Total Infarct Volume Research Articles

The selective estrogen receptor modulator, bazedoxifene, reduces ischemic brain damage in male rat

While the estrogen treatment of stroke is under debate, selective estrogen receptor modulators (SERMs) arise as a promising alternative. We hypothesize that bazedoxifene (acetate, BZA), a third generation SERM approved for the treatment of postmenopausal osteoporosis, reduces ischemic brain damage in a rat model of transient focal cerebral ischemia. For comparative purposes, the neuroprotective effect of 17β-estradiol (E2) has also been assessed. Male Wistar rats underwent 60min middle cerebral artery occlusion (intraluminal thread technique), and grouped according to treatment: vehicle-, E2- and BZA-treated rats. Optimal plasma concentrations of E2 (45.6±7.8pg/ml) and BZA (20.7±2.1ng/ml) were achieved 4h after onset of ischemia, and maintained until the end of the procedure (24h). Neurofunctional score and volume of the damaged brain regions were the main end points. At 24h after ischemia–reperfusion, neurofunctional examination of the animals did not show significant differences among the three experimental groups. By contrast, both E2- and BZA-treated groups showed significantly lower total infarct volumes, BZA acting mainly in the cortical region and E2 acting mainly at the subcortical level. Our results demonstrate that: (1) E2 at physiological plasma levels in female rats is neuroprotective in male rats when given at the acute stage of the ischemic challenge and (2) BZA at clinically relevant plasma levels mimics the neuroprotective action of E2 and could be, therefore, a candidate in stroke treatment.

Bradykinin postconditioning ameliorates focal cerebral ischemia in the rat

The goal of this study is to investigate the effects of bradykinin (BR) postconditioning on cerebral ischemic injury. Transient focal cerebral ischemia was induced in rats by 60min of middle cerebral artery occlusion (MCAO), followed by 3days of reperfusion. BR as a postconditioner at a dose of 150μg/kg was applied intraperitoneally 3, 6, 24 and 48h after MCAO. BR postconditioning significantly reduced total infarct volumes if applied 3h after MCAO by 95%, 6h after MCAO by 80% and 24h after MCAO by 70% in versus vehicle group. Neurological functions were amarked improvement in the BR groups compared to the ischemia group. The number of degenerated neurons in the hippocampal CA1 region was also significantly lower in BR-treated ischemic groups compared to vehicle group. BR postconditioning prevented the release of MnSOD from the mitochondria and reduced the activity of the total SOD and CAT if it is administrated short time after stroke. Our data proves the ischemic tolerance in the brain induced by BR postconditioning resulted as effective agent against as strong an attack as 60min MCAO even when used many hours after ischemia.

Hydrogen Sulfide Protects the Brain Against Ischemic Reperfusion Injury in a Transient Model of Focal Cerebral Ischemia

Hydrogen sulfide (H(2)S), a well-known toxic gas, is regarded as endogenous neuromodulator and plays multiple roles in the central nervous system under physiological and pathological states, especially in secondary neuronal injury. Recent studies have shown relatively high concentrations of hydrogen sulfide (H(2)S) in the brain and also cytoprotective effects of endogenous and exogenous H(2)S in models of in vitro and in vivo ischemic injury. H(2)S protects neurons by functioning as an anti-oxidant, anti-inflammatory, and anti-apoptotic mediator and by improving neurological function. Moreover, it protects neurons from glutamate toxicity. Therefore, the present study aimed to determine whether H(2)S provides protection in transient focal cerebral ischemia. Focal ischemia was induced by 60-min middle cerebral artery occlusion (MCAO), followed by 23-h reperfusion. Saline as a vehicle and NaHS (H(2)S donor; 1 and 5 mg) were intraperitoneally injected (IP) at the beginning of ischemia. Infarct volume, brain edema, and apoptosis were assessed 24 h after MCAO.Treatment with NaHS at doses of 1 and 5 mg markedly reduced total infarct volumes by 29 and 51 %, respectively (P < 0.001). In addition, NaHS at doses of 1 and 5 mg reduced brain edema (P < 0.05) and inhibited apoptosis by decreasing positive TUNEL cells (P < 0.001).The present study shows that treatment with H(2)S reduces brain injuries and postischemic cerebral edema in a dose-dependent manner likely through the blocking programmed cell death.We propose that H(2)S might be a promising therapeutic target for stroke, although more researches are necessary to take into account the potential therapeutic effects of H(2)S in stroke patients.

PD-L1 enhances CNS inflammation and infarct volume following experimental stroke in mice in opposition to PD-1

BackgroundStroke severity is worsened by recruitment of inflammatory immune cells into the brain. This process depends in part on T cell activation, in which the B7 family of co-stimulatory molecules plays a pivotal role. Previous studies demonstrated more severe infarcts in mice lacking programmed death-1 (PD-1), a member of the B7 family, thus implicating PD-1 as a key factor in limiting stroke severity. The purpose of this study was to determine if this protective effect of PD-1 involves either of its ligands, PD-L1 or PD-L2.MethodsCentral nervous system (CNS) inflammation and infarct volume were evaluated in male PD-L1 and PD-L2 knockout (-/-) mice undergoing 60 minutes of middle cerebral artery occlusion (MCAO) followed by 96 hours of reperfusion and compared to wild-type (WT) C57BL/6J mice.ResultsPD-L1-/- and PD-L2-/- mice had smaller total infarct volumes compared to WT mice. The PD-L1-/- and to a lesser extent PD-L2-/- mice had reduced levels of proinflammatory activated microglia and/or infiltrating monocytes and CD4+ T cells in the ischemic hemispheres. There was a reduction in ischemia-related splenic atrophy accompanied by lower activation status of splenic T cells and monocytes in the absence of PD-L1, suggesting a pathogenic rather than a regulatory role for both PD-1 ligands (PD-Ls). Suppressor T cells (IL-10-producing CD8+CD122+ T cells) trafficked to the brain in PD-L1-/- mice and there was decreased expression of CD80 on splenic antigen-presenting cells (APCs) as compared to the WT and PD-L2-/- mice.ConclusionsOur novel observations are the first to implicate PD-L1 involvement in worsening outcome of experimental stroke. The presence of suppressor T cells in the right MCAO-inflicted hemisphere in mice lacking PD-L1 implicates these cells as possible key contributors for controlling adverse effects of ischemia. Increased expression of CD80 on APCs in WT and PD-L2-/- mice suggests an overriding interaction leading to T cell activation. Conversely, low CD80 expression by APCs, along with increased PD-1 and PD-L2 expression in PD-L1-/- mice suggests alternative T cell signaling pathways, leading to a suppressor phenotype. These results suggest that agents (for example antibodies) that can target and neutralize PD-L1/2 may have therapeutic potential for treatment of human stroke.

In vivo monitoring of arterially transplanted bone marrow mononuclear cells in a rat transient focal brain ischemia model using magnetic resonance imaging

Objectives: Intra-arterial transplantation of bone marrow mononuclear cells (BMMCs) effectively improves neuronal function and limits the infarct size. We monitored the fate of BMMCs labeled with super paramagnetic iron oxide (SPIO) until 7 days after the transplantation using high-field magnetic resonance imaging (MRI).Methods: Male Sprague–Dawley rats were subjected to 90-minute focal ischemia using the intraluminal suture technique followed by transplantation of 1×107 BMMCs or vehicle only via the ipsilateral carotid artery immediately after reperfusion. Autologous BMMCs were labeled with SPIO by electroporation prior to ischemia. MRI studies were performed at 1 hour, 24 hours, 3 days, and 7 days after reperfusion on each rat. The total infarct volume and the volume of negative dots were measured on T2-weighted images and T2*-weighted images, respectively.Results: One hour after BMMC transplantation, we confirmed wide spread distribution of BMMCs in the ischemic hemisphere as a negative dot. The volume of negative dot normalized by hemispheric volume decreased rapidly and was seldom seen at the seventh day after transplantation. The infarct volume was significantly smaller in the transplanted group than the vehicle group at 24 hours and 7 days after reperfusion.Discussion: The present study established in vivo monitoring of intra-arterial transplanted SPIO-labeled BMMCs immediately after reperfusion using MRI of a rat transient focal ischemia model. The accumulation of BMMCs in ischemic lesion at the acute stage of ischemia can be part of the conditions to limit the infarct size.

The influence of aging on poststroke depression using a rat model via middle cerebral artery occlusion

Poststroke depression (PSD) is the most frequent psychological sequela following stroke. While previous studies describe the impact of age on brain infarct volume, brain edema, and blood-brain barrier (BBB) breakdown following ischemia, the role of age on PSD has yet to be described. Here, we examine the influence of age on PSD progression in a rat model of PSD by middle cerebral artery occlusion (MCAO). One hundred forty-three rats were divided into three groups. 48 rats 20 weeks of age underwent a sham procedure, 51 rats 20 weeks of age had MCAO, and 44 rats 22-26 months of age had MCAO. Groups were further divided into two subgroups. The first subgroup was used to measure infarct lesion volume, brain edema, and BBB breakdown at 24 h. In the second subgroup at 3 weeks after MCAO, rats were subjected to a sucrose preference test, two-way shuttle avoidance task, forced swimming test, and a brain-derived neurotrophic factor (BDNF) protein level measurement. Total and striatal infarct volume, brain edema, and BBB breakdown in the striatum were increased in older rats, as compared with younger rats. While both old and young rats exhibited depressive-like behaviors on each of the behavioral tests and lower BDNF levels post-MCAO, as compared with control rats, there were no differences between old and young rats. Although older rats suffered from larger infarct volumes, increased brain edema and more BBB disruption following MCAO, the lack of behavioral differences between young and old rats suggests that there was no effect of rat age on the incidence of PSD.

Reperfusion Is a Stronger Predictor of Good Clinical Outcome than Recanalization in Ischemic Stroke

To assess the predictive value of reperfusion indices, recanalization, and important baseline clinical and radiologic scores for good clinical outcome prediction. The study was approved by the local research ethics board. Written consent was obtained from all participants or their caregivers. Baseline computed tomography (CT) perfusion less than 4.5 hours after stroke symptoms, follow-up CT perfusion at 24 hours or less, and 5-7-day magnetic resonance images were obtained for 114 patients. Baseline imaging was assessed blinded to outcome. Recanalization status was determined at follow-up CT angiography. Reperfusion index was calculated on baseline and on follow-up at-risk tissue volume. Kruskal-Wallis, Mann-Whitney rank sum, and Spearman correlation were used for group comparisons and correlation studies. Univariate and multivariate logistic regression tested the association of clinical and imaging parameters with good outcome. Models with and without recanalization and reperfusion were compared by using Akaike information criterion. Reperfusion indices were significantly higher in patients with recanalization than in those without (P < .001). Despite significance of recanalization at univariate analysis, only reperfusion, age, and National Institutes of Health Stroke Scale score were significant after multivariate analysis (P < .01). Time to maximum reperfusion index had the highest accuracy (area under the receiver operating characteristic curve, 0.70) for good outcome, and reperfusion was defined as time to maximum volume of 59% or greater. Patients with reperfusion but no recanalization had significantly lower total infarct volume (P = .001) and infarct growth (P = .004) and had higher salvaged penumbra (P = .009) volumes than patients without reperfusion and recanalization. A final model with reperfusion but not recanalization was the most prognostic model of good clinical outcome. Reperfusion showed stronger association with good clinical outcome than did recanalization.

A Model of Rat Embolic Cerebral Infarction with a Quantifiable, Autologous Arterial Blood Clot

We developed a novel model of a rat embolic cerebral infarction with a quantifiable autologous arterial blood clot. The left femoral artery had 0.15 ml of blood withdrawn and mixed with 10 units of thrombin in 50 μl saline. After 30 min, the clot was suctioned into a 4-French polyvinyl chloride tube. A 24-gage catheter was inserted up through the internal carotid artery via the external carotid artery stump. The 1-cm clot, at a volume of 7.2 mm3, was pushed and inserted into the internal carotid artery via the catheter. After withdrawing the catheter, the ICA blood flow recovered. We checked neurological status after 24 h (neurological free was 15, and worst was 1) and measured the infarction volume by the TTC method. Twelve rats were examined, and five sham-operated rats were included. Two rats were not able to achieve an 80% reduction in CBF. One rat died due to cerebral infarction. The success rate in producing infarction was 83%. The total infarction volume was 368.5 mm3±61.2 se. Median neurological score was 6. Hemorrhagic transformation was not detected. Sham-operated rats revealed no infarction and no neurological deficit. The volume of infarction correlated significantly with the neurological score. We conclude that this embolic stroke model is useful in producing a human, severe cardioembolic cerebral infarction.

Transcranial Laser Therapy and Infarct Volume

Two randomized trials suggested that transcranial laser therapy (TLT) may benefit patients with acute ischemic stroke, although efficacy has not been confirmed. Supportive proof of concept could be demonstrated if TLT reduces the volume of cortical infarction. The NeuroThera Efficacy and Safety Trial-2 (NEST-2) was a randomized trial of TLT versus sham in patients with acute ischemic stroke treated within 24 hours of onset. Infarct volumes were measured quantitatively and semiquantitatively on all protocol-required computed tomography (or MRI, if clinically indicated) scans performed on day 5 (±2). Two approaches assessed treatment effects on cortex: (1) indirectly, by analyzing total infarct volume among patients with clinical presentations suggesting cortical involvement; and (2) directly, by assessing the cortical Alberta Stroke Program Early CT Score (cASPECTS) components (M1-M6, anterior, posterior) on a 0- to 8-point modified scale. A total of 640 subjects had scans (576 computed tomography, 64 MRI) on day 5. The reliability of ASPECTS (intraclass correlation coefficient=0.85) and cASPECTS (intraclass correlation coefficient=0.82) was excellent, and total ASPECTS was correlated with total infarct volume (r=0.71). In the overall study population, there was no impact of TLT on total infarct volume (P=0.30), total ASPECTS (P=0.85), or cASPECTS (P=0.89). Similarly, no effect was seen in any of the following prespecified subgroups selected to indicate cortical involvement: baseline National Institutes of Health Stroke Scale score >10, Oxfordshire Total Anterior Circulation Syndrome, subjects with aphasia or extinction at baseline, or subjects with radiographic involvement of cortex. TLT was not associated with a reduction in overall or cortical infarct volume as measured on computed tomography in the subacute phase.

Opposing roles of PD-1 and PD-L in modulating CNS inflammation and infarct volume following experimental stroke in mice (P1012)

Abstract Stroke severity is worsened by recruitment of inflammatory immune cells into the brain. This process depends in part on T cell activation, in which the B7 family of co-stimulatory molecules plays a pivotal role. Previous studies demonstrated more severe infarcts in mice lacking Programmed Death-1 (PD-1), a member of the B7 family, thus implicating PD-1 as a key factor in limiting stroke severity. The purpose of this study was to determine if this protective effect of PD-1 involves either of its ligands, PD-L1 and PD-L2. Thus, CNS inflammation and stroke severity were evaluated in male PD-L1 and PD-L2 knockout (KO) mice undergoing 60 min middle cerebral artery occlusion (MCAO) followed by 96 h reperfusion. Unexpectedly, PD-L1 KO and to a lesser extent PD-L2 KO mice had reduced levels of activated microglia and infiltrating monocytes and total infarct volume compared to wild-type mice, as well as reduced splenic atrophy and MHC class II expression, suggesting a pathogenic rather than a regulatory role for both PD-ligands. Both positive and negative regulatory roles have been suggested for the PD-ligands in context of various diseases and our novel observations indicate that PD-L1 and PD-L2 might have proinflammatory functions independent of PD-1 binding that worsen stroke outcome. These results suggest the possibility of using antibodies to PD-L1/2 as a potential treatment for human stroke patients.

Improvement of the circulatory function partially accounts for the neuroprotective action of the phytoestrogen genistein in experimental ischemic stroke

We tested the hypothesis that the phytoestrogen genistein protects the brain against ischemic stroke by improving the circulatory function in terms of reduced production of thromboxane A2 and leukocyte–platelet aggregates, and of preserved vascular reactivity. Ischemia-reperfusion (90min-3 days, intraluminal filament) was induced in male Wistar rats, and functional score and cerebral infarct volume were the end points examined. Genistein (10mg/kg/day) or vehicle (β-cyclodextrin) was administered at 30min after ischemia or sham-operation. Production of thromboxane A2 and leukocyte–platelet aggregates, as well as reactivity of carotid artery to U-46619 (thromboxane A2 analogue) and to platelet releasate was measured. At 3 days post-ischemia, both improvement in the functional examination and reduction in the total infarct volume were shown in the ischemic genistein-treated group. Genistein significantly reverted both the increased thromboxane A2 concentration and the increased leukocyte–platelet aggregates production found in samples from the ischemic vehicle-treated group. Both U-46619 and platelet releasate elicited contractions of the carotid artery, which were significantly lower in the ischemic vehicle-treated group. Genistein significantly restored both the decreased U-46619- and the decreased platelet releasate-elicited contractile responses. In conclusion, genistein protects the brain against an ischemia-reperfusion challenge, at least in part, by its beneficial effects on the circulatory function.

N-3 Fatty Acid Rich Triglyceride Emulsions Are Neuroprotective after Cerebral Hypoxic-Ischemic Injury in Neonatal Mice

We questioned if acute administration of n-3 fatty acids (FA) carried in n-3 rich triglyceride (TG) emulsions provides neuroprotection in neonatal mice subjected to hypoxic-ischemic (H/I) brain injury. We examined specificity of FA, optimal doses, and therapeutic windows for neuroprotection after H/I. H/I insult was induced in C57BL/6J 10-day-old mice by right carotid artery ligation followed by exposure to 8% O2 for 15 minutes at 37°C. Intraperitoneal injection with n-3-rich TG emulsions, n-6 rich TG emulsions or saline for control was administered at different time points before and/or after H/I. In separate experiments, dose responses were determined with TG containing only docosahexaenoic acid (Tri-DHA) or eicosapentaenoic acid (Tri-EPA) with a range of 0.1–0.375 g n-3 TG/kg, administered immediately after H/I insult. Infarct volume and cerebral blood flow (CBF) were measured. Treatment with n-3 TG emulsions both before- and after- H/I significantly reduced total infarct volume by a mean of 43% when administered 90 min prior to H/I and by 47% when administered immediately after H/I. In post-H/I experiments Tri-DHA, but not Tri-EPA exhibited neuroprotective effects with both low and high doses (p<0.05). Moreover, delayed post-H/I treatment with Tri-DHA significantly decreased total infarct volume by a mean of 51% when administered at 0 hr, by 46% at 1 hr, and by 51% at 2 hr after H/I insult. No protective effect occurred with Tri-DHA injection at 4 hr after H/I. There were no n-3 TG related differences in CBF. A significant reduction in brain tissue death was maintained after Tri-DHA injection at 8 wk after the initial brain injury. Thus, n-3 TG, specifically containing DHA, is protective against H/I induced brain infarction when administered up to 2 hr after H/I injury. Acute administration of TG-rich DHA may prove effective for treatment of stroke in humans.

The role of the mitochondrial calcium uniporter in cerebral ischemia/reperfusion injury in rats involves regulation of mitochondrial energy metabolism

The mitochondrial calcium uniporter (MCU) maintains intracellular Ca2+ homeostasis by transporting Ca2+ from the cell cytosol into the mitochondrial matrix and is important for shaping Ca2+ signals and the activation of programmed cell death. Inhibition of MCU by ruthenium red (RR) or Ru360 has previously been reported to protect against neuronal death. The aim of the present study was to analyze the mechanisms underlying the effects of MCU activity in a rat model of cerebral ischemia/reperfusion (I/R) injury. Adult male Wistar rats were divided into 4 groups; sham, I/R, I/R + RR and I/R + spermine (Sper) and were subjected to reversible middle cerebral artery occlusion for 2 h followed by 24 h of reperfusion. A bolus injection of RR administered 30 min prior to ischemia was found to significantly decrease the total infarct volume and reduce neuronal damage and cell apoptosis compared with ischemia/reperfusion values. However, treatment with Sper, an activator of the MCU, increased the injury induced by I/R. Analysis of energy metabolism revealed that I/R induced progressive inhibition of complexes I‑IV of the electron transport chain, decreased ATP production, dissipated the mitochondrial membrane potential and increased the generation of reactive oxygen species. Treatment with RR ameliorated the condition, while spermine had the opposite effect. In conclusion, blocking MCU was demonstrated to exert protective effects against I/R injury and this process may be mediated by the prevention of energy failure.

Abstract WMP68: Loss Of Programmed Cell Death Ligand 1 And 2 Reduces Total Infarct Volume In Male Mice Following Experimental Stroke

The programmed cell death 1 receptor (PD-1) is an immunoglobulin superfamily member whose expression is induced in activated T cells, B cells, natural killer cells and monocytes. Binding of PD-1 to either of its two known ligands, programmed cell death ligand 1 and 2 (PD-L1 and PD-L2), induces inhibitory signals that regulate peripheral T cell tolerance and immune homeostasis. We have shown increased PD-1 expression in brain macrophages and glia as well as increased PD-L1 and PD-L2 expression in peripheral and brain B cells in mice after experimental stroke. Furthermore, we have observed increased infarct volume in PD-1 deficient mice, thus potentially implicating the PD-1/PD-L co-inhibitory pathway in post-stroke CNS inflammation. Using PD-L1 and PD-L2 deficient mice, we determined if PD-L1 or PD-L2 was also important in modulating stroke infarct volume. Young adult male wild-type (WT; n=11) and PD-L1 (n=9) and PD-L2 (n=12) knockout (KO) mice underwent 60 min of middle cerebral artery occlusion via intraluminal filament. Laser-Doppler flowmetry was used to monitor cortical perfusion. All mice were euthanized and brains collected at 96 h of reperfusion. Total infarct volume (% corrected contralateral hemisphere) was determined by image analysis of coronal brain slices stained with 2,3,5-triphenyltetrazolium chloride (TTC). Relative laser Doppler flowmetry changes (% baseline) during and after middle cerebral artery occlusion were equivalent among groups. Compared to male WT mice (WT vs. KO), genetic deletion of either PD-L1 (51+3% vs. 20+5%; p&lt;0.001) or PD-L2 (51+3% vs. 35+4%; p=0.007) reduced total infarct volume. We also observed that total infarct volume was smaller in PD-L1 vs. PD-L2 KO mice (20+5% vs. 35+4%; p=0.017). Our findings along with our previous work in PD-1 deficient mice suggest that ligands other than PD-L1 or PD-L2 might be involved with PD-1 signaling in ischemic brain and that PD-L1 and PD-L2 might have other functions independent of PD-1 binding affecting infarct volumes. Future studies will evaluate the roles of PD-1, PD-L1, and PD-L2 in regulatory immune cell subsets that affect ischemic brain outcomes. This study was supported by National Institutes of Health grant NS075887.

Abstract WP274: Novel Platelet-Activating Factor Receptor Antagonists are Highly Neuroprotective in Focal Cerebral Ischemia

INTRODUCTION: Platelet-activating factor (PAF) is a bioactive phospholipid that regulates synaptic activity and when overproduced is a potent mediator of leukocyte functions, including platelet aggregation and inflammation. PAF concentration increases during ischemia-reperfusion and is a mediator of neurotoxicity. The inhibition of this process plays a critical role in neuronal survival. We synthesized various analogues and identified them as PAF receptor antagonists, which were then selected for in vivo experiments to evaluate anti-ischemic activity. METHODS: Physiologically controlled Sprague-Dawley rats received 2 h MCAo by poly-L-lysine-coated intraluminal suture. LAU compounds (LAU-0901, LAU-09015, LAU-09018, LAU-09019, LAU-09020 or LAU-09023; 30 mg/kg) or vehicle (45% cyclodextran) was administered IP at 2 h from onset of MCAo. Neurological status was evaluated during occlusion (at 60 min) and on day 1, 2, 3 and 7. Histopathology was carried out on day 7. RESULTS: Physiological variables were stable and showed no significant differences between groups. All LAU compounds significantly reduced the neurobehavioral deficit compared to the vehicle group, beginning from day 1 and persisting through the 7 day survival period. Histologically, the vehicle group showed large hemispheric infarcts with pan-necrosis. By contrast, LAU-treated rats showed preserved hemispheric structure and small subcortical infarcts. Treatment with LAU (LAU-0901, LAU-09015, LAU-09018, LAU-09019, or LAU-09023) significantly reduced total infarct volume compared to the vehicle group by 68%, 52%, 40%, 51% and 54%, respectively. CONCLUSION: Our results clearly indicate that novel LAU PAF receptor antagonists are highly neuroprotective on behavior and reduction of brain infarction in experimental stroke. These compounds may provide the basis for future therapeutics in patients suffering ischemic stroke. This study was supported by NIH, NINDS grant R01 NS046741 (NGB).

Granulocyte-colony stimulating factor reduces striatal dopamine accumulation caused by cerebral ischemia

ObjectivesGranulocyte-colony stimulating factor (G-CSF) plays important roles in neuroprotection. Acute and massive increases in extracellular dopamine can cause damage to the striatal neurons during cerebral ischemia (CI). We hypothesized that G-CSF may exert neuroprotective effects by reducing dopamine accumulation. Materials and MethodsCI was induced by infusing endothelin-1 (Et-1, 120 pmol in 10 μL saline) into the root of the middle cerebral artery in freely moving rats. G-CSF (200 μg/kg in 0.2 mL saline) was administered immediately after CI induction. ResultsG-CSF reduced dopamine accumulation in the ischemic striatum after CI was induced by Et-1. The total brain infarct volume was reduced by 67% (p < 0.05) at 24 hours after CI. ConclusionThese data suggest that reduced dopamine accumulation may be one of the mechanisms underlying the neuroprotective effects of G-CSF.

Abstracts for Canadian Stroke Conference

Specogna, AV; Patten, SB; Hill, MD; University of Calgary, Calgary, AB Background: Previous studies have examined predictors of treatment costs after Spontaneous Intracerebral Hemorrhage (ICH) in various countries and healthcare systems. Currently however, there is no clear understanding of which factors are associated with cost in any Canadian setting and thus no way of understanding how to plan ICH healthcare spending. Methods: We used a retrospective case series study design and adjusted linear regression to investigate if an association existed between the total cost of ICH hospital care in a Canadian health centre and age (years), gender (female vs. male), Charlson Comorbidity Index (0 vs. ≥1), in-hospital mortality (alive vs. dead), and having surgery (no vs. yes). Economic, treatment, and patient data were obtained from administrative sources. Total inflation-adjusted hospital cost per discharge was estimated in Canadian Dollars and log transformed for all analyses. Results: Analyses were performed using 987 consecutive ICH discharges from 1999 to 2008. The total cost of ICH hospital care was highly variable (mean cost per discharge=$25,613.96±$36,116.33; min $446.81 to max $326,275.10). Older age (β=–0.0057; 95% CI: –0.0103 to –0.0011) and death in hospital (β=–0.4859; 95% CI: –0.6357 to –0.3342) were significantly associated with lower total hospital cost per discharge. Whereas Charlson Comorbidity Index of ≥1 (β=0.6489; 95% CI: 0.5063 to 0.7916) and having surgery (β=1.3552; 95% CI: 1.1893 to 1.5210) were significantly associated with higher total hospital cost per discharge. Gender was not significantly associated with cost (β=0.0727; 95% CI: –0.0679 to 0.2134). Conclusions: To our knowledge this is the first study to examine predictors of ICH treatment costs in Canada. In a Canadian health centre, ICH treatment costs were significantly associated with patient age, level of comorbidity, in-hospital mortality, and surgical treatment. This study provides evidence that it may be reasonable to consider …

Docosahexaenoic Acid Signaling Modulates Cell Survival in Experimental Ischemic Stroke Penumbra and Initiates Long-Term Repair in Young and Aged Rats

BackgroundDocosahexaenoic acid, a major omega-3 essential fatty acid family member, improves behavioral deficit and reduces infarct volume and edema after experimental focal cerebral ischemia. We hypothesize that DHA elicits neuroprotection by inducing AKT/p70S6K phosphorylation, which in turn leads to cell survival and protects against ischemic stroke in young and aged rats.Methods and ResultsRats underwent 2 h of middle cerebral artery occlusion (MCAo). DHA, neuroprotectin D1 (NPD1) or vehicle (saline) was administered 3 h after onset of stroke. Neurological function was evaluated on days 1, 2, 3, and 7. DHA treatment improved functional recovery and reduced cortical, subcortical and total infarct volumes 7 days after stroke. DHA also reduced microglia infiltration and increased the number of astrocytes and neurons when compared to vehicle on days 1 and 7. Increases in p473 AKT and p308 AKT phosphorylation/activation were observed in animals treated with DHA 4 h after MCAo. Activation of other members of the AKT signaling pathway were also observed in DHA treated animals including increases in pS6 at 4 h and pGSK at 24 h. DHA or NPD1 remarkably reduced total and cortical infarct in aged rats. Moreover, we show that in young and aged rats DHA treatment after MCAo potentiates NPD1 biosynthesis. The phosphorylation of p308 AKT or pGSK was not different between groups in aged rats. However, pS6 expression was increased with DHA or NPD1 treatment when compared to vehicle.ConclusionsWe suggest that DHA induces cell survival, modulates the neuroinflammatory response and triggers long term restoration of synaptic circuits. Both DHA and NPD1 elicited remarkable protection in aged animals. Accordingly, activation of DHA signaling might provide benefits in the management of ischemic stroke both acutely as well as long term to limit ensuing disabilities.

Angiotensin II Type 2 Receptor Stimulation Initiated After Stroke Causes Neuroprotection in Conscious Rats

We have demonstrated previously that pretreatment with an angiotensin II type 2 receptor (AT(2)R) agonist is neuroprotective against a subsequent stroke independent of any changes in blood pressure. Therefore, in the current study, we have examined the potential neuroprotective effect of AT(2)R stimulation initiated after stroke induction to mimic the clinical setting. Intracerebroventricular administration of the AT(2)R agonist CGP42112 was commenced 6 hours after an ischemic stroke had been induced in conscious spontaneously hypertensive rats. CGP42112 given over 4 doses in the same rats (3 µg/kg per dose centrally) at 6, 24, 48, and 72 hours after stroke induction reduced total infarct volume (32 ± 13 mm(3) versus vehicle, 170 ± 49 mm(3); P<0.05) and improved motor function. Furthermore, we have demonstrated that AT(2)R stimulation after stroke increased neuronal survival, decreased apoptosis, and caused an increase in the number of activated microglia in the core region of damage. The effects of CGP42112 were partially reversed with the coadministration of an AT(2)R antagonist, PD123319. Thus, the current study has shown for the first time that delayed central AT(2)R stimulation after a cerebral incident is neuroprotective in a conscious rat model of stroke.

Docosahexaenoic acid complexed to human albumin in experimental stroke: neuroprotective efficacy with a wide therapeutic window

BackgroundDocosahexaenoic acid (DHA) complexed to human serum albumin (Alb) is neuroprotective after experimental stroke. Here we tested using lower concentrations of albumin as part of the complex to achieve neuroprotection. We found that lower Alb concentrations extend the therapeutic window of protection beyond 5 h after stroke onset.MethodsSprague–Dawley rats were received 2 h middle cerebral artery occlusion (MCAo). The behavior was evaluated on day 1, 2, 3 and 7 after MCAo. In the dose–response study, animals were given either DHA (5mg/kg), Alb (0.63g/kg), DHA-Alb (5mg/kg + 0.32, 0.63 or 1.25 g/kg) or saline, i.v. 3 h after onset of stroke (n=6-8 per group). In the therapeutic window study, DHA-Alb (5mg/kg + 1.25g/kg) was administered i.v. at either 3, 4, 5, 6 or 7 h after onset of stroke (n=7-9 per group). Alb (1.25g/kg) was given at 3 h or 5 h and saline at 3h after onset of reperfusion. Seven days after MCAo, infarct volumes and number of GFAP, ED-1, NeuN, SMI-71 positive cells and vessels were counted.ResultsModerate DHA-Alb doses (0.63 and 1.25 g/kg) improved neurological scores compared to albumin-treated rats on days 1, 2, 3 and 7. All DHA-Alb doses (0.32, 0.63 and 1.25 g/kg) markedly reduced cortical (by 65-70%), striatal (by 52-63%) and total infarct volumes (by 60-64%) compared to native Alb group. In the therapeutic window study DHA-Alb led to improved neurological score and significant reductions of infarct volumes (especially in the cortical or penumbral region), even when treatment was initiated as late as 7 hours after onset of MCAo.ConclusionsThe DHA-Alb complex affords high-grade neurobehavioral neuroprotection in focal cerebral ischemia, equaling or exceeding that afforded by native Alb or DHA, at considerably moderate doses. It has a broad therapeutic window extending to 7 h after stroke onset. Taken together, these finding support the potential clinical feasibility of administering DHA-Alb therapy to patients with acute ischemic stroke.