Acute Neuronal Injury Research Articles

REVIEW ARTILCE: Cell therapy and stem cells in animal models of motor neuron disorders

AbstractAmyotrophic lateral sclerosis (ALS), spinal bulbar muscular atrophy (or Kennedy's disease), spinal muscular atrophy and spinal muscular atrophy with respiratory distress 1 are neurodegenerative disorders mainly affecting motor neurons and which currently lack effective therapies. Recent studies in animal models as well as primary and embryonic stem cell models of ALS, utilizing over‐expression of mutated forms of Cu/Zn superoxide dismutase 1, have shown that motor neuron degeneration in these models is in part a non cell‐autonomous event and that by providing genetically non‐compromised supporting cells such as microglia or growth factor‐excreting cells, onset can be delayed and survival increased. Using models of acute motor neuron injury it has been shown that embryonic stem cell‐derived motor neurons implanted into the spinal cord can innervate muscle targets and improve functional recovery. Thus, a rationale exists for the development of cell therapies in motor neuron diseases aimed at either protecting and/or replacing lost motor neurons, interneurons as well as non‐neuronal cells. This review evaluates approaches used in animal models of motor neuron disorders and their therapeutic relevance.

Matrix metalloproteinase-9 and urokinase plasminogen activator mediate interleukin-1-induced neurotoxicity

Matrix metalloproteinases (MMPs) are endopeptidases known to mediate acute neuronal injury, but it is unclear whether these proteases are induced by the primary insult or by inflammation associated with injury. We have reported recently that interleukin-1 (IL-1) induces neurotoxicity by an astrocyte-dependent mechanism. The aim of the present study was to test the hypothesis that MMPs mediate IL-1 neurotoxicity in rat, glial–neuronal cocultures. IL-1β induced the release of astrocytic MMP-9 in cocultures, whilst an antagonist of MMP-9 inhibited IL-1β-induced neuronal death. Urokinase plasminogen activator (uPA) was constitutively expressed on neuronal membrane fractions, and amiloride (an antagonist of uPA) or plasminogen activator inhibitor (PAI)-1 significantly reduced IL-1β-induced neurotoxicity. Thus, neuronal uPA contributes to IL-1 neurotoxicity, and may be responsible for activating MMP-9 released from IL-1-primed astrocytes. In summary, IL-1-induced neurotoxicity is dependent on extracellular protease activity, and these mechanisms may contribute to neuronal cell death in CNS diseases.

The autopsy in sudden unexpected adult death: Epilepsy

Summary There are up to 500 epilepsy-related deaths annually in the UK, many of which are unwitnessed. Likely mechanisms for sudden and unexpected death in epilepsy (SUDEP) are cerebrogenic cardiac arrhythmias, or central respiratory depression occurring during the peri-ictal period. Pathologists should be informed of the circumstances of the death, severity of seizures, seizure control and the certainty of the clinical diagnosis of epilepsy; this allows accurate clinicopathological correlation. SUDEP autopsies include neuropathological assessment, histological examination of other organs and toxicology, and require the elimination of other causes of sudden death. Macroscopic (non-fatal) abnormalities described in SUDEP include evidence of previous cerebral injury, hippocampal sclerosis and cerebellar atrophy. Histological examination may reveal neuronal loss and gliosis consistent with seizure-related brain injury. Hippocampal sclerosis shows subfield-specific patterns of neuronal loss, granule cell dispersion and mossy fibre sprouting. Rarely, acute neuronal injury is seen as evidence of a more recent cerebral event. This article discusses the pathological findings and possible mechanisms in SUDEP, and future directions for pathology-based research.

Destruction and reconstruction: Hypoxia and the developing brain

Preterm infants have a high rate of neurodevelopmental handicap. Recent imaging studies have revealed that adverse outcomes are strongly associated with reduced brain growth and neural complexity in later life. Increasing data suggest that these chronic deficits primarily reflect acute neuronal and glial injury sustained during adverse in utero events, such as exposure to severe hypoxia-ischemia and inflammation. In the present review we examine recent evidence that this chronic impairment is partly due to upregulation of physiological apoptosis, related to input deprivation, and output isolation secondary to acute white and gray matter damage and axonal injury. However, progenitor cells in the subventricular zone (SVZ) are also vulnerable to injury, and loss of part of this critical population likely further compromises brain development. Based on these concepts the impact of proposed interventions such as induced hypothermia and endogenous growth factors are likely to be complex, but potentially offer focused ways of improving the outcomes of premature birth.

Mitofusin 2 Protects Cerebellar Granule Neurons against Injury-induced Cell Death

Of the GTPases involved in the regulation of the fusion machinery, mitofusin 2 (Mfn2) plays an important role in the nervous system as point mutations of this isoform are associated with Charcot Marie Tooth neuropathy. Here, we investigate whether Mfn2 plays a role in the regulation of neuronal injury. We first examine mitochondrial dynamics following different modes of injury in cerebellar granule neurons. We demonstrate that neurons exposed to DNA damage or oxidative stress exhibit extensive mitochondrial fission, an early event preceding neuronal loss. The extent of mitochondrial fragmentation and remodeling is variable and depends on the mode and the severity of the death stimuli. Interestingly, whereas mitofusin 2 loss of function significantly induces cell death in the absence of any cell death stimuli, expression of mitofusin 2 prevents cell death following DNA damage, oxidative stress, and K+ deprivation induced apoptosis. More importantly, whereas wild-type Mfn2 and the hydrolysis-deficient mutant of Mfn2 (Mfn2(RasG12V)) function equally to promote fusion and lengthening of mitochondria, the activated Mfn2(RasG12V) mutant shows a significant increase in the protection of neurons against cell death and release of proapoptotic factor cytochrome c. These findings highlight a signaling role for Mfn2 in the regulation of apoptosis that extends beyond its role in mitochondrial fusion.

Dynamic Regulation of Acid-Sensing Ion Channels by Extracellular and Intracellular Modulators

Changes of extracellular pH values can have profound effects on neuronal function. For example, the low pH (also called acidosis) generated in brain ischemia causes acute neuronal injury. For years the receptors that detect pH variations surrounding neurons and their physiological/pathological importance remain uncertain. The recent finding that acidosis activates a distinct family of membrane ion channels, the acid-sensing ion channels (ASICs) in both peripheral and central neurons has dramatically changed the view of acidosis-associated signaling and provided a new strategy for therapeutic inventions. Although proton is the only known agonist for the activation of ASICs, a variety of extracellular and intracellular signaling molecules can modulate the activities of ASICs and have profound influence on the functions of these channels in both physiological and pathological processes. The goal of this article is therefore to provide a comprehensive review of the modulators of ASICs that adapt ASIC activity to changes of extracellular and intracellular environments.

Progress in shivering control

Hypothermia is a potent neuroprotectant and induced hypothermia holds great promise as a therapy for acute neuronal injury. Thermoregulatory responses, most notably shivering, present major obstacles to therapeutic temperature management. A review of thermoregulatory physiology and strategies aimed at controlling physiologic responses to hypothermia is presented.

Roles of Volume-Sensitive Chloride Channel in Excitotoxic Neuronal Injury

Excitotoxicity is associated with stroke, brain trauma, and a number of neurodegenerative disorders. In the brain, during excitotoxic insults, neurons undergo rapid swelling in both the soma and dendrites. Focal swellings along the dendrites called varicosities are considered to be a hallmark of acute excitotoxic neuronal injury. However, it is not clear what pathway is involved in the neuronal anion flux that leads to the formation and resolution of excitotoxic varicosities. Here, we assessed the roles of the volume-sensitive outwardly rectifying (VSOR) Cl- channel in excitotoxic responses in mouse cortical neurons. Whole-cell patch-clamp recordings revealed that the VSOR Cl- channel in cultured neurons was activated by NMDA exposure. Moreover, robust expression of this channel on varicosities was confirmed by on-cell and nystatin-perforated vesicle patch techniques. VSOR channel blockers, but not blockers of GABA(A) receptors and Cl- transporters, abolished not only varicosity resolution after sublethal excitotoxic stimulation but also necrotic death after sustained varicosity formation induced by prolonged NMDA exposure in cortical neurons. The present slice-patch experiments demonstrated, for the first time, expression of the VSOR Cl- channels in somatosensory pyramidal neurons. NMDA-induced necrotic neuronal death in slice preparations was largely suppressed by a blocker of the VSOR Cl- channel but not of the GABA(A) receptor. These results indicate that VSOR Cl- channels exert dual, reciprocal actions on neuronal excitotoxicity by serving as major anionic pathways both for varicosity recovery after washout of an excitotoxic stimulant and for persistent varicosity formation under prolonged excitotoxic insults leading to necrosis in cortical neurons.

Spatio-temporal expression of cysteinyl leukotriene receptor-2 mRNA in rat brain after focal cerebral ischemia

Cysteinyl leukotrienes (CysLTs) induce inflammatory responses mediated by activating CysLT 1 and CysLT 2 receptors. We have recently reported that CysLT 1 receptor expression is increased in rat brain after focal cerebral ischemia and the increased expression is spatio-temporally related to acute neuronal injury and late astrocyte proliferation. Here we report spatio-temporal expression of CysLT 2 receptor mRNA in rat brain after focal cerebral ischemia induced by 30 min of middle cerebral artery occlusion. We found that the neuron density was gradually decreased or disappeared in the ischemic core and boundary zone during 14 days after reperfusion, and the astrocyte population in the boundary zone was increased 3–14 days after reperfusion. In the ischemic core, the expression of CysLT 2 receptor mRNA was increased at 6, 12 and 24 h and then recovered at 3, 7 and 14 days after reperfusion. In the boundary zone, the expression was significantly increased 3, 7 and 14 days after reperfusion. The results suggest that CysLT 2 receptor may be related to the acute neuronal injury and late astrocyte proliferation in the ischemic brain.

Cysteinyl leukotriene receptors CysLT1and CysLT2are upregulated in acute neuronal injury after focal cerebral ischemia in mice

To determine whether cysteinyl leukotriene receptors (CysLT1 and CysLT2) are upregulated in acute neuronal injury after focal cerebral ischemia in mice, and to confirm CysLT1 receptor localization. After permanent focal cerebral ischemia was induced by middle cerebral artery occlusion (MCAO), neurological deficits and neuron loss were determined at various time points within 48 h. The mRNA expressions of CysLT1 and CysLT2 receptors in the brain were analyzed by RT-PCR. CysLT1 receptor localization was detected by double immunofluorescence. Neurological deficits and neuron loss were found 6, 24 and 48 h after MCAO. The mRNA expressions of both CysLT1 and CysLT2 receptors were upregulated in the ischemic hemisphere 1, 24, and 48 h after MCAO with peaks at 24 h. The CysLT1 receptor was selectively localized in neurons 24 h after MCAO. CysLT1 and CysLT2 receptors are upregulated in acute neuronal injury after focal cerebral ischemia, and the CysLT1 receptor is localized in neurons after ischemia.

Neurochemical aftermath of amateur boxing

BACKGROUND: Little solid information is available on the possible risks for neuronal injury in amateur boxing.OBJECTIVE: To determine whether amateur boxing and severity of hits are associated with elevated levels of biochemical markers for neuronal injury in cerebrospinal fluid.DESIGN: Longitudinal study.SETTING: Referral center specializing in evaluation of neurodegenerative disorders.PARTICIPANTS: Fourteen amateur boxers (11 men and three women) and 10 healthy male nonathletic control subjects.INTERVENTIONS: The boxers underwent lumbar puncture 7–10 days and 3 months after a bout. The control subjects underwent LP once.MAIN OUTCOME MEASURES: Neurofilament light protein, total tau, glial fibrillary acidic protein, phosphorylated tau, and beta‐amyloid protein 1–40 (Abeta([1–40])) and 1–42 (Abeta([1–42])) concentrations in cerebrospinal fluid were measured.RESULTS: Increased levels after a bout compared with after 3 months of rest from boxing were found for two markers for neuronal and axonal injury, neurofilament light protein (mean±SD, 845±1140 vs 208±108 ng/L; P=0.008) and total tau (mean±SD, 449±176 vs 306±78 ng/L; P=0.006), and for the astroglial injury marker glial fibrillary acidic protein (mean±SD, 541±199 vs 405±138 ng/L; P=0.003). The increase was significantly higher among boxers who had received many hits (>15) or high‐impact hits to the head compared with boxers who reported few hits. In the boxers, concentrations of neurofilament light protein and glial fibrillary acidic protein, but not total tau, were significantly elevated after a bout compared with the nonathletic control subjects. With the exception of neurofilament light protein, there were no significant differences between boxers after 3 months of rest from boxing and the nonathletic control subjects.CONCLUSIONS: Amateur boxing is associated with acute neuronal and astroglial injury. If verified in longitudinal studies with extensive follow‐up regarding the clinical outcome, analyses of cerebrospinal fluid may provide a scientific basis for medical counseling of athletes after boxing or head injury.

The HSV-2 protein ICP10PK prevents neuronal apoptosis and loss of function in an in vivo model of neurodegeneration associated with glutamate excitotoxicity

Excessive glutamate receptor activation results in neuronal death, a process known as excitotoxicity. Intrastriatal injection of N-methyl- d-aspartate (NMDA) is a model of excitotoxicity. We used this model to examine whether excitotoxic injury is inhibited by the anti-apoptotic herpes simplex virus type 2 (HSV-2) protein, ICP10PK, delivered by the replication incompetent HSV-2 vector, ΔRR. Intrastriatal ΔRR administration (2500 plaque forming units) was nontoxic and did not induce microglial activation 5 days after injection. Intrastriatal injection of ΔRR with NMDA or 4 h after NMDA injection showed increased neuronal survival and decreased mitochondrial damage compared to injection of NMDA alone. Neuroprotection was due to the inhibition of NMDA-induced apoptosis through ERK activation. ΔRR-treated mice did not develop NMDA-associated behavioral deficits. The data suggest that ΔRR is a promising platform for treatment of acute neuronal injury.

Methylene Blue Decreases Ischemia-Reperfusion (I/R)-Induced Spinal Cord Injury: An in vivo Study in an I/R Rabbit Model

Objectives: To evaluate the effects of intravenous methylene blue (MB) administration on ischemia-reperfusion (I/R) injury of the spinal cord (SC). Methods: 16 rabbits were randomly assigned either to group M (n = 8; receiving MB, intervention group) or group C (n = 8; control group) and underwent a 30-min period of SC ischemia by clamping the abdominal aorta between the left renal artery and the aortic bifurcation. 15 min before clamping, rabbits received either intravenous MB (10 mg/kg; group M) or normal saline (group C). The two groups were compared 24 h postoperatively both histologically and for neurological function, using a Tarlov score. Measurements to determine levels of malondialdehyde (MDA) and glutathione (GSH) in the SC tissue were also performed. Results: Neurological impairment and spinal tissue MDA levels were significantly lower in animals treated with MB (p < 0.001). In contrast, spinal GSH levels were significantly higher in group M (p < 0.001). Histological examination revealed that the integrity of the SC was better preserved in the MB group, whereas cords from the control group exhibited evidence of acute neuronal injury. Conclusions: The prophylactic use of MB reduces neurological injury and improves clinical outcomes in the rabbit SC I/R model. These effects are probably mediated by the drug’s antioxidant properties.

From tau to toxicity: Emerging roles of NMDA receptor in Alzheimer's disease

Glutamate toxicity through NMDA receptor channels has long been central to the understanding of acute neuronal injury. Recent studies implicate similar events in chronic neurodegenerative diseases. Here, we analyze some of the most intriguing evidence for NMDA receptor-mediated cellular dysfunction and propose a mechanism by which hyperactive NMDA receptors might lead to neurofibrillary degeneration in Alzheimer's disease.

Association of Serial Biochemical Markers With Acute Ischemic Stroke

Biochemical markers of acute neuronal injury may aid in the diagnosis and management of acute ischemic stroke. Serum samples from the National Institute for Neurological Disorders and Stroke (NINDS) recombinant tissue plasminogen activator Stroke Study were analyzed for the presence of 4 biochemical markers of neuronal, glial, and endothelial cell injury. These biochemical markers, myelin basic protein (MBP), neuron-specific enolase (NSE), S100beta, and soluble thrombomodulin, were studied for an association with initial stroke severity, infarct volume, and functional outcome. In the original NINDS study, serum samples were drawn from all patients on presentation to the Emergency Department and at approximately 2 and 24 hours after initiation of study therapy. In this analysis, stored serum samples were available for 359 patients; 107 patients had samples for all 3 time points. Serum marker concentrations were measured by ELISA techniques. We examined the relation between serum concentrations of each marker and the degree of baseline neurological deficit, functional outcome, and infarct size on computed tomography at 24 hours and the effect of fibrinolytic therapy. Higher 24-hour peak concentrations of MBP, NSE, and S100beta were associated with higher National Institutes of Health Stroke Scale baseline scores (r=0.186, P<0.0001; r=0.117, P=0.032; and r=0.263, P<0.0001, respectively). Higher peak concentrations of MBP and S100beta (r=0.209, P<0.0001; r=0.239, P<0.0001) were associated with larger computed tomography lesion volumes. Patients with favorable outcomes had smaller changes in MBP and S100beta (P<0.05) concentrations in the first 24 hours. Soluble thrombomodulin was not associated with any severity or outcome measure. This study corroborates previous work demonstrating correlations of MBP, NSE, and S100beta with clinical and radiographic features in acute stroke. Despite significantly better outcomes in the tissue plasminogen activator-treated group, we found no difference in the early release of the 4 biomarkers between treatment groups. Further study will define the role of biomarkers in acute stroke management and prognostication.

Neurochemical Aftermath of Amateur Boxing

Little solid information is available on the possible risks for neuronal injury in amateur boxing. To determine whether amateur boxing and severity of hits are associated with elevated levels of biochemical markers for neuronal injury in cerebrospinal fluid. Longitudinal study. Referral center specializing in evaluation of neurodegenerative disorders. Fourteen amateur boxers (11 men and 3 women) and 10 healthy male nonathletic control subjects. The boxers underwent lumbar puncture 7 to 10 days and 3 months after a bout. The control subjects underwent LP once. Neurofilament light protein, total tau, glial fibrillary acidic protein, phosphorylated tau, and beta-amyloid protein 1-40 (Abeta([1-40])) and 1-42 (Abeta([1-42])) concentrations in cerebrospinal fluid were measured. Increased levels after a bout compared with after 3 months of rest from boxing were found for 2 markers for neuronal and axonal injury, neurofilament light protein (mean +/- SD, 845 +/- 1140 ng/L vs 208 +/- 108 ng/L; P = .008) and total tau (mean +/- SD, 449 +/- 176 ng/L vs 306 +/- 78 ng/L; P = .006), and for the astroglial injury marker glial fibrillary acidic protein (mean +/- SD, 541 +/- 199 ng/L vs 405 +/- 138 ng/L; P = .003). The increase was significantly higher among boxers who had received many hits (>15) or high-impact hits to the head compared with boxers who reported few hits. In the boxers, concentrations of neurofilament light protein and glial fibrillary acidic protein, but not total tau, were significantly elevated after a bout compared with the nonathletic control subjects. With the exception of neurofilament light protein, there were no significant differences between boxers after 3 months of rest from boxing and the nonathletic control subjects. Amateur boxing is associated with acute neuronal and astroglial injury. If verified in longitudinal studies with extensive follow-up regarding the clinical outcome, analyses of cerebrospinal fluid may provide a scientific basis for medical counseling of athletes after boxing or head injury.

Chronically Increased Transforming Growth Factor-β1 Strongly Inhibits Hippocampal Neurogenesis in Aged Mice

There is increasing evidence that hippocampal learning correlates strongly with neurogenesis in the adult brain. Increases in neurogenesis after brain injury also correlate with improved outcomes. With aging the capacity to generate new neurons decreases dramatically, both under normal conditions and after injury. How this decrease occurs is not fully understood, but we hypothesized that transforming growth factor (TGF)-beta1, a cell cycle regulator that rapidly increases after injury and with age, might play a role. We found that chronic overproduction of TGF-beta1 from astrocytes almost completely blocked the generation of new neurons in aged transgenic mice. Even young adult TGF-beta1 mice had 60% fewer immature, doublecortin-positive, hippocampal neurons than wild-type littermate controls. Bromodeoxyuridine labeling of dividing cells in 2-month-old TGF-beta1 mice confirmed this decrease in neuro-genesis and revealed a similar decrease in astrogenesis. Treatment of early neural progenitor cells with TGF-beta1 inhibited their proliferation. This strongly suggests that TGF-beta1 directly affects these cells before their differentiation into neurons and astrocytes. Together, these data show that TGF-beta1 is a potent inhibitor of hippocampal neural progenitor cell proliferation in adult mice and suggest that it plays a key role in limiting injury and age-related neurogenesis.

P1-370: Monitoring the biochemical aftermath of boxing by cerebrospinal fluid analyses

There is an increasing body of evidence that professional boxing can lead to acute and chronic brain injury with dementia. The possible risks of amateur boxing are largely unknown. We aimed at investigating levels of biochemical markers specific for central nervous system damage in cerebrospinal fluid (CSF) from amateur boxers. We collected CSF from 14 boxers (11 males and 3 females, age 22 years SD=3.8) one week after boxing and following a 3 month resting period. CSF was analyzed for total–tau (T–tau), hyperphosphorylated–tau (P–tau), Aβ40 and Aβ42, neurofilament–light (NFL) and glial fibrillary acidic protein (GFAP). Healthy non–boxers (n=10, all male, age 30 years, SD=6.3) were included as controls. Mean levels of GFAP and NFL, markers for astrocytes and neuronal axons, were significantly increased after bout (541, SD=199 and 845.0, SD=1143, respectively) compared with after rest (405, SD=138 and 208, SD=108, p=0.003 and p=0.008, respectively) and controls (402, SD=88.8 ng/L and <125 ng/L, p=0.037 and p<0.0001, respectively). The increase in CSF NFL after bout was also related to number and severity of hits. No significant differences after fight or rest were found for any of the other biomarkers, except for T–Tau which was higher after bout compared with rest (449, SD=176 and 306, SD=78.1 ng/L, respectively, p=0.003). To our knowledge, this is the first study showing that acute neuronal and axonal injury caused by amateur boxing can be directly identified and monitored. CSF analyses may serve as the scientific basis for medical counselling of athletes after boxing.

Cerebral Hypoxia and Ischemia: The Forensic Point of View: A Review

In cases with suspected brain anoxia/ischemia and hypoxia/hypoxemia a neuropathological investigation should give additional information to elucidate the cause of death and its pathophysiological mechanisms. Primary ischemic brain damage is associated with morphological and biochemical alterations. While acute ischemic neuronal injury reveals axon sparing and selective neuronal lesions due to the release of large quantities of glutamate, late neuronal death is associated with antiapoptotic growth factors, and decreased expression of microtubule-associated proteins and tubulin. On the morphological level ischemia can be detected by necrosis of neurons, proliferation of microglia, and astrocytes in vulnerable regions of the brain. In cases of permanent ischemia the so-called pale nervous cell injury is observed, in cases of partial perfusion the so-called dark nerve cell injury and apoptosis are detectable. In spite of the considerable advantages of recent research, presently there is no reliable qualitative marker to ascertain death due to acute hypoxic or ischemic events.

Feasibility and Safety of Delivering Xenon to Patients Undergoing Coronary Artery Bypass Graft Surgery While on Cardiopulmonary Bypass

Postoperative neurocognitive deficit is prevalent after cardiac surgery. Xenon may prevent or ameliorate acute neuronal injury, but it also may aggravate injury during cardiac surgery by increasing bubble embolism. Before embarking on a randomized clinical trial to test the safety and efficacy of xenon for postoperative neurocognitive deficit, we undertook a phase I study to investigate the safety of administering xenon to patients undergoing coronary artery bypass grafting while on cardiopulmonary bypass and to assess the practicability of our xenon delivery system. Sixteen patients scheduled for coronary artery bypass grafting surgery with hypothermic cardiopulmonary bypass gave their informed consent to participate in an open-label dose-escalation study (0, 20, 35, 50% xenon in oxygen and air). Xenon was delivered throughout surgery using both a standard anesthetic breathing circuit and the oxygenator. Gaseous and blood xenon partial pressures were measured five times before, during, and after cardiopulmonary bypass. Middle cerebral artery Doppler was used to assess embolic load, and major organ system function was assessed before and after surgery. Middle cerebral artery Doppler showed no evidence of increased emboli with xenon. Patients receiving xenon had no major organ dysfunction: Troponin I and S100beta levels tended to be lower in patients receiving xenon. Up to 25 l xenon was used per patient. Xenon partial pressure in the blood tracked the delivered concentration throughout. Xenon was safely and efficiently delivered to coronary artery bypass grafting patients while on cardiopulmonary bypass. Prevention of nervous system injury by xenon should be tested in a large placebo-controlled, randomized clinical trial.