariate analysis was performed using bone necrosis as the dependent variable. Among the 372 patients (mean age 48.6 years, 57.4% males) who received 414 bone flaps during the observation period, 134 (36.0%) had a diffuse traumatic brain injury, 69 (18.5%) had subarachnoid hemorrhage, 58 (15.6%) had cerebral infarction, 56 (15.1%) had extraaxial bleeding, 43 (11.6%) had intracerebral bleeding, and 12 (3.2%) had a neoplasm. Surgical relevant Type II bone flap necrosis occurred in 85 patients (22.8%) and 91 bone flaps, after a median time of 15 months (interquartile range [IQR], 10-33 months). In a multivariate analysis with Type II necrosis as the dependent variable, bone flap fragmentation with 2 (OR 3.35, 95% CI 1.59-7.01, p < 0.002) or more fragments (OR 24.00, 95% CI 10.13-56.84, p < 0.001), shunt-dependent hydrocephalus (OR 1.76, 95% CI 0.99-3.12, p = 0.04), and a younger age (OR 0.98, 95% CI 0.96-0.99, p = 0.004) was associated with a higher risk for the development of an aseptic bone flap necrosis. In patients undergoing bone flap reinsertion after craniotomy, aseptic bone necrosis is an underestimated problem during long-term follow-up. Especially in younger patients with an expected good neurological recovery and a fragmented bone flap, an initial allograft should be considered because of an increased risk for aseptic bone flap necrosis.

The effects of lornoxicam on brain edema and blood brain barrier following diffuse traumatic brain injury in rats

In this experiment, the effects of lornoxicam on brain edema and the blood brain barrier (BBB) following diffuse traumatic brain injury (TBI) were studied. Twenty adult male Wistar albino rats were anesthetized, and experimental closed head trauma was induced by the Marmarou method. After head injury, the rats were randomly divided into two groups: Group I was the control group, to which 2 ml saline was administered intraperitoneally, and Group II was the lornoxicam group, to which 2 ml 1.3 mg kg-1 lornoxicam was administered intraperitoneally. Twenty-four hours after head trauma, 99 mTc pentetate (DTPA) was injected at a dose of 37 MBq, and posterior planar images of each rat were obtained using an Infinia gamma camera. After imaging of BBB permeability, brain tissues were dissected from the cranium. The brain water content (BWC) of each sample was calculated using the wet-dry method. The lesion/background (L/b) ratio of Group I was 3.76±0.46 and 3.02±0.66 for early (5th min) and late (60th min) imaging, respectively. In Group II, the L/b ratios were 3.52±0.96 and 2.63±0.63 for early and late imaging, respectively (p>0.05). BWC was 79.6±2.5% and 77.5±1.1% for Groups I and II, respectively (p<0.05). In this rat model of TBI, lornoxicam reduced brain edema but did not affect BBB permeability.

The effects of lornoxicam on neuroprotection following diffuse traumatic brain injury in rats

In this study, the effects of lornoxicam on the prevention of secondary brain injury via the apoptotic pathway were studied in a rat model of head injury. Thirty adult male Wistar albino rats were anesthetized, and experimental closed head trauma was induced by allowing a 450 g weight to fall two meters onto a metallic disk fixed to the intact skull. After head injury, the rats were randomly divided into two groups: Group I (n=15) rats were administered 2 mL saline intraperitoneally (controls); Group II (n=15) rats were administered 2 mL 1.3 mg kg-1 lornoxicam intraperitoneally. Brain tissue samples were divided into two pieces by interhemispheric incision for biochemical and histological analysis. TUNEL positivity was seen in neuroglia cells of the brain cortex in both groups. While the immunoreactivities of caspase 8, 9 and Fas/ Fas ligand were similar in both groups, the immunoreactivity of caspase 3 was greater in Group I than Group II. MDA was significantly lower in Group II than in Group I (p < 0.05). The decrease in SOD level was higher in Group I than Group II. Lornoxicam did not prevent apoptosis in this rat model of brain trauma but causes a decrease.

Differences in brain edema and intracranial pressure following traumatic brain injury across the estrous cycle: Involvement of female sex steroid hormones

It has been shown that sex steroid hormones have profound neuroprotective effects in experimental traumatic brain injury (TBI). Because the endogenous hormone levels are proven to differ with estrous cycle stage, we evaluated whether estrous cycle stage affects various outcomes following diffuse TBI. TBI was induced by Marmarou's method in normal cycling and in ovariectomized rats with physiologically relevant restoration of hormonal levels by hormone capsule implantation. Intracranial pressure (ICP) and cerebral perfusion pressure (CPP) were measured before and different times after TBI and brain edema was assessed at 24h after trauma. Results indicated that after TBI, water content (WC) in traumatic proestrous (TP) rats was less than the one in traumatic non-proestrous (TNP) and ovariectomized (TOVX) and also in high estradiol (HE) and progesterone (HP) was statistically less than in TBI untreated groups.There was no significant difference in WC between high doses hormone treated and TP and also between TNP, TOVX, low estradiol (LE) and progesterone (LP) groups. At 4h and 24h after trauma, there was a significant difference in ICP between TP, HE and HP compared to TNP and other TBI nontreated groups. Also in these times, the CPP increased in TP and hormone treated groups compared with TOVX, but the difference between TNP and TOVX was not significant. The results indicate that the estrous cycle has a prominent role in TBI outcome's and the difference in female sex steroid levels might be the reason of the different neuroprotective effects in proestrous and non-proestrous groups.

Association between reduced expression of hippocampal glucocorticoid receptors and cognitive dysfunction in a rat model of traumatic brain injury due to lateral head acceleration

Expression of hippocampal glucocorticoid receptors (GRs) and spatial learning and memory were observed in rat model of diffuse traumatic brain injury (TBI) due to lateral head acceleration with an aim at investigating the relation between GRs expression and cognitive deficits. Immunohistochemical staining, Western blotting, and RT-PCR indicated that down-regulation of GRs expression occurred in the hippocampus among TBI-rats which demonstrated reduced performance of learning and memory in Morris water maze. As the GRs expression bounced up, the cognitive function approached to normal. It is concluded that reduced expression of hippocampal GRs was closely associated with learning and memory deficits in TBI-rats. Hippocampal GRs was involved in the biochemical mechanisms of cognitive deficits after TBI.

The effects of Zinc Protoporphyrin and Heme on the expression of HO-1 in cochlear and the change of auditory brainstem response on diffuse traumatic brain injury model of rats

Objective To explore the effects of Zinc Protoporphyrin and Heme on the expression of HO-1 in cochlear and the change of auditory brainstem response on diffuse traumatic brain injury model of rats.Methods Diffuse traumatic brain injury model of rats were established and randomly divided into thirteen groups.Then auditory brainstem response examination,light microscope,immunohistochemistry technique were used to evaluate the change of auditory brainstem response and the expression of HO-1 in cochlear.Results The differences of auditory brainstem response threshold and latency of wave between the experimental and the normal control group were obvious(P＜0.05).The expression of HO-1 in the control group was normal,whereas there were obvious changes of inner ear HO-1 expression in the traumatic groups.The grey value of HO-1 expression in trauma group,Znpp group and heme group was significantly associated with auditory function change(P＜0.05).Conclusion There were influence of Zinc Protoporphyrin and Hemeon the inner ear HO-1 expression and the change of auditory brainstem response with diffuse traumatic brain injury model of rats.The protective effect of heme on auditory function may be associated with the increased expression of HO-1. Key words: Diffuse traumatic brain injury; Zinc Protoporphyrin; Heme; Cochlear; Heme oxygenase; Auditory Brainstem Response

The DECRA Trial and Decompressive Craniectomy in Diffuse Traumatic Brain Injury: Is Decompression Really Ineffective?

To assess quality of life of patients who underwent decompressive craniectomy (DC) for traumatic brain injury and satisfaction of caregivers with outcomes.This cross-sectional study was conducted at a tertiary care urban center in Pakistan. All patients with severe traumatic brain injury who underwent DC and survived >6 months were included. Outcomes were assessed using 2 scales: Glasgow Outcome Scale Extended and Quality of Life After Traumatic Brain Injury (QOLIBRI). The proforma was translated and validated into the national language. Patient caregivers were interviewed to ask if they would opt for DC in a similar situation in future.The study comprised 40 patients, including 35 male (88%) and 5 female (12%) patients. Mean age of patients was 26.5 ± 9.5 years. Mean Glasgow Coma Scale score at presentation was 8.34 ± 3.22. Median follow-up was 12 months (range, 6–18 months). Mean Glasgow Outcome Scale Extended score was 5.35 ± 1.9, which correlates with an unfavorable outcome. Mean QOLIBRI score was 59.65 ± 21.27. Family members of 38 (95%) patients were content with their decision to give consent for DC in their patients. Spearman correlation for different domains of QOLIBRI and Glasgow Outcome Scale Extended was statistically significant for all parameters except social relationship.Mean QOLIBRI score of patients undergoing DC was 59.65 ± 21.27. Most caregivers (95%) were satisfied with their decision to consent for DC. Patient-reported health-related quality of life assessment is necessary to assess impact of traumatic brain injury.

Rod microglia: elongation, alignment, and coupling to form trains across the somatosensory cortex after experimental diffuse brain injury

BackgroundSince their discovery, the morphology of microglia has been interpreted to mirror their function, with ramified microglia constantly surveying the micro-environment and rapidly activating when changes occur. In 1899, Franz Nissl discovered what we now recognize as a distinct microglial activation state, microglial rod cells (Stäbchenzellen), which he observed adjacent to neurons. These rod-shaped microglia are typically found in human autopsy cases of paralysis of the insane, a disease of the pre-penicillin era, and best known today from HIV-1-infected brains. Microglial rod cells have been implicated in cortical ‘synaptic stripping’ but their exact role has remained unclear. This is due at least in part to a scarcity of experimental models. Now we have noted these rod microglia after experimental diffuse brain injury in brain regions that have an associated sensory sensitivity. Here, we describe the time course, location, and surrounding architecture associated with rod microglia following experimental diffuse traumatic brain injury (TBI).MethodsRats were subjected to a moderate midline fluid percussion injury (mFPI), which resulted in transient suppression of their righting reflex (6 to 10 min). Multiple immunohistochemistry protocols targeting microglia with Iba1 and other known microglia markers were undertaken to identify the morphological activation of microglia. Additionally, labeling with Iba1 and cell markers for neurons and astrocytes identified the architecture that surrounds these rod cells.ResultsWe identified an abundance of Iba1-positive microglia with rod morphology in the primary sensory barrel fields (S1BF). Although present for at least 4 weeks post mFPI, they developed over the first week, peaking at 7 days post-injury. In the absence of contusion, Iba1-positive microglia appear to elongate with their processes extending from the apical and basal ends. These cells then abut one another and lay adjacent to cytoarchitecture of dendrites and axons, with no alignment with astrocytes and oligodendrocytes. Iba1-positive rod microglial cells differentially express other known markers for reactive microglia including OX-6 and CD68.ConclusionDiffuse traumatic brain injury induces a distinct rod microglia morphology, unique phenotype, and novel association between cells; these observations entice further investigation for impact on neurological outcome.

Attenuation of brain edema and spatial learning deficits by the inhibition of NADPH oxidase activity using apocynin following diffuse traumatic brain injury in rats

Diffuse brain injury (DBI) is a leading cause of mortality and disability among young individuals and adults worldwide. In specific cases, DBI is associated with permanent spatial learning dysfunction and motor deficits due to primary and secondary brain damage. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) is a major complex that produces reactive oxygen species (ROS) during the ischemic period. The complex aggravates brain damage and cell death following ischemia/reperfusion injury; however, its role in DBI remains unclear. The present study aimed to investigate the hypothesis that levels of NOX2 (a catalytic subunit of NOX) protein expression and the activation of NOX are enhanced following DBI induction in rats and are involved in aggravating secondary brain damage. A rat model of DBI was created using a modified weight-drop device. Our results demonstrated that NOX2 protein expression and NOX activity were enhanced in the CA1 subfield of the hippocampus at 48 and 72h following DBI induction. Treatment with apocynin (50mg/kg body weight), a specific inhibitor of NOX, injected intraperitoneally 30min prior to DBI significantly attenuated NOX2 protein expression and NOX activation. Moreover, treatment with apocynin reduced brain edema and improved spatial learning function assessed using the Morris water maze. These results reveal that treatment with apocynin may provide a new neuroprotective therapeutic strategy against DBI by diminishing the upregulation of NOX2 protein and NOX activity.

Neuronal damage and functional deficits are ameliorated by inhibition of aquaporin and HIF1α after traumatic brain injury (TBI)

The present study, using a rodent model of closed-head diffuse traumatic brain injury (TBI), investigated the role of dysregulated aquaporins (AQP) 4 and 9, as well as hypoxia inducible factor −1α(HIF-1α) on brain edema formation, neuronal injury, and functional deficits.TBI was induced in adult (400–425g), male Sprague–Dawley rats using a modified Marmarou's head impact-acceleration device (450g weight dropped from 2m height). Animals in each treatment group were administered intravenous anti-AQP4 or -AQP9 antibodies or 2-Methoxyestradiol (2ME2, an inhibitor of HIF-1α) 30min after injury. At 24h post-TBI, animals (n=6 each group) were sacrificed to examine the extent of brain edema by water content, as well as protein expression of AQP and HIF-1α by Western immune-blotting. At 48-hours post-TBI, neuronal injury (n=8 each group) was assessed by FluoroJade (FJ) histochemistry. Spatial learning and memory deficits were evaluated by radial arm maze (n=8 each group) up to 21days post-TBI.Compared to non-injured controls, significant (p<0.05) increases in the expression of AQP4 and −9 were detected in the brains of injured animals. In addition, significant (p<0.05) brain edema after TBI was associated with increases (p <0.05) both in neuronal injury (FJ labeling) and neurobehavioral deficits. Selective inhibition of either AQP4 or −9, or HIF-1α significantly (p<0.05) decreased the expression of the proteins. In addition, inhibition of the AQPs and HIF-1α significantly (p<0.05) ameliorated brain edema, as well as the number of injured neurons in cortical layers II/III and V/VI, striatum and hippocampal regions CA1/CA3. Finally, compared to the non-treated TBI animals, AQP or HIF-1α inhibition significantly (p<0.01) improved neurobehavioral outcomes after TBI.Taken together, the present data supports a causal relation between HIF-AQP mediated cerebral edema, secondary neuronal injury, and tertiary behavioral deficits post-TBI. The data further suggests that upstream modulation of the molecular patho-trajectory effectively ameliorates both neuronal injury and behavioral deficits post-TBI.

DECRA Investigators' Response to “The Future of Decompressive Craniectomy for Diffuse Traumatic Brain Injury” by Honeybul et al.

DECRA Investigators' Response to “The Future of Decompressive Craniectomy for Diffuse Traumatic Brain Injury” by Honeybul et al.

Detection of hemorrhagic and axonal pathology in mild traumatic brain injury using advanced MRI: Implications for neurorehabilitation

There is a need to more accurately diagnose milder traumatic brain injuries with increasing awareness of the high prevalence in both military and civilian populations. Magnetic resonance imaging methods may be capable of detecting a number of the pathoanatomical and pathophysiological consequences of focal and diffuse traumatic brain injury. Susceptibility-weighted imaging (SWI) detects heme iron and reveals even small venous microhemorrhages occurring in diffuse vascular injury. Diffusion tensor imaging (DTI) reveals axonal injury by detecting alterations in water flow in and around injured axons. The overarching hypothesis of this paper is that newer, advanced MR imaging generates sensitive biomarkers of regional brain injury which allows for correlation with clinical signs and symptoms. Studies involving subjects with a history of traumatic brain injury as well as healthy, non-trauma controls were used. Analysis involved comparison of TBI patients' imaging results with healthy controls as well as correlation of imaging findings with clinical measures of injury severity. An additional animal study of Sprague-Dawley albino rats compared imaging results with histopathological findings after the animals were sacrificed and stained for b-APP. SWI revealed small foci of hemosiderin for some patients while aggregate lesion volume on SWI correlated with clinical injury severity indices. Similarly, DTI showed striking group differences for fractional anisotropy over the white matter globally, while tract and voxel-based regional results colocalized with SWI and FLAIR lesions in some cases and correlated with clinical deficits. For the rats, correlations were seen between imaging findings and staining of axonal injury. Animal data gave important tissue correlations with imaging results. SWI and DTI are commercially available sequences that can improve the diagnostic and prognostic ability of the trauma clinician. These biomarkers of regional brain injury which are present in imaging shortly after acute injury and persist indefinitely can inform clinicians and researchers about not only injury severity but also which neurobehavioral systems were injured. Analogous to stroke rehabilitation, having an understanding of the distribution of brain injury should ultimately allow for development of more effective rehabilitation strategies and more efficient clinical interventional trials.

Morphological and genetic activation of microglia after diffuse traumatic brain injury in the rat

Traumatic brain injury (TBI) survivors experience long-term post-traumatic morbidities. In diffuse brain-injured rats, a chronic sensory sensitivity to whisker stimulation models the agitation of TBI survivors and provides anatomical landmarks across the whisker-barrel circuit to evaluate post-traumatic neuropathology. As a consequence of TBI, acute and chronic microglial activation can contribute to degenerative and reparative events underlying post-traumatic morbidity. Here we hypothesize that a temporal sequence of microglial activation states contributes to the circuit pathology responsible for post-traumatic morbidity, and test the hypothesis by examining microglial morphological activation and neuroinflammatory markers for activation states through gene expression and receptor-binding affinity. Adult male, Sprague–Dawley rats were subjected to a single moderate midline fluid percussion injury (FPI) or sham injury. Microglial activation was determined by immunohistochemistry, quantitative real-time PCR and receptor autoradiography in the primary somatosensory barrel field (S1BF) and ventral posterior medial nucleus (VPM) of the thalamus at 7 and 28days following FPI. Morphological changes indicative of microglial activation, including swollen cell body with thicker, shrunken processes, were evident in S1BF and VPM at 7 and 28days post-injury. Principally at 7days post-injury in VPM, general inflammatory gene expression (major histocompatibility complex I, major histocompatibility complex II, translocator protein 18kDa [TSPO]) is increased above sham level and TSPO gene expression confirmed by receptor autoradiography. Further, CD45, a marker of classical activation, and TGF-βI, an acquired deactivation marker, were elevated significantly above sham at 7days post-injury. Daily administration of the anti-inflammatory ibuprofen (20mg/kg, i.p.) significantly reduced the expression of these genes. Evidence for alternative activation (arginase 1) was not observed. Thus, these data demonstrate concomitant classical activation and acquired deactivation phenotypes of microglia in diffuse TBI in the absence of overt contusion or cavitation. Anti-inflammatory treatment may further alleviate the neuropathological burden of post-traumatic inflammation.

Early cerebral perfusion pressure augmentation with phenylephrine after traumatic brain injury may be neuroprotective in a pediatric swine model*

Cerebral perfusion pressure<40 mm Hg following pediatric traumatic brain injury has been associated with increased mortality independent of age, and current guidelines recommend maintaining cerebral perfusion pressure between 40 mm Hg-60 mm Hg. Although adult traumatic brain injury studies have observed an increased risk of complications associated with targeting a cerebral perfusion pressure>70, we hypothesize that targeting a cerebral perfusion pressure of 70 mm Hg with the use of phenylephrine early after injury in the immature brain will be neuroprotective. Animals were randomly assigned to injury with a cerebral perfusion pressure of 70 mm Hg or 40 mm Hg. Diffuse traumatic brain injury was produced by a single rapid rotation of the head in the axial plane. Cerebral microdialysis, brain tissue oxygen, intracranial pressure, and cerebral blood flow were measured 30 min-6 hrs postinjury. One hour after injury, cerebral perfusion pressure was manipulated with the vasoconstrictor phenylephrine. Animals were euthanized 6 hrs posttraumatic brain injury, brains fixed, and stained to assess regions of cell injury and axonal dysfunction. University center. Twenty-one 4-wk-old female swine. Augmentation of cerebral perfusion pressure to 70 mm Hg resulted in no change in axonal dysfunction, but significantly smaller cell injury volumes at 6 hrs postinjury compared to cerebral perfusion pressure 40 (1.1% vs. 7.4%, p<.05). Microdialysis lactate/pyruvate ratios were improved at cerebral perfusion pressure 70 compared to cerebral perfusion pressure 40. Cerebral blood flow was higher in the cerebral perfusion pressure 70 group but did not reach statistical significance. Phenylephrine was well tolerated and there were no observed increases in serum lactate or intracranial pressure in either group. Targeting a cerebral perfusion pressure of 70 mm Hg resulted in a greater reduction in metabolic crisis and cell injury volumes compared to a cerebral perfusion pressure of 40 mm Hg in an immature swine model. Early aggressive cerebral perfusion pressure augmentation to a cerebral perfusion pressure of 70 mm Hg in pediatric traumatic brain injury before severe intracranial hypertension has the potential to be neuroprotective, and further investigations are needed.

Increased Intracranial Pressure after Diffuse Traumatic Brain Injury Exacerbates Neuronal Somatic Membrane Poration but not Axonal Injury: Evidence for Primary Intracranial Pressure-Induced Neuronal Perturbation

Increased intracranial pressure (ICP) associated with traumatic brain injury (TBI) is linked to increased morbidity. Although our understanding of the pathobiology of TBI has expanded, questions remain regarding the specific neuronal somatic and axonal damaging consequences of elevated ICP, independent of its impact on cerebral perfusion pressure (CPP). To investigate this, Fischer rats were subjected to moderate TBI. Measurements of ICP revealed two distinct responses to injury. One population exhibited transient increases in ICP that returned to baseline levels acutely, while the other displayed persistent ICP elevation (>20 mm Hg). Utilizing these populations, the effect of elevated ICP on neuronal pathology associated with diffuse TBI was analyzed at 6 hours after TBI. No difference in axonal injury was observed, however, rats exhibiting persistently elevated ICP postinjury revealed a doubling of neurons with chronic membrane poration compared with rats exhibiting only transient increases in ICP. Elevated postinjury ICP was not associated with a concurrent increase in DNA damage; however, traditional histological assessments did reveal increased neuronal damage, potentially associated with redistribution of cathepsin-B from the lysosomal compartment into the cytosol. These findings indicate that persistently increased ICP, without deleterious alteration of CPP, exacerbates neuronal plasmalemmal perturbation that could precipitate persistent neuronal impairment and ultimate neuronal death.

Glycerol accumulation in edema formation following diffuse traumatic brain injury

Traumatic brain injury (TBI) induces brain edema via water and glycerol transport channels, called aquaporins (AQPs). The passage of glycerol across brain cellular compartments has been shown during edema. Using a modified impact/head acceleration rodent model of diffuse TBI, we assessed the role of hypoxia inducible factor (HIF)-1alpha in regulating AQP9 expression and glycerol accumulation during the edema formation. Adult (400–425 g) male Sprague–Dawley rats received a closed head injury with a weight drop (450 g, 2-m height) and were allowed to survive up to 48 hours. Some rat groups were administered 2-methoxyestradiol (2ME2, a HIF-1alpha inhibitor) 30 minutes after injury and were euthanized at 4 and 24 hours after injury. Brain edema was measured directly by water content, and glycerol concentration was determined by the Cayman Glycerol Assay. HIF-1alpha and AQP9 protein levels were assessed by Western immunoblotting. This study demonstrated a significant (P<0·05) increase in brain water content at 4–48 hours following impact. Cerebral glycerol was significantly (P<0·05) up-regulated at as early as 1 hour and remained at high levels for up to 48 hours. Similarly, significant (P<0·05) increases in HIF-1alpha and AQP9 protein levels were found at 1 hour and up to 48 hours after injury. Compared to untreated but injured rats, inhibition of HIF-1alpha by 2ME2 significantly (P<0·05) reduced the TBI-induced AQP9 up-regulation. This reduction was temporally associated with significant (P<0·05) decreases in both edema and glycerol accumulation. The data suggested an associated induction of HIF-1alpha, AQP9, and extracellular glycerol accumulation in edema formation following diffuse TBI. The implication of HIF-1alpha and AQP9 underlying TBI-induced edema formation offers possibilities for novel TBI therapies.

A Substance P Antagonist Improves Outcome in Female Sprague Dawley Rats Following Diffuse Traumatic Brain Injury

Over the past decade it has become increasingly clear from work in experimental models that the secondary injury cascade following traumatic brain injury (TBI) may differ between males and females [1, 2], with females exhibiting a different temporal profile of edema [1] and neurodegeneration compared to males [2]. Furthermore significant neuroprotection was noted in male, but not female rodents treated with posttraumatic hypothermia [3], or a dopamine agonist [4]. This highlights the need to ensure that experimental treatments are equally efficacious in both genders before proceeding to clinical trials that are performed in mixed populations. Previous research in our laboratory found that substance P (SP) plays an integral role in the secondary injury cascade following TBI in males [5]. SP is a member of the tachykinin family of neuropeptides, with its release causing the development of neurogenic inflammation characterised by vasodilation, plasma extravasation, and tissue swelling [6]. These effects are principally mediated by the binding of SP to the NK1 tachykinin receptor [6]. Following impact-acceleration TBI in male rodents SP levels were found to increase. Furthermore, inhibition of SP, through treatment with an NK1 receptor antagonist reduced vasogenic edema formation and axonal injury, with a corresponding improvement in motor and cognitive outcome [5, 7]. However, it is not known whether SP plays a similar role in females following TBI. As such, the effects of an NK1 receptor antagonist, N-acetyl-L-tryptophan (NAT) on outcome following TBI in female rodents were investigated. Female Sprague Dawley rats were injured using the impactacceleration model of diffuse TBI, as previously described [5]. At 30 mins postinjury rats were treated intravenously with either 2.5 mg/kg of NAT or an equal volume of saline. Sham animals were surgically prepared but not injured. Motor deficits were assessed on rotarod as previously described [5]. For histological analysis, rats were transcardially perfused fixed, the brains removed and processed. Slides were labeled with APP (1:1,000), SP (1:2,000), or albumin (1:20,000). To objectively analyse levels of SP and albumin, Ruifrok and Johnston’s color deconvolution method was employed on 2 slides per animal to determine the amount of DAB and hence antigen that was present, as previously described [8]. Axonal injury was determined by counting the number of APP immunopositive lengths within the corpus callosum. For analysis of edema the specific gravity of the brain was determined by placing it in a Percoll density gradient, with this measurement then converted to% water as previously described [9]. All data were analyzed using a one or two-way ANOVA as appropriate, followed by Bonferonni t-tests using Graphpad Prism software. SP immunoreactivity was assessed postinjury to determine whether levels were increased, as previously reported in males (Figures 1A and B). Indeed, at 24 h postinjury increased SP immunoreactivity was observed, particularly apparent in perivascular nerve fibers and astrocytic processes in vehicle treated rats. In contrast low levels of SP immunoreactivity were observed in shams. Furthermore color deconvolution analysis demonstrated a significant increase in%DAB weight in vehicle treated rats, with 28.0 ± 3.6% compared to 16.6 ± 2.8% in sham controls (P < 0.01).

Novel Treatment for Traumatic Brain Injury (S47.004)

Objective: We hypothesized that targeting ETrA could mitigate the deleterious effects of TBI. Background Traumatic brain injury results in multiple pathologies, including significant reduction in cerebral blood flow. Our laboratory have shown that endothelin-1 plays a major role in the induction of hypoperfusion. Furthermore, we demonstrated that the endothelin receptor A (ETrA) is upregulated as early as 4 hours post TBI and maintains upregulation up to 48 hours, thus temporally corresponding with the state of hypoperfusion. Design/Methods: Using a rodent model of diffuse traumatic brain injury (acceleration impact in Sprague-Dawley rats), we tested the effects of IV administration of a selective ETrA antagonist, Clazosentan administered at various times post TBI on CBF (as measured up to 48 hours post injury using arterial spin labeling MRI, n=6 per group) and behavioral outcome (measured up to 20 days post injury using a radial arm maze-spatial learning task, n=12-15 per group) following injury. Results: Our results indicate that when Clazosentan is administered at 30 minutes, 2 hours, or 4 hours after TBI there is a marked decrease in the extent of TBI-induced hypoperfusion. However, administration at 12 hours post-TBI resulted in variable effects (i.e., some animals improved, some worsened, and some remained the same). The most effective paradigm was a dual-administration approach in which drug was given at 2 and 24 hours post-TBI. Behavioral results recapitulated the blood flow results, showing that there was a trend towards improved behavioral outcome when Clazosentan was administered at 2 hours, however a significant improvement was seen when given at 2 and 24 hours. Conclusions: These results suggest that selective ETrA antagonism may be effective in mitigating the deleterious effects of TBI. Furthermore, it appears that our results help to define the “window of opportunity of drug delivery”. Supported by: NIH-NS064976(CK,PI), VARR&D-RX000224(CK, PI), and American Academy of Neurology(MK). Disclosure: Dr. Kaufman has nothing to disclose. Dr. Tiesma has nothing to disclose. Dr. Kreipke has received research support from Veterans Administration and National Institute of Health.

Substance P-induced changes in cell genesis following diffuse traumatic brain injury

Inhibition of substance P (SP) activity through the use of NK1 receptor antagonists has been shown to be a promising neuroprotective therapy following traumatic brain injury (TBI). Conversely, recent research has implicated SP in the stimulation of neurogenesis, suggesting that the neuropeptide has the potential to promote recovery following TBI. This study characterised the effects of SP and the NK1 antagonist, n-acetyl tryptophan (NAT), on cell proliferation following diffuse TBI. Adult male Sprague–Dawley rats were injured using the impact acceleration model of TBI and randomly assigned to one of five treatment groups: sham, vehicle control, NAT alone, SP alone or SP with NAT. Cellular proliferation was assessed with immunostaining for bromodeoxyuridine (BrdU) and cell-specific markers. Infusion of SP (±NAT) promoted cellular proliferation in the subventricular zone and dentate gyrus following TBI. This increase was largely associated with microglial proliferation and did not correspond with functional improvements. These results suggest that NAT treatment results in neuroprotection following TBI, mediated in part via inhibition of microglia.

Evaluation of the effects of treatment with sAPPα on functional and histological outcome following controlled cortical impact injury in mice

Treatment with sAPPα, the product of non-amyloidogenic processing of the amyloid precursor protein (APP) has been shown to be protective following diffuse traumatic brain injury (TBI), by improving motor outcome and reducing axonal injury. However the effects of treatment with sAPPα following a focal TBI have yet to be determined. To investigate this, mice were subjected to a controlled cortical impact injury and treated with either sAPPα or its vehicle at 30min post-injury. Following treatment with sAPPα the mice showed a significant improvement in motor and cognitive function early following injury, as determined on the ledged beam and Barnes Maze, respectively, relating to a more rapid rate of recovery. However the effect of treatment with sAPPα was not as dramatic as that seen previously following a diffuse injury. Nonetheless, these improvements in functional outcome were acompanied by a small but significant improvement in the amount of cortical and hippocampal at 7 days post-injury, and provide further support for the efficacy of sAPPα as a potential neuroprotective agent following TBI.