Hypoxic-ischemic Stroke Research Articles

Postmortem radiological imaging and autopsy of a manually strangled dog with a neck twist

• Tracheal rupture with ischemic stroke was documented in a dog abuse case. • Excessive gas buildup caused by an air hunger effect was discussed. • PMCT was more appropriate to study the subtle bone structures to diagnose fractures. • Radiological imaging assists in determining a cause and manner of death in prosecuting animal crime. Forensic imaging studies were used to supplement the postmortem examination of a poodle-type dog that died of an unknown cause in an animal crime investigation. The carcass was powdered and the neck was twisted to the left. At the death scene, the police felt crepitation while moving the carcass to the autopsy room. They wondered whether the ribs and cervical bones of the carcass had been broken. Conventional X-rays revealed air pockets along the trachea and also an abundance of air in the subcutaneous tissue and body cavities. Three-dimensional volume-rendered images created from postmortem computed tomography confirmed that the hyoid bone, cervical vertebrae, and ribs were all intact. The findings during the autopsy were focally extensive intradermal and intramuscular hemorrhages and bruises on the right side of the body, tracheal rupture, and substantial brain congestion and edema. The results of the imaging and autopsy findings referred to asphyxiation and hypoxic ischemic stroke in the manner of death caused by blunt force trauma to the neck. During the police inquiry, the accused confessed to strangling the dog with her bare hands.

Astrocytes Protect Human Brain Microvascular Endothelial Cells from Hypoxia Injury by Regulating VEGF Expression.

Hypoxic-ischemic stroke has been associated with changes in neurovascular behavior, mediated, in part, by induction of the vascular endothelial growth factor (VEGF). The objective of this study was to investigate the effects of human astrocytes on the proliferation, apoptosis, and function of human microvascular endothelial cells (hBMEC) in vitro. Human microvascular endothelial cells (hBMEC) and human normal astrocytes (HA-1800) were used to establish in vitro cocultured cell models. The coculture model was used to simulate hypoxic-ischemic stroke, and it was found that astrocytes could promote hBMEC proliferation, inhibit apoptosis, reduce cell damage, and enhance antioxidant capacity by activating the VEGF signaling pathway. When VEGF is knocked out in astrocytes, the protective effect of astrocytes on hBMEC was partially lost. In conclusion, our study confirms the protective effect of hBMEC and laid a foundation for the study of hypoxic-ischemic stroke.

BDNF Pretreatment of Human Embryonic-Derived Neural Stem Cells Improves Cell Survival and Functional Recovery after Transplantation in Hypoxic–Ischemic Stroke

Intra-arterial neural stem cell (NSC) therapy has the potential to improve long-term outcomes after stroke. Here we evaluate if pretreatment of NSCs with brain-derived neurotrophic factor (BDNF) prior to transplantation improves cell engraftment and functional recovery following hypoxic-ischemic (HI) stroke. Human embryonic-derived NSCs with or without BDNF pretreatment (1 h, 100 ng/ml) were transplanted 3 days after HI stroke. Functional recovery was assessed using the horizontal ladder test. Cell engraftment was evaluated using bioluminescence imaging (BLI) and histological counts of SC121(+) cells. Fluoro-Jade C (FJC) and NeuN stains were used to evaluate neuroprotection. The effect of BDNF on NSCs was analyzed using a migration assay, immunocytochemistry, Luminex proteomic assay, and RT-qPCR.BLI analysis demonstrated significantly higher photon flux in the BDNF-treated NSC group compared to untreated NSC (p = 0.049) and control groups (p = 0.0021) at 1 week after transplantation. Immunohistochemistry confirmed increased transplanted cell survival in the cortex (p = 0.0126) and hippocampus (p = 0.0098) of animals injected with BDNF-treated NSCs compared to untreated NSCs. Behavioral testing revealed that the BDNF-treated NSC group demonstrated increased sensorimotor recovery compared to the untreated NSC and control groups (p < 0.001) over the 1-month period (p < 0.001) following transplantation. A significant improvement in performance was found in the BDNF-treated NSC group compared to the control group at 14, 21, and 28 (p < 0.05) days after transplantation. The cortex and hippocampus of the BDNF-treated NSC group had significantly more SC121(+) NSCs (p = 0.0125, p = 0.0098), fewer FJC(+) neurons (p = 0.0370, p = 0.0285), and a higher percentage of NeuN(+) expression (p = 0.0354) in the cortex compared to the untreated NSC group. BDNF treatment of NSCs resulted in significantly greater migration to SDF-1, secretion of M-CSF, VEGF, and expression of CXCR4, VCAM-1, Thrombospondins 1 and 2, and BDNF. BDNF pretreatment of NSCs results in higher initial NSC engraftment and survival, increased neuroprotection, and greater functional recovery when compared to untreated NSCs.

Hippocampal Adult Neurogenesis Is Maintained by Neil3-Dependent Repair of Oxidative DNA Lesions in Neural Progenitor Cells

Accumulation of oxidative DNA damage has been proposed as a potential cause of age-related cognitive decline. The major pathway for removal of oxidative DNA base lesions is base excision repair, which is initiated by DNA glycosylases. In mice, Neil3 is the main DNA glycosylase for repair of hydantoin lesions in single-stranded DNA of neural stem/progenitor cells, promoting neurogenesis. Adult neurogenesis is crucial for maintenance of hippocampus-dependent functions involved in behavior. Herein, behavioral studies reveal learning and memory deficits and reduced anxiety-like behavior in Neil3(-/-) mice. Neural stem/progenitor cells from aged Neil3(-/-) mice show impaired proliferative capacity and reduced DNA repair activity. Furthermore, hippocampal neurons in Neil3(-/-) mice display synaptic irregularities. It appears that Neil3-dependent repair of oxidative DNA damage in neural stem/progenitor cells is required for maintenance of adult neurogenesis to counteract the age-associated deterioration of cognitive performance.

Stratification substantially reduces behavioral variability in the hypoxic–ischemic stroke model

Stroke is the most common cause of long-term disability, and there are no known drug therapies to improve recovery after stroke. To understand how successful recovery occurs, dissect candidate molecular pathways, and test new therapies, there is a need for multiple distinct mouse stroke models, in which the parameters of recovery after stroke are well defined. Hypoxic–ischemic stroke is a well-established stroke model, but behavioral recovery in this model is not well described. We therefore examined a panel of behavioral tests to see whether they could be used to quantify functional recovery after hypoxic–ischemic stroke. We found that in C57BL/6J mice this stroke model produces high mortality (approximately one-third) and variable stroke sizes, but is fast and easy to perform on a large number of mice. Horizontal ladder test performance on day 1 after stroke was highly and reproducibly correlated with stroke size (P < 0.0001, R2 = 0.7652), and allowed for functional stratification of mice into a group with >18% foot faults and 2.1-fold larger strokes. This group exhibited significant functional deficits for as long as 3 weeks on the horizontal ladder test and through the last day of testing on automated gait analysis (33 days), rotarod (30 days), and elevated body swing test (EBST) (36 days). No deficits were observed in an automated activity chamber. We conclude that stratification by horizontal ladder test performance on day 1 identifies a subset of mice in which functional recovery from hypoxic–ischemic stroke can be studied.

Delayed Administration of a Small Molecule Tropomyosin-Related Kinase B Ligand Promotes Recovery After Hypoxic–Ischemic Stroke

Stroke is the leading cause of long-term disability in the United States, yet no drugs are available that are proven to improve recovery. Brain-derived neurotrophic factor stimulates neurogenesis and plasticity, processes that are implicated in stroke recovery. It binds to both the tropomyosin-related kinase B and p75 neurotrophin receptors. However, brain-derived neurotrophic factor is not a feasible therapeutic agent, and no small molecule exists that can reproduce its binding to both receptors. We tested the hypothesis that a small molecule (LM22A-4) that selectively targets tropomyosin-related kinase B would promote neurogenesis and functional recovery after stroke. Four-month-old mice were trained on motor tasks before stroke. After stroke, functional test results were used to randomize mice into 2 equally, and severely, impaired groups. Beginning 3 days after stroke, mice received LM22A-4 or saline vehicle daily for 10 weeks. LM22A-4 treatment significantly improved limb swing speed and accelerated the return to normal gait accuracy after stroke. LM22A-4 treatment also doubled both the number of new mature neurons and immature neurons adjacent to the stroke. Drug-induced differences were not observed in angiogenesis, dendritic arborization, axonal sprouting, glial scar formation, or neuroinflammation. A small molecule agonist of tropomyosin-related kinase B improves functional recovery from stroke and increases neurogenesis when administered beginning 3 days after stroke. These findings provide proof-of-concept that targeting of tropomyosin-related kinase B alone is capable of promoting one or more mechanisms relevant to stroke recovery. LM22A-4 or its derivatives might therefore serve as "pro-recovery" therapeutic agents for stroke.

Abstract 199: Pre-treatment Of Human Embryonic-derived Neural Progenitor Cells With Bdnf Increases Migration Of Cells To The Brain And Elicits Functional Recovery In A Mouse Model Of Hypoxic-ischemia

Intro: Increasing cell migration through upregulation of chemokine and adhesion molecule receptors could improve intravascular cell treatment for stroke and BDNF has been shown to induce these pathways. Therefore we tested whether BDNF cell-pretreatment would improve cell migration and functional recovery in an experimental stroke model. Methods: hES-derived NPCs (5x10 5 in 5µl saline) pre-treated with BDNF for 5 hours and harboring a reporter gene construct containing renilla luciferase and eGFP in serum free media, non-treated hES-derived NPCs (5×10 5 in 5[l saline) in media, and media control with BDNF were delivered to the brain via the ipsilateral carotid artery at 3 days after hypoxic-ischemic stroke in NODSCID mice (n=11/group). Cell engraftment was monitored by in-vivo bioluminescence imaging (BLI). The ladder test was used to assess behavioral recovery throughout a 4 week time course. Brain homogenates from animals at 28 days were analyzed using RT-qPCR for common chemokines, adhesion molecules, and neurotrophins. Mechanisms of cell migration were evaluated by assessing cell receptor expression of chemokines and adhesion molecules on hES-derived NPC and by analyzing the change in expression profile in the mouse brain at 3 days after stroke. Boyden-chamber migration assays were used to evaluate cell migratory potential in vitro . RESULTS: One day after cell transplantation the subset of animals transplanted with BDNF-pretreated cells showed significantly higher BLI signal at 1 (p=0.021) and 7 days after transplantation (p=0.002). Histological analysis also revealed engraftment of hES-derived NPC at 1 week after transplantation. Behavioral assessment revealed significant functional recovery in the BDNF pre-treated group throughout the 28 day time course (ANOVA, p&lt;0.05). BDNF-pretreatment of hES-derived NPCs upregulated CXCR4 expression 12.5 times and in vitro led to significantly greater migration in response to CXCL12 (CXCR4 ligand) compared to untreated cells. At 28 days after transplantation, neurotrophic factors IL6, IL10, Ntrk1 were upregulated 3.3, 3.4, and 3.3 times. Common T-cell and neutrophil cytokine receptors IL8rb, IL8ra and IL1a were all downregulated, while several chemokines that increase migration of inflammatory cells were downregulated including CCL2, CCL5, CCL8, and CCL12. Anti-Angiogenic factor Adamts8 was also downrgulated in the brains of animals transplanted with BDNF pre-treated cells. Lastly, MMP3, MMP8, and MMP9 were downregulated at 28 days after stroke indicating increased blood brain barrier integrity. Conclusion: Intravascular transplantation of BDNF pre-treated hES-derived NPCs elicits functional gains via increased migration of cells, immunomodulation, increased BBB integrity, and by influencing the upregulation of neur0protective factors.

Exogenous growth hormone attenuates cognitive deficits induced by intermittent hypoxia in rats

Sleep disordered breathing (SDB), which is characterized by intermittent hypoxia (IH) during sleep, causes substantial cardiovascular and neurocognitive complications and has become a growing public health problem. SDB is associated with suppression of growth hormone (GH) secretion, the latter being integrally involved in the growth, development, and function of the CNS. Since GH treatment is able to attenuate neurocognitive deficits in a hypoxic-ischemic stroke model, GH, GH receptor (GHR) mRNA expression, and GH protein expression were assessed in rat hippocampus after exposures to chronic sustained hypoxia (CH, 10% O(2)) or IH (10% O(2) alternating with 21% O(2) every 90 s). In addition, the effect of GH treatment (50 μg/kg daily s.c. injection) on erythropoietin (EPO), vascular endothelial growth factor (VEGF), heme oxygenase-1 (HO-1), and GLUT-1 mRNA expression and neurobehavioral function was assessed. CH significantly increased GH mRNA and protein expression, as well as insulin-like growth factor-1 (IGF-1). In contrast, IH only induced a moderate increase in GH mRNA and a slight elevation in GH protein at day 1, but no increases in IGF-1. CH, but not IH, up-regulated GHR mRNA in the hippocampus. IH induced marked neurocognitive deficits compared with CH or room air (RA). Furthermore, exogenous GH administration increased hippocampal mRNA expression of IGF-1, EPO, and VEGF, and not only reduced IH-induced hippocampal injury, but also attenuated IH-induced cognitive deficits. Thus, exogenous GH may provide a viable therapeutic intervention to protect IH-vulnerable brain regions from SDB-associated neuronal loss and associated neurocognitive dysfunction.

Iron in the Brain

After completing this article, readers should be able to: 1. Describe the role of iron in proteins involved in neuronal and glial energy metabolism, myelin production, and monoamine neurotransmitter metabolism. 2. Indicate the neonates who are at risk for iron deficiency. 3. Explain how iron deficiency affects the biochemistry, structure, signaling molecules, and electrophysiology of the hippocampus. 4. Describe the primary causes of iron overload in the neonate. 5. Explain why the preterm infant is at particular risk for iron deficiency. This article is designed to acquaint readers with the role of iron in early brain development. Several important principles that govern the effects of nutrients on the immature brain are discussed, followed by an explanation of the roles of iron during development. Because the developing brain is sensitive to iron overload as well as iron deficiency, both are discussed in the context of human studies and in model systems. The first guiding principle of nutrient-brain interactions during development is the concept of timing, dose, and duration. (1) The positive or negative effects of any nutrient depend on which brain regions or processes are developing at the time that nutrient status is altered and the requirements of those regions for the nutrient during that time of development. Generally, the more rapidly an area is developing, the more vulnerable it is. Dependence on specific nutrients during phases of rapid proliferation or differentiation results in the second principle, a regionalization of brain effects during periods of nutrient deficiency. In the case of iron deficiency, certain brain areas that are dependent on iron for proper development are more vulnerable than areas that are less dependent on iron. Additionally, these same brain areas are likely to be more vulnerable at certain time points than at others. To exert a maximal effect on regional brain development, the timing of iron …

Bax expression in mammalian neurons undergoing apoptosis, and in Alzheimer's disease hippocampus

Recent studies indicate that the proto-oncogene Bax, and other related proteins (eg Bcl-2) may play a major role in determining whether cells will undergo apoptosis under conditions which promote cell death. Increased expression of Bax has been found to promote apoptosis, while over-expression of Bcl-2 can inhibit apoptosis. To investigate the role of Bax in nerve cell death in the rat brain we examined the level of Bax expression in cells undergoing apoptosis, using a hypoxic-ischemic stroke model. We found that Bax was expressed at high levels in the nuclei of neurons in the hippocampus, cortex, cerebellum, and striatum on the control side, and that Bax levels increased in hippocampal neurons undergoing apoptosis on the stroke side, and then declined (correlating with cell loss). In the Alzheimer's disease hippocampi we found a concentrated localisation of Bax in senile plaques, which correlated with the localisation of β-amyloid protein in adjacent sections from the same brains. β-Amyloid positive plaques are thought to contribute to the Alzheimer's disease process, possibly via an apoptotic mechanism, and this may occur via an increase in Bax in these areas. Bax was also strongly stained in τ-positive tangles in Alzheimer's disease hippocampi, suggesting Bax may play a role in tangle formation. In addition, we observed a loss of Bax expression in the dentate granule cells of Alzheimer's disease hippocampi compared with moderate Bax expression in control hippocampi, and this loss may be related to the survival of these neurons in Alzheimer's disease. Finally, we observed substantially different staining patterns of Bax using three different commercially available antisera to Bax, indicating the need for caution when interpreting results in this area.