Nerve growth factor plays a role in the neurotherapeutic effect of a CD45+ pan-hematopoietic subpopulation derived from human umbilical cord blood in a traumatic brain injury model.

BACKGROUND AIMS: Human umbilical cord blood (HUCB) is an important source of stem cells for therapy of hematopoietic disorders and is a potential therapy for various neurological disorders, including traumatic brain injury (TBI). The expression of nerve growth factor (NGF) and its receptors TrkA, p75NTR and α9ß1 integrin on an HUCB CD45+ pan-hematopoietic subpopulation was investigated in the context of its neurotherapeutic potential after TBI. METHODS: NGF and its receptors were detected on CD45+ cells by reverse transcriptase polymerase chain reaction, flow cytometry analysis and confocal microscopy. CD45+ cells were stimulated by TBI brain extracts, and NGF levels were measured by enzyme-linked immunosorbent assay. TBI mice were divided into six groups for xenogeneic intravenous transplantation, 1 day post-trauma, with 1 × 106 CD45+ cells untreated or treated with the anti-NGF neutralizing antibody K252a, a TrkA antagonist; VLO5, an α9ß1 disintegrin; or negative (vehicle) and positive (NGF) controls. RESULTS: The HUCB CD45+ subpopulation constitutively expresses NGF and its receptors, mainly TrkA and p75NTR and minor levels of α9ß1. In vitro experiments provided evidence that trauma-related mediators from brain extracts of TBI mice induced release of NGF from HUCB CD45+ cell cultures. HUCB CD45+ cells induced a neurotherapeutic effect in TBI mice, abrogated by cell treatment with either anti-NGF antibody or K252a, but not VLO5. CONCLUSIONS: These findings strengthen the role of NGF and its TrkA receptor in the HUCB CD45+ subpopulation's neurotherapeutic effect. The presence of neurotrophin receptors in the HUCB CD45+ pan-hematopoietic subpopulation may explain the neuroprotective effect of cord blood in therapy of a variety of neurological disorders.

Regional sensitivity to neuroinflammation: in vivo and in vitro studies.

BACKGROUND: Neuroinflammation is involved in several acute-onset neuropathologies such as meningitis, encephalitis, stroke, and traumatic brain injury as well as in neurodegenerative diseases. All of these patholologies are associated with cognitive deficits. Using a model of pure neuroinflammation (intracisternal injection of endotoxin in mice), we tested the hypothesis that brain regions involved in cognition are the most vulnerable to inflammatory insults, and this vulnerability is an inherent property of neocortical neurons. METHODS: Mice (n = 10/group) injected with endotoxin (LPS) or saline in the cisterna magna underwent neurobehavioral and cognitive testing followed by quantitative autoradiographic assessment of regional neuroinflammation with [3H]PK11195, an established marker of microgliosis. In parallel, cocultures of cortical and striatal neurons taken from embryonic day 19 rat embryos or postnatal day 1 mice expressing green fluorescent protein were exposed for 24 h to the proinflammatory cytokine TNFalpha, glutamate, or a combination of the two agents. RESULTS: LPS-treated mice exhibited significant deficits in memory and significant increases in specific PK11195 binding in cortical and hippocampal regions, but not in striatum. Cultured neurons of cortical origin showed significantly lower survival rate relative to striatal neurons in response to TNFalpha, glutamate, or a combination of the two agents. Furthermore, TNFalpha exerted neuroprotective rather than neurotoxic effects in the striatal but not in the cortical neurons. CONCLUSIONS: These results suggest that the cortex is inherently more sensitive than the striatum to the deleterious effects of neuroinflammation, and may offer an explanation for the preponderance of cognitive deficits in neuropathologies with a neuroinflammatory component.

Histone deacetylase inhibitor ITF2357 is neuroprotective, improves functional recovery, and induces glial apoptosis following experimental traumatic brain injury.

Despite efforts aimed at developing novel therapeutics for traumatic brain injury (TBI), no specific pharmacological agent is currently clinically available. Here, we show that the pan-histone deacetylase (HDAC) inhibitor ITF2357, a compound shown to be safe and effective in humans, improves functional recovery and attenuates tissue damage when administered as late as 24 h postinjury. Using a well-characterized, clinically relevant mouse model of closed head injury (CHI), we demonstrate that a single dose of ITF2357 administered 24 h postinjury improves neurobehavioral recovery from d 6 up to 14 d postinjury (improved neurological score vs. vehicle; P< or =0.05), and that this functional benefit is accompanied by decreased neuronal degeneration, reduced lesion volume (22% reduction vs. vehicle; P< or =0.01), and is preceded by increased acetylated histone H3 levels and attenuation of injury-induced decreases in cytoprotective heat-shock protein 70 kDa and phosphorylated Akt. Moreover, reduced glial accumulation and activation were observed 3 d postinjury, and total p53 levels at the area of injury and caspase-3 immunoreactivity within microglia/macrophages at the trauma area were elevated, suggesting enhanced clearance of these cells via apoptosis following treatment. Hence, our findings underscore the relevance of HDAC inhibitors for ameliorating trauma-induced functional deficits and warrant consideration of applying ITF2357 for this indication.

Dynamic changes in the recovery after traumatic brain injury in mice: effect of injury severity on T2-weighted MRI abnormalities, and motor and cognitive functions.

Memory and neurobehavioral dysfunctions are among the sequelae of traumatic brain injury (TBI). The Neurological Severity Score (NSS) includes 10 tasks and was previously designed to assess the functional status of mice after TBI. The object recognition task (ORT) measures specific episodic memory and is expressed by the percent time spent by an animal at a novel, unfamiliar object (Discrimination Index [DI]). It is an ideal tool for evaluating cognitive function after TBI. The present study sought to validate the use of the NSS and ORT in severe and mild focal TBI in mice, and to confirm that the spontaneous recovery and the radiological abnormalities, shown by T2-weighted magnetic resonance imaging (MRI), are dependent upon injury severity. Mice were subjected to severe and mild closed head injury (NSS at 1 h 7.52 +/- 0.34 and 4.62 +/- 0.14, respectively). NSS was evaluated for 25 days and showed a decrease by 3.86 +/- 0.26 and 2.54 +/- 0.35 units in the severely and mildly injured mice, respectively. ORT revealed DI in severely injured group of 51.7 +/- 6.15%, (vs approximately 75-80% in uninjured animal) on day 3 and 66.2 +/- 6.81% on day 21. In contrast, the mildly injured mice did not show cognitive impairment throughout the same period. The damage seen by MRI at 24 h after injury, strongly correlated with NSS(1h) (R = 0.87, p < 0.001). We conclude that NSS is a reliable tool for evaluation of neurological damage in head-injured mice, NSS(1h) predicts the motor dysfunction, cognitive damage, and brain-damage characteristics as depicted by T2-weighted MRI. The combined assessment of neurobehavioral and cognitive function along with MRI is most useful in evaluating recovery from injury, especially when testing effectiveness of novel treatments or genetic manipulations.

Differential neuroprotective properties of endogenous and exogenous erythropoietin in a mouse model of traumatic brain injury.

Both heat acclimation (HA) and post-injury treatment with recombinant human erythropoietin (Epo, rhEpo, exogenous Epo) are neuroprotective against traumatic brain injury (TBI). Our previous data demonstrated that HA-induced neuroprotection includes improved functional recovery and reduced cerebral edema formation. Additionally, in earlier Western-blot analyses, we found that HA mice display increased expression of the specific erythropoietin receptor (EpoR) and of hypoxia-inducible factor-1 alpha (HIF-1 alpha), the inducible subunit of the transcription factor, which regulates Epo gene expression, but not of Epo itself. In light of this, the aim of the current study was threefold: (1) to assess Epo expression in the trauma area and hippocampus following HA, rhEpo administration, or combined HA-rhEpo treatment, using immunohistochemical methods that offer enhanced anatomical resolution; (2) to examine the effects of endogenous and exogenous Epo on edema formation in normothermic (NT) mice; and (3) to evaluate the effects of exogenous Epo administration on neuroprotective outcome measures in HA animals. HA induced enhanced expression of endogenous Epo in the trauma area and the hippocampus. Treatment with anti-Epo antibody given to NT mice increased edema formation, whereas rhEpo induced no beneficial effect. Cognitive performance testing and immunohistochemical findings reinforced HA and rhEpo as separate protective interventions but showed no advantage to combining the two strategies. We therefore suggest that HA-induced neuroprotection is shaped by pre-existing mediators but cannot be modified by post-injury treatment aimed at increasing the levels of neuroprotective agents.

D-cycloserine improves functional recovery and reinstates long-term potentiation (LTP) in a mouse model of closed head injury.

Traumatic brain injury triggers a massive glutamate efflux, activation of NMDA receptor channels, and cell death. Recently, we reported that NMDA receptors in mice are down-regulated from hours to days following closed head injury (CHI), and treatment with NMDA improved recovery of motor and cognitive functions up to 14 d post-injury. Here we show that a single injection of a low dose of D-cycloserine (DCS), a partial NMDA receptor agonist, in CHI mice 24 h post-injury, resulted in a faster and greater recovery of motor and memory functions as assessed by neurological severity score and object recognition tests, respectively. Moreover, DCS treatment of CHI mice led to a significant improvement of hippocampal long-term potentiation (LTP) in the CA1 region that was completely blunted in CHI control mice. However, DCS did not improve CHI-induced impairment in synaptic glutamate release measured by paired pulse facilitation (PPF) ratio in hippocampal CA1 region. Finally, CHI-induced reduction of brain-derived neurotrophic factor (BDNF) was fully restored following DCS treatment. Since DCS is in clinical use for other indications, the present study offers a novel approach to treat human brain injury.

Akt phosphorylation is required for heat acclimation-induced neuroprotection.

Long-term heat exposure, known as heat acclimation (HA; 30 days at 34 +/- 1 degrees C) is neuroprotective against traumatic brain injury. Acclimated mice were previously found to display improved functional recovery as well as an increase in the levels of the specific erythropoietin receptor. As the activation of this receptor is known to facilitate functional recovery on one hand and the phosphorylation and activation of Akt, an intracellular kinase which regulates anti-apoptotic pathways on the other, in this study we investigated whether HA affects Akt phosphorylation prior to and following injury and whether this step is required for development of HA-induced neuroprotection. Akt phosphorylation was blocked using Triciribine (TCN), a compound shown to block the phosphorylation process without affecting upstream effectors of this kinase, and several post-injury functional end-point measures were subsequently evaluated. Acclimation led to a post-injury increase in the levels of phosphorylated Akt, resulting in higher levels when compared with normothermic controls at 4 h post-injury (63.6 +/- 5.2% and 42.7 +/- 3.7%, respectively, p

Altered cytokine expression and sustained hypothermia following traumatic brain injury in heat acclimated mice.

Long-term exposure to moderate ambient heat (heat acclimation, HA, 30 days at 34+/-1 degrees C) provides protection toward a variety of stressors including traumatic brain injury. As previous studies suggested an anti-inflammatory effect of HA and given the ability of augmented pre-injury anti-inflammatory cytokine expression to harbor neuroprotection and to attenuate early post-injury expression of pro-inflammatory mediators, we hypothesized that HA-induced neuroprotection may involve enhanced pre-injury expression of anti-inflammatory mediators or a reduction in post-injury TNF alpha (TNFalpha) expression. Since the attenuation of inflammatory-associated entities has also been linked to mild hypothermia, an established neuroprotective paradigm, the effect of HA on post-injury body temperature was also studied. HA mice and normothermic (NT) counterparts were examined using a closed head injury model. Cytokines were measured within the ipsilateral cortex. Pre-injury protein levels of anti-inflammatory interleukins 10 and 4 (IL-10, IL-4) were quantified by enzyme-linked immunosorbent assays (ELISA). mRNA and protein levels of TNFalpha were quantified during the initial 2 h post-injury using semi-quantitative and real-time polymerase chain reaction (sqRT-PCR and qRT-PCR) or ELISA, respectively. Rectal temperatures were measured. HA induced augmented pre-injury IL-10 expression and a post-injury reduction in TNFalpha mRNA levels, as well as altered expression dynamics of TNFalpha protein. TNFalpha protein levels decreased relative to the sham state in HA mice only. HA mice displayed sustained post-injury hypothermia, namely significantly lower body temperature at 4 h post-injury. Given the evidence on the neuroprotective nature of hypothermia and anti-inflammatory cytokines, we suggest that these changes may contribute to HA-induced neuroprotection.

CuZn-SOD deficiency, rather than overexpression, is associated with enhanced recovery and attenuated activation of NF-kappaB after brain trauma in mice.

Superoxide-dismutases (SOD) catalyze O2- conversion to hydrogen peroxide (H2O2) and with other antioxidant enzymes and low molecular weight antioxidants (LMWA) constitute endogenous defense mechanisms. We first assessed the effects of SOD1 levels on outcome after closed head injury (CHI) and later, based on these results, the effects of SOD1 deficiency on cellular redox homeostasis. Superoxide-dismutase 1-deficient (SOD1-/-) and -overexpressing (transgenic (Tg)) mice and matched wild-type (WT) controls were subjected to CHI and outcome (neurobehavioral and memory functions) was assessed during 14 days. Brain edema, LMWA, and SOD2 activity were measured along with histopathological analysis. Transactivation of nuclear factor-kappa B (NF-kappaB) was evaluated by electromobility shift assay. Mortality, motor, and cognitive outcome of Tg and WT mice were comparable. Mortality and edema were similar in SOD1-/- and WT mice, yet, unexpectedly, SOD1-/- displayed better neurobehavioral recovery (P<0.05) at 14 days after CHI. Basal LMWA were higher in the cortex and liver of SOD1-/- mice (P<0.05) and similar to WT in the cerebellum. Five minutes after CHI, cortical LMWA decreased only in SOD1-/- mice. One week after CHI, SOD2 activity decreased fourfold in WT cortex (P<0.001), but was preserved in the SOD1-/-. Constitutive NF-kappaB transactivation was comparably low in SOD1-/- and WT; however, CHI induced a robust NF-kappaB activation that was absent in SOD1-/- cortices (P<0.005 versus WT). At the same time, immunohistochemical analysis of brain sections revealed that astrogliosis and neurodegeneration were of lesser severity in SOD1-/- mice. We suggest that SOD1 deficiency impairs H2O2-mediated activation of NF-kappaB, decreasing death-promoting signals, and leading to better outcome.

Heat acclimation increases hypoxia-inducible factor 1alpha and erythropoietin receptor expression: implication for neuroprotection after closed head injury in mice.

Experimental evidence indicates that long-term exposure to moderately high ambient temperature (heat acclimation, HA) mediates cross-tolerance to various types of subsequently applied stress. The transcriptional activator hypoxia-inducible factor 1 (HIF-1) has been implicated in playing a critical role in HA. It also regulates the expression of Erythropoietin (Epo), whose neuroprotective effects have been shown in a variety of brain injuries. The aim of the present study was to examine whether HA exerts a beneficial effect on the outcome of closed head injury (CHI) in mice and to explore the possible involvement of HIF-1 and Epo in this process. Heat acclimated mice and matched normothermic controls were subjected to CHI or sham surgery. Postinjury motor and cognitive parameters of acclimated mice were compared with those of controls. Mice were killed at various time points after injury or sham surgery and brain levels of HIF-1alpha, the inducible subunit of HIF-1, Epo, and the specific erythropoietin receptor (EpoR) were analyzed by Western immunoblotting. Motor and cognitive functions of acclimated mice were significantly better than those of controls. Heat acclimation was found to induce a significant increase in expression of nuclear HIF-1alpha and EpoR. The EpoR/Epo ratio was also significantly higher in acclimated mice as compared with controls. Nuclear HIF-1alpha and EpoR were higher in the acclimated group at 4 h after injury as well. The improved outcome of acclimated mice taken together with the basal and postinjury upregulation of the examined proteins suggests the involvement of this pathway in HA-induced neuroprotection.

Inosine improves functional recovery after experimental traumatic brain injury.

Despite years of research, no effective therapy is yet available for the treatment of traumatic brain injury (TBI). The most prevalent and debilitating features in survivors of TBI are cognitive deficits and motor dysfunction. A potential therapeutic method for improving the function of patients following TBI would be to restore, at least in part, plasticity to the CNS in a controlled way that would allow for the formation of compensatory circuits. Inosine, a naturally occurring purine nucleoside, has been shown to promote axon collateral growth in the corticospinal tract (CST) following stroke and focal TBI. In the present study, we investigated the effects of inosine on motor and cognitive deficits, CST sprouting, and expression of synaptic proteins in an experimental model of closed head injury (CHI). Treatment with inosine (100 mg/kg i.p. at 1, 24 and 48 h following CHI) improved outcome after TBI, significantly decreasing the neurological severity score (NSS, p<0.04 vs. saline), an aggregate measure of performance on several tasks. It improved non-spatial cognitive performance (object recognition, p<0.016 vs. saline) but had little effect on sensorimotor coordination (rotarod) and spatial cognitive functions (Y-maze). Inosine did not affect CST sprouting in the lumbar spinal cord but did restore levels of the growth-associated protein GAP-43 in the hippocampus, though not in the cerebral cortex. Our results suggest that inosine may improve functional outcome after TBI.

Neuroprotection after traumatic brain injury in heat-acclimated mice involves induced neurogenesis and activation of angiotensin receptor type 2 signaling.

Long-term exposure of mice to mild heat (34°C±1°C) confers neuroprotection against traumatic brain injury (TBI); however, the underling mechanisms are not fully understood. Heat acclimation (HA) increases hypothalamic angiotensin II receptor type 2 (AT2) expression and hypothalamic neurogenesis. Accumulating data suggest that activation of the brain AT2 receptor confers protection against several types of brain pathologies, including ischemia, a hallmark of the secondary injury occurring following TBI. As AT2 activates the same pro-survival pathways involved in HA-mediated neuroprotection (e.g., Akt phosphorylation, hypoxia-inducible factor 1α (HIF-1α), and brain-derived neurotrophic factor (BDNF)), we examined the role of AT2 in HA-mediated neuroprotection after TBI. Using an AT2-specific antagonist PD123319, we found that the improvements in motor and cognitive recovery as well as reduced lesion volume and neurogenesis seen in HA mice were all diminished by AT2 inhibition, whereas no significant alternations were observed in control mice. We also found that nerve growth factor/tropomyosin-related kinase receptor A (TrkA), BDNF/TrkB, and HIF-1α pathways are upregulated by HA and inhibited on PD123319 administration, suggesting that these pathways play a role in AT2 signaling in HA mice. In conclusion, AT2 is involved in HA-mediated neuroprotection, and AT2 activation may be protective and should be considered a novel drug target in the treatment of TBI patients.

Neurotherapeutic effect of cord blood derived CD45+ hematopoietic cells in mice after traumatic brain injury.

Treatment of traumatic brain injury (TBI) is still an unmet need. Cell therapy by human umbilical cord blood (HUCB) has shown promising results in animal models of TBI and is under evaluation in clinical trials. HUCB contains different cell populations but to date, only mesenchymal stem cells have been evaluated for therapy of TBI. Here we present the neurotherapeutic effect, as evaluated by neurological score, using a single dose of HUCB-derived mononuclear cells (MNCs) upon intravenous (IV) administration one day post-trauma in a mouse model of closed head injury (CHI). Delayed (eight days post-trauma) intracerebroventricular administration of MNCs showed improved neurobehavioral deficits thereby extending the therapeutic window for treating TBI. Further, we demonstrated for the first time that HUCB-derived pan-hematopoietic CD45 positive (CD45(+)) cells, isolated by magnetic sorting and characterized by expression of CD45 and CD11b markers (96-99%), improved the neurobehavioral deficits upon IV administration, which persisted for 35 days. The therapeutic effect was in a direct correlation to a reduction in the lesion volume and decreased by pre-treatment of the cells with anti-human-CD45 antibody. At the site of brain injury, 1.5-2 h after transplantation, HUCB-derived cells were identified by near infrared scanning and immunohistochemistry using anti-human-CD45 and anti-human-nuclei antibodies. Nerve growth factor and vascular endothelial growth factor levels were differentially expressed in both ipsilateral and contralateral brain hemispheres, thirty-five days after CHI, measured by enzyme-linked immunosorbent assay. These findings indicate the neurotherapeutic potential of HUCB-derived CD45(+) cell population in a mouse model of TBI and propose their use in the clinical setting of human TBI.

Hypoxia-inducible factor 1 is essential for spontaneous recovery from traumatic brain injury and is a key mediator of heat acclimation induced neuroprotection.

Heat acclimation (HA), a well-established preconditioning model, confers neuroprotection in rodent models of traumatic brain injury (TBI). It increases neuroprotective factors, among them is hypoxia-inducible factor 1α (HIF-1α), which is important in the response to postinjury ischemia. However, little is known about the role of HIF-1α in TBI and its contribution to the establishment of the HA protecting phenotype. Therefore, we aimed to explore HIF-1α role in TBI defense mechanisms as well as in HA-induced neuroprotection. Acriflavine was used to inhibit HIF-1 in injured normothermic (NT) or HA mice. After TBI, we evaluated motor function recovery, lesion volume, edema formation, and body temperature as well as HIF-1 downstream transcription targets, such as glucose transporter 1 (GLUT1), vascular endothelial growth factor, and aquaporin 4. We found that HIF-1 inhibition resulted in deterioration of motor function, increased lesion volume, hypothermia, and reduced edema formation. All these parameters were significantly different in the HA mice. Western blot analysis and enzyme-linked immunosorbent assay showed reduced levels of all HIF-1 downstream targets in HA mice, however, only GLUT1 was downregulated in NT mice. We conclude that HIF-1 is a key mediator in both spontaneous recovery and HA-induced neuroprotection after TBI.

The role and dynamics of ß-catenin in precondition induced neuroprotection after traumatic brain injury.

Preconditioning via heat acclimation (34°C 30 d) results in neuroprotection from traumatic brain injury due to constitutive as well as dynamic changes triggered by the trauma. Among these changes is Akt phosphorylation, which decreases apoptosis and induces HIF1α. In the present study we investigated the Akt downstream GSK3ß/ß-catenin pathway and focused on post injury alternations of ß catenin and its impact on the cellular response in preconditioned heat acclimated mice. We found that the reduction in motor disability is accompanied with attenuation of depressive like behavior in heat acclimated mice that correlates with the GSK3ß phosphorylation state. Concomitantly, a robust ß catenin phosphorylation is not followed by its degradation, or by reduced nuclear accumulation. Enhanced tyrosine phosphorylation of ß catenin in the injured area weakens the ß catenin-N cadherin complex. Membrane ß catenin is transiently reduced in heat acclimated mice and its recovery 7 days post TBI is accompanied by induction of the synaptic marker synaptophysin. We suggest a set of cellular events following traumatic brain injury in heat acclimated mice that causes ß catenin to participate in cell-cell adhesion alternations rather than in Wnt signaling. These events may contribute to synaptogenesis and the improved motor and cognitive abilities seen heat acclimated mice after traumatic brain injury.

The anti-inflammatory drug carprofen improves long-term outcome and induces gliogenesis after traumatic brain injury.

Traumatic brain injury (TBI) initiates acute and chronic inflammatory processes involving cyclooxygenase-2 (COX-2), which may have detrimental effects on outcome and especially on brain regeneration. Therefore we aimed to study whether carprofen, a COX-2 inhibitor, would improve outcome and increase neurogenesis after TBI. TBI was induced in Sabra mice that were then treated with vehicle or carprofen for 7 days. Functional outcome was evaluated with the Neurological Severity Score (NSS).Cytokine levels were assessed 4 h post-TBI and water content was measured 24 h post TBI. Mice were given BrdU to label newborn cells for 10 days. The animals were killed 90 days post-TBI and the lesion size as well as newborn cell fate were assessed. Carprofen significantly reduced lesion size (p=0.002), decreased water content in the lesioned cortex (p=0.03), reduced the number of microglia in the lesioned cortex (p<0.0001), and lowered the levels of proinflammatory cytokines (IL-1ß, p=0.03; IL-6, p=0.02). Carprofen led to significantly larger improvements in functional outcome (p≤0.008) which were durable over 90 days. Carprofen also induced a threefold increase in the proliferation of new cells in the peri-lesion area (p≤0.002), but newborn cells differentiated mainly into glia in both groups. Carprofen is neuroprotective and induces cell proliferation and gliogenesis after TBI. Treatment with carprofen is consistently associated with better functional outcome. Our results imply that anti-inflammatory drugs may represent novel therapeutic options for TBI.

Combination of vascular endothelial and fibroblast growth factor 2 for induction of neurogenesis and angiogenesis after traumatic brain injury.

Fibroblast growth factor 2 (FGF2) and vascular endothelial growth factor (VEGF) are potent mitogens for endogenous neural stem cells (eNSC) and also induce angiogenesis. We infused the individual factors or their combination into the lateral ventricles of mice for 7 days after traumatic brain injury (TBI) in order to evaluate the effects on functional outcome and on eNSC proliferation and differentiation. The results show that VEGF induced a significant increment in the number of proliferating eNSC in the subventricular zone and in the perilesion cortex and that combination of FGF2 and VEGF did not augment the effects of VEGF alone. Fate analysis showed that most newborn cells differentiated into astrocytes and oligodendroglia while only a few cells differentiated into neurons. Functional outcome was significantly better in mice treated with VEGF, FGF2, or their combination as compared to vehicle. Injury size was significantly reduced only in mice treated with VEGF suggesting additional neuroprotective effects for VEGF. Combination therapy did not have an additive effect on outcome or neuronal differentiation. In conclusion, FGF2-VEGF combination does not augment neurogenesis and angiogenesis or reduce lesion volumes after TBI compared with individual factors. This may suggest the existence of a ceiling effect for brain regeneration.

Vascular endothelial growth factor increases neurogenesis after traumatic brain injury.

Activation of endogenous stem cells has been proposed as a novel form of therapy in a variety of neurologic disorders including traumatic brain injury (TBI). Vascular endothelial growth factor (VEGF) is expressed in the brain after TBI and serves as a potent activator of angiogenesis and neurogenesis. In this study, we infused exogenous VEGF into the lateral ventricles of mice for 7 days after TBI using mini-osmotic pumps to evaluate the effects on recovery and functional outcome. The results of our study show that VEGF significantly increases the number of proliferating cells in the subventricular zone and in the perilesion cortex. Fate analysis showed that most newborn cells differentiated into astrocytes and oligodendroglia and only a few cells differentiated into neurons. Functional outcome was significantly better in mice treated with VEGF compared with vehicle-treated animals after TBI. Injury size was significantly smaller at 90 days after TBI in VEGF-treated animals, suggesting additional neuroprotective effects of VEGF. In conclusion, VEGF significantly augments neurogenesis and angiogenesis and reduces lesion volumes after TBI. These changes are associated with significant improvement in recovery rates and functional outcome.

Heat acclimation provides sustained improvement in functional recovery and attenuates apoptosis after traumatic brain injury.

Heat acclimation (HA) offers functional neuroprotection in mice after traumatic brain injury (TBI). This study further characterizes endogenous neuroprotection acquired by HA (34+/-1 degrees C, 30 d) after TBI. We establish here the ability of HA to induce sustained functional benefits and to reduce activation of apoptotic pathways. Neurobehavioral recovery, assessed by the Neurological Severity Score, was greater in HA mice up to 8 days after injury as compared with normothermic controls (P<0.05) and lesion volume was also smaller in the HA group (P<0.05). Reduced apoptotic cell death in HA mice was confirmed using caspase-3 activity measurements and immunohistochemistry. To investigate the underlying molecular pathways, expression levels of intrinsic apoptotic pathway-related proteins were examined. HA mice displayed higher mitochondrial levels of antiapoptotic Bcl-xL, accompanied by lower proapoptotic Bad levels and decreased cytochrome c release, suggesting a higher apoptotic threshold. Taken together with our previous reports, indicating increased Akt phosphorylation and antioxidative capacity, alongside with reduced tumor necrosis alpha levels after TBI in HA animals, the current results support the involvement of an antiapoptotic effect in HA-induced neuroprotection. Current results warrant further study as TBI-induced apoptosis may persist over weeks after injury, possibly providing a target for belated therapeutic intervention.