XVII. European Stroke Conference
Nice, France
XVII. European Stroke Conference
Nice, France
Poster Session: Experimental studies
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
01
Functional evaluation in experimental embolic stroke: Comparison of a compound neurological assessment with conventional scoring systems
M. Nedelmann
P. Reuter
B. Alessandri
O. Kempski
T. Gerriets
Justus Liebig-University, Giessen
GERMANY
Background: The embolic MCA occlusion model in rats is used for recanalisation studies in acute stroke. Next to morphological lesion size, the assessment of functional outcome may improve the value of this animal model. In this study we compare a novel compound assessment scale with 2 widely used scoring systems based on motor disability. Methods: Male Wistar rats were submitted to MCA clot embolism (n=14) or sham surgery (n=7). Follow-up period was 6 days. Blinded outcome assessment consisted of testing for ten different motor and non-motor (i.e., sensory and coordinatory) deficits (NeuroScore) that are not evaluated by more conventional outcome scores. The results were compared with histology and with two scoring systems on motor functions that are widely used in routine experimental testing (5-point scale described by Zea Longa et al 1989; 6-point scale by Zausinger et al 2000). Results: Neurological impairment as scored with the NeuroScore gave significant results throughout the observation period (p<0.001). The degree of impairment significantly correlated with infarct volume (p<0.001; R=0.80). Both the 5-point and the 6-point scale significantly detected impairment. However, the 5-point scale did not correlate with histology (p=0.28; R=0.30). The 6-point scale revealed significant correlation, however, compared to the NeuroScore, the strength of correlation was consistently lower (p<0.05; R=0.51). Discussion: In this study, a novel outcome test is presented that provides quantitative and objective tools to test functional impairment in rats following embolic stroke. Compared to established tests, it shows superiority with regard to discrimination between various degrees of lesion volume.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
02
Inhibition of p53-dependent Neuronal Apoptosis after Focal Cerebral Ischemia
K.Y.Park
G.M.Kim
C.S.Chung
Y.C.Youn
O.S.Kwon
J.H.Min
Chung-Ang University Hospital
SOUTH KOREA
Background: We sought to evaluate the mechanism of p53-mediated activation of proapoptotic cascades and the effect of p53 suppression on downstream gene activation and subsequent neuronal cell apoptosis in focal cerebral ischemia. Methods: Middle cerebral artery occlusion (MCAO) was induced by the intraluminal sutre model for 90 minutes in male Sprague-Dawley rats. p53 DNA binding activity and the subsequent expression of Puma, Noxa, and caspase-3 were evaluated at different time points. Antisense oligodeoxynucleotide (ODN) to p53 was infused into the right cerebral ventricle. We studied the regional and cellular localization of ODN in the rat brain. The effects of antisense ODN to p53 on subsequent Puma, Noxa, and Caspase-3 expression were examined at different time points after MCAO. The brains were processed for immunohistochemical staining with TUNEL and cresyl violet and assessed for the number of apoptotic neurons and the volume of infarction. Results: After transient MCAO, p53 DNA binding activity was increased. The expression of Puma, Noxa, and caspase-3 was also increased and the mitochondrial translocation of Puma and Noxa was observed. Insulted neurons expressed Puma and Noxa on immunohistochemical staining in the ischemic cortical region. The expression of Puma or Noxa was overlapped in the caspase-3 or TUNEL-positive cells. The immunoreactivity of FITC-labeled ODN was enhanced in neurons in the brain after injection of ODN. Infusion of antisense ODN to p53 inhibited Puma and Noxa expression after transient MCAO. The number of TUNEL-positive cells and the amount of active caspase-3 expression were significantly reduced in antisense ODN injection group (p<0.05). The size of infarct volume, measured by cresyl violet staining at 24 hours after transient MCAO, was also significantly decreased in antisense ODN injection group (p<0.05). Conclusions: Our results suggest that the ischemic activation of p53 regulates the subsequent expression of the BH3-only proapoptotic cascades and antisense ODN to p53 inhibits selected downstream proapoptotic events.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
03
Delayed Erythropoietin Treatment in Animal Model of Ischemic Stroke
H.Y.Kim
S.H.Koh
Y.B.Lee
K.Y.Lee
Y.J.Lee
J. Kim
H.T.Kim
W.Y.Jang
H.D.Park
S.H.Kim
College of Medicine, Hanyang University
Gachon University of Medicine and Science, Gil Hospital
SOUTH KOREA
Background Ischemic brain injury results from a complex pathophysiological sequence. It has been reported that Erythropoietin (EPO) treatment in early ischemic phase can reduce infarct volume and improve behavioral function via anti-inflammatory and antiapoptotic effects. Moreover, EPO stimulates the proliferation and differentiation of endogenous neuronal progenitor cells. Therefore, we investigated to ascertain whether these beneficial effects of EPO can be induced by delayed repeated injection. Methods The left middle cerebral artery (MCA) of Sprague-Dawley (SD) male rats was occluded for 2 hours. Eighty SD rats were divided into 4 groups; group 1(S-S) saline injection weekly since acute stage, group 2(E-S) EPO injection once in acute stage, group 3(S-E) EPO injection weekly from 7 days since cerebral infarction, group 4(E-E) EPO injection weekly since acute stage. Behavioral tests were performed sequentially. Brain caspase-3, pAkt, COX-2, HSTF-1, and GFAP activities were determined by Western blots and immunohistochemistries at 24 hours. Additionally, brain nestin activity was estimated at 6 weeks. Results The mean infarct volume was decreased in EPO injection group at 24 hours. Western blots and immunohistochemistries at 24 hours showed increased pAkt, decreased caspase-3, COX-2, and GFAP activities in EPO treatment group at 24 hours. Enhanced nestin activity was detected in E-E, E-S, and S-E groups at 6 weeks. However, improved behavior functions were noted only in E-E and E-S groups. Discussion Despite of neurogenic property shown in our results, its effect could not influence on behavioral improvement. Therefore, we cautiously postulate that early treatment of EPO to reduce the inflammation and apoptosis would be more important in restoring neurobehavioral function.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
04
Experimental cell therapy of stroke - preclinical evaluation of transplantation modalities
D.C.Wagner
U.R.Schmidt
A. Foerschler
M. Kamprad
A. Kranz
D. Egger
F. Emmrich
J. Boltze
Fraunhofer Institute for Cell Therapy and Immunology
GERMANY
Objectives The relevance of cell therapies in neurological affections has been shown in several preclinical studies. These auspicious results should be confirmed in clinical trials, but there are a couple of open questions. In our experiment we transplanted human umbilical cord blood cells (HUCBC) within different time points to evaluate the appropriate therapeutic time window. Furthermore, we compared the efficacy of intravenous and intracerebral transplantation. Materials and Methods Permanent middle cerebral artery occlusion (MCAO) was conducted in 61 male spontaneously hypertensive rats. A cell suspension of 10E6 HUCBC was injected intravenously at different time points upon MCAO: 4h (n=8), 24h (n=8), 72h (n=7), 120h (n=8) and 14d (n=8). The control group (n=8) received vehicle solution 24h following MCAO. Intrastriatal transplantation of 3x10E5 HUCBC (n=8) took place 24h upon MCAO, the corresponding control group (n=6) received vehicle solution intrastriatally. Functional outcome was examined by three behavioural tests (RotaRod, BeamWalk and modified Neurological Severity Score) from Day 1 to 30 regularly. Infarct volumetry was performed via magnetic resonance investigations on Days 1, 8 and 29. Immunohistochemical methods were used to investigate localization and differentiation of transplanted cells and to analyze glial scarring. Results We observed a significant reduction of sensorimotor defects when cell transplantations took place within the first 72h following experimental brain ischemia. These results were supported by a degradation of glial scarring and a decreased tissue lost. Transplantation 120h upon stroke induced a significant improvement of sensorimotor deficits without a measurable effect on glial activation and tissue lost. Transplantation after a two week period failed to show any beneficial effects. Local transplantation of HUCBC 24h upon stroke showed a significant improvement within all examination routines. We found no evidence for significant differences between local and systemic administration of HUCBC 24h following stroke.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
05
U - shaped response to blood pressure lowering in spontaneously hypertensive rats treated with candesartan after experimental acute stroke.
W. Kozak
A. Kozak
M.H.Johnson
H.F.Elewa
S.C.Fagan
The University of Georgia
College of Pharmacy
USA
Background: Hypertension is common both as a risk factor for, and a reaction to, ischemic stroke. We have shown that candesartan at a dose of 1 mg/kg injected into normotensive rats at reperfusion decreased the acute stroke-induced elevation of mean arterial blood pressure (MAP) and improved neurologic outcomes. The aim of the present study was to determine if the same beneficial response could be achieved in spontaneously hypertensive rats (SHR). Methods: SHRs were subjected to middle cerebral artery occlusion (MCAO) for 3 hours followed by reperfusion. Intravenous injections of three doses of candesartan (0.1 mg/kg, 0.3 mg/kg and 1.0 mg/kg), and saline as control, were administered 5 minutes after artery reopening in separate groups of animals. Blood pressure was monitored by means of telemetry. Neurologic function, infarct size, edema formation and hemoglobin content in the ischemic hemisphere were evaluated 24 h after MCAO. Response to blood pressure lowering in the acute stroke period was compared between Wistars and SHRs. Results: MAP of SHR increased immediately upon MCAO from 135 (baseline) to 190 mmHg, and remained elevated until reperfusion. Candesartan reduced BP in a dose-dependent manner (p<0.0001), with a drop below baseline after a dose of 1.0 mg/kg. Infarct size (p=0.0004), edema formation (p=0.023) and hemoglobin content (p=0.0092) were significantly lower than saline in the candesartan 0.3 mg/kg dose group only. In addition, functional outcome was significantly improved in the candesartan 0.3 mg/kg group (p<0.01). SHR animals were more sensitive to the BP lowering effects of candesartan than normotensive controls and tended to experience dramatic and prolonged BP lowering with the highest dose. Conclusion: We conclude that intermediate doses of candesartan provide neurovascular protection after temporary MCAO in spontaneously hypertensive rats. Preexisting hypertension increases the likelihood of hypotension after BP lowering in the acute stroke period.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
06
Inducible nitric oxide synthase expression is responsible for vasogenic edema after experimental intracerebral hemorrhage
S.W.Jeong
D.E.Kim
J.Y.Kim
D.W.Kim
Dongguk University International Hospital, Dongguk University College of Medicine
SOUTH KOREA
BACKGROUND AND PURPOSE: Brain edema after intracerebral hemorrhage (ICH) is an important prognostic factor. Inducible nitric oxide synthase (iNOS) is not normally expressed in brain, but induced after ischemia, and known to be involved in secondary neuronal injury. However, its significance and temporal expression profile in intracerebral hemorrhage were unknown. We investigated temporal profile of iNOS expression, and tested whether iNOS influenced vasogenic edema after ICH. METHODS: Experimental ICH was induced by intrastriatal stereotaxic administration of bacterial collagenase. In first experiment, C57BL/6 mice were sacrificed at day 0(n=3), 1(n=3), 3(n=3), 7(n=3) after ICH. We investigated the induction of iNOS after ICH on protein and mRNA level by immunohistochemistry, western blot assay and RT-PCR. Second, C57BL/6 mice (n = 8) and iNOS knockout mice (n = 10) were used. Evans blue in PBS was infused into mice via tail vein as a blood brain barrier (BBB) permeability tracer at 48hr after ICH. Brain tissue was obtained at 72 hrs after ICH. BBB permeability by Evans Blue dye was measured. RESULTS: The result of RT-PCR showed the mRNA, which was not expressed in normal rats, were induced in collagenase-induced ICH rats. The induction of iNOS mRNA was maximal at 1day after ICH, then started to decrease at 3 days and nearly normalized at 7 days after ICH. The maximal expression of iNOS protein after ICH was observed at 1 day. Immunoreactivity was detected in cells around the lesion of hemorrhage. Brain Evans blue leakage concentration of whole brain was 217.5±6.84 ng in C57BL/6 control and 204.5±3.35ng in iNOS knockout mice (p=0.03). CONCLUSIONS: We have demonstrated that the induction of iNOS mRNA was maximal at 1day after ICH, and brain vasogenic edema was reduced in iNOS knockout mice compared with littermate. These findings suggest that iNOS modulation could be an anti-edematic therapy for ICH.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
07
Expression of the transcription factor olig1 and the demyelination in the white matter after focal cerebral ischemia in the rat
Y. Yong-bo Zhang
X. Xiao-Yu Gao
D. De-Xin Wang
H. Hong Zhao
J. Ji-Mei Li
Beijing Friendship Hospital, Capital Medical University
CHINA
Objective The transcription factor olig1 is related with the differentiation and maturation of oligodendrocytes, and has an important role in the formation and repair of myelin sheath in multiple sclerosis. This study is to explore the olig1 expression after focal cerebral ischemia in rats, and to identify the relationship between the expression of olig1 and the white matter damage after ischemia. Methods Sprague-Dawley male rats were subjected to 2 hours of middle cerebral artery occlusion by an intraluminal thread and divided randomly into different groups. The expression of olig1 was performed by immunohistochemistry and the relative quantification PCR. Myelin sheath was characterized by Luxol fast blue-periodic acid Schiff Staining (LFB-PAS) and the integrated optical densities (IODs) ratio in the external capsule of both hemispheres was calculated to reflect the severity of white matter damage. Results Olig1 was mainly present in the cytoplasm of oligodendrocytes not only in normal brain but also at the edge of infarction. The olig1 expression level decreased from 6h after ischemia, and gradually decreased over time to the lowest at 7d(P<0.001). From 14d, it began to increase and was higher than that in sham-operation group at 21d (P<0.001). Myelin sheath lost their LFB-PAS stabilization and IODs ratio decreased also at 6h after ischemia, and also gradually decreased over time to the lowest at 14d(P<0.001).There were no significant difference in IODs ratio between 14d group and 21d group(P>0.05). Conclusions The expression of olig1 has temporal changes after cerebral ischemia. The decrease of the olig1 expression was coincidence with the demyelination in the white matter after focal cerebral ischemia in the rat. So maybe olig1 participates in the pathogenesis of cerebral white matter lesion and plasticity after ischemia.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
08
Targeted Brain Delivery of Z-DEVD-FMK Containing Chitosan-PEG-BIO-SA/OX26 Nanoparticles Reduces Infarct Volume by Caspase-3 Inhibition in Experimental Ischemia
H. Karatas
Y. Aktas
O. Pinarbasili
Y. Ozdemir
M. Yemisci
Y. Capan
T. Dalkara
Institute of Neurological Sciences and Psychiatry, Hacettepe University
TURKEY
The inhibition of the caspase-3 enzyme is reported to increase neuronal cell survival following cerebral ischemia. The peptide Z-DEVD-FMK is a specific caspase inhibitor, which significantly reduces vulnerability to the neuronal cell death. However this molecule is unable to cross the blood brain barrier (BBB). Thus the development of an effective delivery system is needed to provide sufficient drug concentration into the brain to prevent cell death. Using the avidin (SA)-biotin (BIO) technology, we describe here the design of chitosan (CS) nanospheres conjugated with polyethyleneglycol (PEG) bearing OX26 whose affinity for the transferrin receptor may trigger receptor-mediated transport across the BBB. Our previous study revealed that these novel nanoparticles (np) appear to be promising carriers for the transport of the anticaspase peptide Z-DEVD-FMK into the brain. The purpose of this study was to prepare Z-DEVD-FMK loaded CS-PEG-BIO/SA-OX26 np and to evaluate in vivo efficiency of this nano-carrier system in decreasing infarct volume. CS-PEG-BIO np were prepared according to the procedure previously described by our group. Various concentrations of Z-DEVD-FMK were incorporated into CS-PEG-BIO np. Fluorescence labelled CS-PEG-BIO/SA-OX26 np (1mg in 200µL saline) were injected intravenously to swiss albino mice before ischemia/reperfusion. Intraluminal filament model was used to induce ischemia and filament was pulled back to get reperfusion. After 2 h of ischemia and 24 h of reperfusion the infarct volume was calculated. Z-DEVD-FMK was successfully associated into the np and the incorporated amount of the peptide was increased from 200ng/mL to 800ng/mL. Nanoparticle treatment significantly decreased infarct volume when compared to sham by the loaded amount of the peptide. These data demonstrate that the Z-DEVD-FMK loaded CS-PEG-BIO-OX26/SA np sufficiently penetrate through BBB to efficiently inhibit the caspase-3 activity after cerebral ischemia and provide neuroprotection.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
09
Imaging Cellular Responses to Experimental Focal Cerebral Ischemia in Rats by Magnetic Resonance Imaging
Y.M.YANG
X.Y.FENG
Z.W.YAO
Hua Shan Hospital of FuDan University
CHINA
Background: Cerebral ischemia educes a strong cellular response that involves glia cell activation as well as infiltration of hematogenous leukocytes within and around the primary infarct. The present study is conducted to demonstrate the feasibility of using the multi-mode MR techniques, including diffusion tensor imaging to provide useful information that might aid understanding of cellular responses to experimental cerebral ischemia in rats. Materials and Methods: Rats (n=6) were subjected to transient middle cerebral artery occlusion to assess the changes of MR T2, apparent diffusion coefficient (ADC) and fractional anisotropy (FA) in ischemic core and boundary zone at 1 day and 2 weeks. The correspondent histological features at weeks 2 were showed by cresly-violet and luxol fast blue stain. Results: At day 1 after ischemia, T2 value in the infarction elevated, ADC and FA declined. The signal intensity in all MR images was homogeneous in the ischemic core and boundary zones. At weeks 2, on the ischemic core, T2 value elevated obviously, the ADC reversed above normal values, and FA declined much more. Compared with ischemic core, the signal of the boundary zones presented that T2 and ADC signal reduced to the corresponding intensity of the contralateral region, and the previously declined FA recovered. The histological sections demonstrated the liquefaction necrosis was the predominant feature in the ischemic core, and the cellular infiltration was evident in the ischemic boundary zone at weeks 2 after ischemia. Regions with dense cellular infiltration in histological sections showed an excellent spatial correlation with the iso-intensity areas on T2, ADC and FA maps in all animals. Conclusions: Our data indicated that FA, as one of new MR parameters, accompanying T2 and ADC appeared as useful measurements to identify the cellular infiltration from liquefaction in the ischemic core in chronic infarction. The imaging presentation should contribute to the development of neo-adjuvant therapies targeting inflammatory responses to focal ischemia.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
10
Effects of intracerebroventricular infusion of angiotensin-(1–7) on bradykinin formation and the kinin receptor expression after focal cerebral ischemia-reperfusion in rats
Y.D.Zhang
J. LU
nanjing brain hospital, nanjing medical university
CHINA
BACKGROUND AND PURPOSE: Accumulating evidence suggests the angiotensin-(1–7) [Ang-(1-7)] is an active member of the central nervous system (CNS) renin-angiotensin system. We evaluated the possibility that intracerebroventricular exogenous Ang-(1-7) infusion could participate in the potentiation of bradykinin (BK) release and the kinin receptor expression in ischemic brain parenchyma after focal cerebral ischemia-reperfusion in rats. METHODS AND RESULTS: The middle cerebral artery occlusion (MCAO) and sham-operation models were prepared, continuous administrated with Ang-(1-7) (1pmol/0.5µl/h, 100pmol/0.5µl/h, 10nmol/0.5µl/h) or artificial cerebrospinal fluid (aCSF) (0.5µl/h) by implanted Alzet osmotic minipump into lateral cerebral ventricle after reperfusion in male Sprague-Dawley (SD) rats, and thus were divided into sham operation group (sham operation + aCSF), aCSF treatment group (MCAO+aCSF) and Ang-(1-7) treatment groups [MCAO + Ang-(1-7)] at three doses with low (1pmol/0.5µl/h), medium (100pmol/0.5µl/h) and high (10nmol/0.5µl/h) doses. The five groups were further divided into 6 subgroups for observation at 3, 6, 12, 24, 48 and 72 hours after reperfusion following a 90-min MCAO. The levels of bradykinin in ischemic tissues were measured by ELISA . RT-PCR and western blot were used to determine mRNA and protein level of the kinin receptors in ischemic tissues. Cerebral infarction resulted in a significant increase of bradykinin formation at 3 and 6 hours compared with sham operation group after reperfusion, whereas medium and high-dose Ang-(1-7) infusion markedly enhanced bradykinin levels at 3, 6, 12, 24 and 48 hours after reperfusion. Medium and high-dose Ang-(1-7) infusion treatment markedly increased kinin B2 receptor mRNA and protein expression, whereas only high-dose Ang-(1-7) infusion induced upregulating the expression of B1 receptor. Low-dose Ang-(1-7) infusion did not modify both the kinin B1 and B2 receptors expression compared with aCSF treatment group in ischemic brain parenchyma after focal cerebral ischemia-reperfusion.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
11
Meta-analysis and Meta-regression analysis of the efficacy of Granulocyte-colony stimulating factor in animal models of focal cerebral ischemia
J. Minnerup
J. Heidrich
J. Wellmann
A. Rogalewski
A. Schneider
W. R.Schäbitz
University of Münster
GERMANY
Background and Purpose: Recent reports describe the efficacy of the hematopoietic growth factor granulocyte-colony-stimulating factor (G-CSF) in animal stroke models. Early clinical multicenter trials evaluating the effect of G-CSF in acute stroke and pilot clinical trials for the subacute phase are ongoing. To guide further development, a meta-analysis was performed assessing the effects of G-CSF on infarct size and sensorimotor deficits. Methods: Using electronic and manual searches of the literature, we identified studies describing the efficacy of G-CSF in animal models of focal cerebral ischemia. Two reviewers independently selected studies and extracted data on study quality, G-CSF doses, time of administration, and outcome measured as infarct volume and/or sensorimotor deficit. Data from all studies were pooled using meta-regression analyses. Results: Thirteen studies including 277 animals for infarct size calculation and 258 animals for assessment of sensorimotor deficit met the criteria for inclusion. Overall efficacy of G-CSF regarding infarct size reduction was 42%. Meta regression analysis revealed a 0.8% (p<0.0001) decrease in infarct size per one µg/kg increase in G-CSF-dose. Infarct volume decreased by 3.0% (p<0.0001) with every hour of delayed treatment initiation within the first 24 hours. Sensorimotor deficits categorized into 3 subgroups were improved between 24% and 40%. Conclusion: Our findings consolidate G-CSF as a drug that both reduces infarct size and enhances functional recovery. These effects are dose-dependent. In contrast to most other neuroprotectants a beneficial outcome may also be achieved when treatment is delayed
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
12
Oxygen Therapy does not Increase Free Oxygen Radical-induced Brain Damage in Experimental Cerebral Ischemia
R. Veltkamp
G. Wolferts
L. Sun
University Heidelberg
GERMANY
Background: Experimental studies suggest a therapeutic potential of normobaric (NBO) and hyperbaric oxygen (HBO) in ischemic stroke. A major concern, however, is that oxygen therapy enhances production of reactive oxygen species (ROS) and thereby induces cell death. We examined the effect of oxygen therapy on ROS production and in a ROS susceptible mouse strain. Methods: Mice underwent filament-induced middle cerebral artery (MCAO) for 90 min. Thirty min after filament introduction, animals were placed in a HBO chamber and breathed either air, 100% O2 at ambient pressure (NBO), or at 3.0 bar (HBO) for 1 h. Superoxide (SO-) production was quantified 2 h after reperfusion using hydroethidine fluorescence. Effect of cotreatment with the ROS scavenger PBN on infarct size was evaluated 24 h after MCAO. Efficacy of HBO was also studied in heterozygous SOD-2 (+/-) mice. Results: Interhemispheric ratio (ischemic/nonischemic) of SO- formation detected by hydroethidine fluorescence was significantly lower in HBO (0.97 +/- 0.13) than in NBO (1.13 +/- 0.08) and air (1.34 +/- 0.10) treated mice (p<0.05; ANOVA). HBO (33 +/- 9 mm3) had significantly smaller infarct volume than air treated wild type mice (51 +/- 18), but cotreatment of HBO+PBN failed to reduce infarct size further (39 +/- 14). As previously described, SOD-2 (+/-) mice had significantly larger infarcts than wild type control. Nevertheless, HBO significantly reduced infarct volume in this ROS susceptible strain compared to air. Conclusions: Intraischemic treatment with HBO reduces rather than enhances SO- production in the ischemic hemisphere. As HBO was protective even in mice which are susceptible to ROS, it does not appear to increase ROS-induced brain injury. Supported by Deutsche Forschungsgemeinschaft (VE 196/2-2)
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
13
Early stage investigations of ultrasmall superparamagnetic iron oxide particles (USPIO)-induced signal changes after permanent middle cerebral artery occlusion in mice.
V. Desestret
J.C.Brisset
E. Devillard
S. Moucharrafie
S. Nataf
J. Honnorat
N. Nighoghossian
Y. Berthezene
M. Wiart
Laboratoire Creatis-LRMN, CNRS, UMR 5220
FRANCE
Background: Interest of ultrasmall superparamagnetic iron oxide particles (USPIO) for monitoring neuroinflammation with magnetic resonance imaging (MRI) has been demonstrated. Nevertheless, the interpretation of MR signal changes after intravenous USPIO injection at early stages of focal cerebral ischemia remains controversial. Methods: Here, we confronted MRI enhancement at 7T and histological iron particles distribution from 6 to 24 hours after permanent middle cerebral artery occlusion (pMCAO) induced by electrocoagulation in mice. USPIO (Ferumoxtran-10, Guerbet, France) were injected 5h post-injury. Mice were scanned before injection and then either 1h and 19h after. We isolated mononuclear cells of peripheral blood for cytospin iron staining. After the last MR scan, mice were sacrificed for histology to depict: iron with Prussian blue staining and phagocytes by immunochemistry against F4/80. For in vitro evaluation of cell labelling, free-floating monocytic cells were derived from bone-marrow cultures and incubated for 24h with USPIO. Results: We observed four areas of early signal changes after USPIO injection on gradient echo T1 weighted imaging. Gadolinium enhancement indicated early blood brain barrier (BBB) disruption confirmed by histological immunoglobulin staining. On histology, iron staining was mostly associated to the vascular and the cerebrospinal fluid (CSF) compartments. Only brain phagocytes in contact with CSF have already phagocytosed iron particles. USPIO-loaded cells were not detected in the blood of injured mice within the first 24h. Cultured phagocytes incubated with USPIO did not show significant iron labelling. Discussion: Our study suggests that early brain USPIO enhancements are mainly caused by unspecific mechanisms such as BBB leakage and intravascular trapping.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
14
Increased Cerebral Metabolism in Excersise-induced Neuroprotection after Stroke
M. Guo
J.L.Caron
D.F.Jimenez
Y. Ding
University of Texas Health Sciences Center at San Antonio
USA
It has been reported that physical exercise increases the metabolism rate in muscles. The purpose of this study was to test whether pre-ischemic exercise alter cerebral metabolism in association with reduced brain injury after stroke. ADP/ATP ratio and key molecules in glycolysis, including glucose transporter 1(GLUT1), glucose transporter 3 (GLUT3), phosphofructokinase (PFK), and lactate dehydrogenase (LDH) were investigated. Adult male Sprague Dawley rats were subjected to a 30 minute exercise on a treadmill each day for 3 weeks. Stroke was induced by a 2-hour middle cerebral artery (MCA) intraluminal filament occlusion in exercised rats and controls. Brain infarct volume was determined by Nissl staining. ADP/ATP ratio, the gene and protein expressions of GLUT1, GLUT3, PFK, and LDH were determined in the MCA territory using bioluminescent assay, real time PCR and Western Blot. In association with significantly (p<0.05) reduced brain infarct volume and neurological deficits after stroke, pre-ischemic exercise initially increased ADP/ATP ratio, as well as the levels of mRNA and protein of GLUT1, GLUT3, PFK, and LDH. Interestingly, ADP/ATP ratio in stroke remained significantly low level in the exercised rats as compared to that in non-exercised rats. The increase in GLUT1, GLUT3, PFK, and LDH after physical exercise was further significantly (p<0.05) enhanced during ischemia and reperfusion at 6, 12, and 24 hours. Our results indicated that physical exercise increases the metabolism rate in brain, and that the metabolic function in brain after stroke can be enhanced by the pre-ischemic exercise, leading to reduction in brain injury.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
15
IDENTIFICATION AND CHARACTERIZATION OF NEUROSERPIN IN HUMAN SERUM AS A DIAGNOSTIC MARKER FOR ACUTE ISCHEMIC STROKE.
R. Rodríguez-González
T. Sobrino
I. Cristobo
O. Hurtado
P. Ramos-Cabrer
O. Moldes
J. Agulla
D. Brea
I. Lizasoain
J. Castillo
Clinical Neuroscience Research Lab. Hospital Clínico Universitario. Santiago de Compostela
SPAIN
Background: Neuroserpin, a natural t-PA inhibitor, has shown neuroprotective effects in experimental stroke models. Nevertheless, its clinical application for acute stroke has not yet been tested. Our purpose was to identify neuroserpin in human serum samples in order to analyze its utility as a diagnostic marker in acute stroke. Methods: Thirty subjects matched for age and sex were included: fifteen with ischemic stroke, five with hemorrhagic stroke, five with Parkinson´s disease and another five healthy control subjects. Of the ischemic stroke patients, five received t-PA treatment, five had low infarct volume (2-10 cc) and another five had high infarct volume (35-120 cc). For all stroke patients, blood samples were collected at admission and at 24 hours from stroke onset. Serum neuroserpin levels were analyzed by Western-blot. Results: Neuroserpin was identified in the serum of all subjects. Differences were found between ischemic stroke patients and control subjects (p<0.05). Among ischemic stroke patients, differences were also found between high and low infarct volume groups (p<0.05). Both control subjects and low infarct volumes showed higher neuroserpin levels. Although not statistically significant, non t-PA treated, hemorrhagic stroke and Parkinson’s disease patients showed higher neuroserpin levels than ischemic stroke group. No differences were found between samples at admission and at 24 hours from stroke onset. Conclusion: Neuroserpin may represent a diagnostic biomarker for ischemic stroke, specially useful in patients treated with t-PA. However, larger studies with more sensitive molecular techniques like ELISA are necessary to confirm these preliminary results.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
16
Invasive versus non-invasive monitoring of brain tissue oxygenation
S. Griffith
J.L.Caron
R. Coleman
M. Stiegelman
J. Salazar
R. DiGeronimo
Dept. of Neurosurgery, University of Texas Health Science Center at San Antonio and Dept of Pediatrics and Surgery, Wilford Hall USAF Medical Center
USA
Accurate measurement of brain tissue tension of oxygen (PbtO2) is important in neurosurgical critical care prevention and treatment of anoxic brain injury. Invasive monitoring of PbtO2 with the Licox® intracranial probe is used in neurosurgery ICUs. Near-infrared spectroscopy can also measure regional brain saturation of oxygen (rSO2) with Somanetics®. The purpose of this study was to examine for congruency the measurement of cerebral oxygenation by comparing the rSO2 as measured by Somanetics® to the PbtO2 as measured by Licox® during periods of hypothermia and circulation arrest. Methods: Immature Yorkshire swine were assigned to one of two study groups (n=6/group): (I) Control at 18°C (no cerebral perfusion) or (II) cerebral perfusion at 18oC. Animals underwent 60 minutes of hypothermia, followed by 4 hours of recovery. Cerebral perfusion was stopped by placing the animal on cardiopulmonary bypass. Cerebral oxygenation was measured using regional near-infrared spectroscopy (rSO2 index; Somanetics®) and direct brain tissue oxygen tension by surgically implanted cerebral catheters (pBtO2 (mmHg); Licox®). Results: There was good correlation between rSO2 and pBtO2 measurements at baseline. During hypothermia with absence of cerebral perfusion, there was a drop in the rSO2 index from 63.6 ± 6.3 to 28.7 ± 9.2. The PbtO2 correlation was 0.01 ± .005 relative units. With continued cerebral perfusion rSO2 index changed from 67.4 ± 7.5 to 71.4 ± 9.4. The PbtO2 correlation was 1.29 ± .30 relative units. Upon return to baseline conditions, PbtO2 analysis revealed persistently lower values in the control group until 2 hrs post-recovery despite normalization of the rSO2 (0.53 ± 0.02 RU for 1 hour post recovery and 0.79 ± 0.23 for 2 hours post recovery). Conclusions: Both Somanetics® (rSO2) and Licox® (PbtO2) show consistent measurement of cerebral oxygenation at baseline conditions. Invasive Licox® monitoring indicates a persistently lower level of cerebral oxygenation than does the non-invasive Somanetics® monitor during hypoperfusion and initial return to baseline levels of perfusion.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
17
The MR image and ultrastructure observation of the secondary damage in substantia nigra following single side middle cerebral artery occlusion
Y.M.YANG
X.Y.FENG
Z.W.YAO
Hua Shan Hospital of FuDan University
CHINA
Objective:In this present study, we will conduct to observe the changes in substantia nigra following focal cerebral ischemia in the middle cerebral artery territory by magnetic resonance imaging and electron microscopy methods. Materials and Methods: Healthy male Sprague-Dawley rats (n=28) were divided into three groups randomly: MCAO group (n=12), MCAO/R(reperfusion) group (n=12) and control group (n=4). The perfusion CT scan, 3D Time-Of-Flight MRA and MR T2WI, DWI and T2 map sequences were performed. The image analysis was made between the primary ischemic region and substantia nigra, and the MR imaging was interpreted by the ultrastructure observation. Results: The perfusion CT CBF image after MCAO 20min showed the decrease of the blood flow perfusion in MCA territory, but in substantia nigra, blood flow perfusion between two sides had no significant difference. By MRA, the occlusion and recirculation of the single side middle cerebral artery was validated, and the signal intensity of bilateral posterior cerebral arteries as the supply artery of substantia nigra was equal each other. After cerebral ischemia, the primary lesions in cortex and striatum were imaged by MRI from 6h to 14d. In substantia nigra, when the primary lesion was so obviously seen on 6h and 1d, no changes on MR imaging were detected. But on 3d, the ipsilateral substantia nigra showed obviously higher signal intensity on T2WI and lower ADC value than the contralateral substantia nigra, with the astrocytic end-feet and the neuron swollen. On 7d and 14d, the gliosis was significant feature in pathology, but no changes of the MR signal in the sunstantia nigra could be observed. Neither the brains of sham-operated animals showed any noteworthy abnormality. Conclusions: The MR change in the ipsilateral substantia nigra following focal cerebral ischemia of the middle cerebral artery territory was one of the secondary damage in the ischemic exofocal position. MR could reflect the pathological change in some extent, and image the secondary damage in ipsilateral substantia nigra in vivo.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
18
Relationship between primary motor cortex activation and activity of daily living (ADL) motor and ADL process skills in first three months post stroke patients
J. Jansa
U. Puh
Z. Sicherl
K. Angleitner
A. Mesec
D. Suput
University Medical Centre,
Department for neurology
Zaloska 2, 1000 Ljubljana, Slovenia
SLOVENIA
Aim The aim of the study was to evaluate the relationship between the recovery of M1 cortex and ADL motor and ADL process skills. Methods Patients: We included 12 patients, aged 65±15yrs, 2-8 days after first ischaemic stroke. All had partial impaired function of the right (8) or left (4) upper extremity. All were right handed. Assessment tools: We used Assessment of Motor and Process Skills (AMPS) and fMRI. AMPS is an observational assessment providing information about the quality of ADL motor and ADL process skills while a patient carries out a meaningful task. It consists of 16 motor and 20 process skills. A 1.5 T GE Signa scanner was used to collect fMRI data of brain cortex activation during each hand’s functional tasks (thumb-to-fingers opposition, transposition of small objects). Intensity of M1 cortex activation was calculated for each brain hemisphere using AFNI software. AMPS software was used for AMPS data. Spearman correlation coefficients was calculated between M1 activation of each brain hemisphere and ADL motor and ADL process skills. Procedure: Patients were assessed prior and post 2 weeks functional training according to motor learning programme and followed at three months. Results: In week 1 there was a correlation between motor skills and M1 activation in unaffected hemisphere while using hemiparetic (r=0.70) and nonhemiparetic (r=0.64) hand; there was also a correlation between process skills and M1 in the dominant hemisphere during hemiparetic (r=0.60) and nonhemiparetic (r=0.60) hand performance. Patients improve in both skills at week three and month three post stroke. There was no correlation between M1 and skills at week three. At month 3 there was only a correlation between process skills and M1 in affected hemisphere during hemiparetic hand performance (r=0.80). All correlations were at p < 0,05. Conclusions: Results implicate that quality of ADL motor and process skills relate to M1 activation in both hemispheres in early stroke. At month 3 quality of ADL process skills relate to activation in affected hemispheres only.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
19
Inflammatory Response after Occlusion-Reperfusion-Injury to the Brain
F. Leypoldt
M. Gelderblom
K. Steinbach
E. Tolosa
T. Arumugam
C. Gerloff
R. Martin
T. Magnus
Experimental Research in Stroke and Inflammation (ERSI) Department of Neurology, University Hospital Hamburg-Eppendorf
GERMANY
schemic stroke is a devastating disease that represents the third leading cause of death in the western world. Early restoration of blood flow remains the treatment of choice for limiting brain injury following stroke but reperfusion of the ischemic brain can augment the inflammatory response that may cause additional injury to the cerebral microcirculation and adjacent brain tissue. Post-ischemic inflammation is a dynamic process involving a complicated spatially and temporally regulated interaction among various inflammatory cells and molecules. Objectives: The elucidation of the temporal distribution of inflammatory cell influx into the brain and their pattern of activation after ischemic stroke is not well characterized. Although T-cells have been shown to be an essential part of the post-ischemic tissue damage, not much is known about the dynamics of antigen presenting cells, e.g. dendritic cells, monocytes and resident microglia in this setting. Methods: The temporal distribution and activation patterns of different inflammatory cells after temporal middle cerebral artery occlusion (tMCAO) in mice was examined by whole brain FACS characterization of CD3, CD4, CD8, CD45, B220, CD11b, CD11c, NK1.1 and activation markers such as CD69, CD25, MHCII, B7-1 and B7-2. Results: Beginning 6 hours after tMCAO and peaking at 72 hours a strong ipislateral but not contralateral increase in activated dendritic cells was observed. In parallel, CD4 and CD8 positive lymphocytes appeared ipis- and contralateral after 12 hours and peaked at 72 hours. Conclusion: After experimental stroke, there is a significant increase in specialized antigen presenting cells ipislateral in the brain. These seem to communicate with other invading inflammatory cells such as lymphocytes. The further characterization of the molecular mechanisms and modulation of these could provide valuable insights into regulation of postischemic inflammation. These analyses might eventually allow the therapeutic shaping of the immune response towards a more restorative response rather than towards a detrimental immune response.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
20
Identification of novel gene expression associated with vascular remodeling in atherosclerosis plaque development using laser capture microdissection
M.M.Turu
J. Krupinski
L. Badimon
A. Luque
M. Slevin
Cardiovascular Research Centre, CSIC-ICCC, Hospital de la Santa Creu i SAnt Pau, Barcelona
SPAIN
Background and purpose: Neovascularization is an important feature of plaque development. Neovessels infiltrating to the intima from the adventicia are often malformed, leaky and contribute to instability of carotid plaques. Identification of the factors responsible for initiating and propagating angiogenesis will provide the bases for novel therapeutic strategies. Methods: Here, we used laser capture microdissection, real-time PCR and our own designed angiogenenic microfluidic cards to compare active (cd105/flt-1 positive) intimal sites of blood vessel formation with quiescent, non-active areas (n=10). Results: We have identified several novel de-regulated angiogenic genes including G-CSF and Notch-3 and have determined a pro/anti-angiogenic profile for these areas. Discussion: Modulation of the pro-angiogenic switch might reduce development of complicated haemorrhagic plaques and subsequent thrombosis.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
21
AUTOLOGOUS BONE MARROW MONONUCLEAR CELL TRANSPLANTATION IN PATIENTS WITH ISCHEMIC STROKE
G.R.de Freitas
V. Battistella
L.M. Barbosa da Fonseca
B. Gutfien
R. C.Goldemberg
E. Wajnberg
D. Mercante
R. Mendez-Otero
C. André
Universidade Federal do Rio de Janeiro
BRAZIL
Objective: The main goal of this study is to assess the safety and exequibility of bone marrow mononuclear cell (BMMC) transplantation in patients with ischemic stroke in the territory of the middle cerebral artery within 90 days after the beginning of symptoms. In addition, we tested whether the injected cells can be visualized in the brain with SPECT scan. Methods: A total of 10 patients will be included in the study. Up to now 5 patients with cerebral infarcts within the middle cerebral artery territory, and NIHSS between 4 and 13 were treated with BMMC according to a defined protocol (NCT00473057). All patients were men (ages 24, 65, 47, 65, 57; NIHSS 7, 9, 4, 13 and 9, respectively) and had a stroke 67, 82, 62, 72 and 59 days prior to the cell transplantation, respectively. Bone marrow biopsy with BMMC separation was performed and 10% of the cells were marked with tecnecium and reinjected in the middle cerebral artery (5 x 108 ; 1,25 x 108 ; 3,9 x 108; 4 x 108 and 3,2 x 108, respectively) via femoral artery. Single photon emission computed tomography (SPECT) was done two hours after the procedure and whole body scan and planar images were taken 24h after the injection. Eletroencephalogram was performed between the second and the seventh day after autologous transplantation. Patients were evaluated with blood tests, NIHSS and Rankin scales and Barthel index, 3, 7, 30, 60, 90 and 120 days after autologous transplantation. Results: None of the patients treated showed any complication during the procedure. Three patients have images of cell presence in the area of ischemia in SPECT and in the whole body scan. Three patients have already completed the protocol (120 days) whithout any adverse events. Conclusion: Our preliminary results show that BMMC transplantation in patients with ischemic stroke appears to be feasible and safe. We also showed that BMMC can reach the ischemic area and remain in this region at least for 24 hours.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
22
Ocular Microtremor in Acute Stroke
N. Collins
M. al-Kalbani
S. Walsh
M. Martin
G. Boyle
J. Harbison
D. Coakley
Dept. Medical Gerontology, Trinity College Dublin
IRELAND
Background Ocular microtremor (OMT) is a minute high frequency eye movement present in all individuals. OMT frequency is related to brainstem function and is reduced in neurological disorders such as coma and brainstem death. OMT frequency can be measured rapidly and easily at the bedside, using a relatively inexpensive portable device, and may be a useful adjunct in stroke diagnosis. The aim of this case-control study was to investigate whether OMT frequency was reduced in patients with stroke diagnosed clinically and radiologically, when compared to neurologically normal subjects. Methods OMT frequency was measured using a piezoelectric transducer in patients diagnosed with an acute stroke by the specialist stroke service, based on clinical and radiological evidence. OMT measurements were performed within two weeks of stroke onset, the majority being performed within seven days. Results We matched 20 stroke patients (cases) with 20 neurologically normal controls. The stroke group had a significantly lower OMT frequency (mean 73.0Hz; median 75.2Hz; 95% CI: 68.8Hz, 77.2Hz) than the control group (mean 88.9Hz; median 89.0Hz; 95% CI: 86.6Hz, 91.2Hz) (p < 0.001). Following correction for age using multivariate analysis of variance, an average OMT value of 75.3Hz (95% CI: 71.7Hz, 78.9Hz) was obtained in the stroke group compared with 86.6Hz (95% CI: 83.0Hz, 91.2Hz) in the control group (p < 0.001). Area under the receiver operating characteristic (ROC) curve (AUC) demonstrated that OMT could distinguish between stroke cases and controls (AUC 0.973, p < 0.001). An OMT value of </= 82.5Hz was observed in 18/20 of the stroke patients and 1/20 of the controls, affording a sensitivity of 90% and specificity of 95% for differentiation between stroke cases and controls in this study. Conclusions Ocular microtremor frequency was reduced in patients with confirmed stroke compared to neurologically normal subjects. The results suggest that OMT activity may be of value in the investigation of stroke.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
23
Core Temperature Versus Intracerebral Temperature Monitoring In Hypothermia For Neuroprotection
S. Griffith
J.L.Caron
R. Coleman
M. Stiegelman
J. Salazar
R. DiGeronimo
University of Texas Health Science Center at San Antonio and Wilford Hall USAF Medical Center
USA
Background: Hypothermia is neuroprotective in the management of ischemic stroke and confers advantages over pharmacologic agents. Core temperature readings are relied upon clinically for cerebral hypothermic neuroprotection. There is little data correlating core temperature with actual brain temperature. We present data comparing core and actual brain temperature in a swine hypothermia model. Methods: Immature Yorkshire swine (n=7) were implanted with cerebral catheters for direct measurement of intracranial temperature (Licox®). Esophageal and rectal probes were placed for core temperature readings. With other physiologic variables controlled, baseline temperature measurements were obtained. All animals were then subjected to 60 minutes of hypothermia, followed by 4 hours of recovery. Both cerebral and core temperatures were continuously monitored. Results: At baseline temperatures, regression analysis of both esophageal and rectal temperatures revealed good correlation with cerebral temperatures (Brain vs Esophageal R=0.903, R2=0.817, p<.001 and Brain vs Rectal R=0.823, R2=0.677, p<.001). During hypothermia, esophageal temperature correlated significantly better with actual cerebral temperature than rectal temperature (Brain vs Esophageal R=0.371, R2=0.138, p.028 and Brain vs Rectal R=0.051, R2=0.003, p.772). Overall correlations during normothermia and hypothermia revealed brain vs esophageal (R = 0.958, R2 = 0.917, p < .001), and brain vs rectal temperature (R = 0.928, R2 = 0.860, p < .001), respectively. Conclusion: Core temperature alone may underestimate the level of cerebral hypothermia. More work needs to be done to better understand the relationship between core and cerebral temperature in hypothermia.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
24
Protective Effect of Simvastatin on PC12 Cells in Oxygen and Glucose Deprivation Injury
Z.L.Guo
S.D.Chen
the Medical College of Shanghai Jiaotong University, Shanghai 200025
CHINA
Objective: To investigate the protective effect of simvastatin pretreatment with different concentration on PC12 cells in oxygen and glucose deprivation injury. Methods: PC12 cells were treated under oxygen and glucose deprivation(OGD) for one hour and reperfusion for 24 hours. They were divided into the normal control group, OGD group and OGD with simvastatin group which were pretreated one hour prior to OGD by different concentration(the final concentration was 0.01µmol/L, 0.1µmol/L, 1µ;mol/L, 1µmol/L).The contents of NO, IL-1β and TNFαamong all groups were measured after the reperfusion model of PC12 cells was established. The cell viability was assayed by MTT, and the cell apoptosis rate was determined by Annexin V. Results: Contents of NO, IL-1βand TNFα; of OGD group were significantly increased than those of the other groups(P<0.01). The cell viability was obviously decreased in OGD group than that in the other groups (P<0.01). The cell apoptosis rate was also significantly increased in OGD group. No significant statistically difference was found between normal control group and OGD with simvastatin group (P>0.05). Conclusion: Simvastatin pretreatment with different concentration has protective effect on PC12 cells in OGD in vitro. KEY WORDS: Simvastatin, PC12 cell, OGD, Reperfusion
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
25
Cytokine treatment reduces ADAMTS-13 expression in central nervous system cells in vitro: possible involvement in transient ischaemic attack (TIA)?
G.A.Frentzou
K. Harkness
G. Haddock
N. Woodroofe
A. Cross
Sheffield Hallam University
UNITED KINGDOM
Deficiency in ADAMTS-13, the thirteenth member of the ADAMTS family, is associated with a variety of thrombotic microangiopathies, with decreased Von Willebrand factor (VWF)-cleaving activity including thrombotic thrombocyotpenic purpura (TTP). Furthermore, ADAMTS-13 levels are also shown to be reduced in patients undergoing major surgery and non symptomatic carriers of TTP. VWF levels are known to be increased during stroke. In the present study ADAMTS-13 expression was investigated in four different central nervous system derived cell lines, SHSY-5Y (human neuroblastoma), U373 (human astroglioma), CHME-3 (human microglial) and hCMEC/D3 (human brain endothelial) in response to cytokine treatment. The cells were cultured until confluent and then plated at a density of 100000 cells/ well in a 24 well plate. Further to this the cells were treated with a range of concentrations (0-100 ng/ml) of IL-1 beta, TNF alpha, IL-6 and ADAMTS-13 expression was measured by real-time PCR. All the cell lines expressed ADAMTS-13 in different levels. IL-1 beta was shown to down-regulate ADAMTS-13 mRNA expression in human astroglioma (U373) and human microglial cells (CHME-3). Surprisingly, TNF alpha in low concentrations was shown to up-regulate ADAMTS-13 expression, in human neuroblastoma cells (SHSY-5Y). Treatment of cells with IL-6 showed no significant differences in ADAMTS-13 expression. We suggest that under conditions of a TIA or stroke, where cytokine expression is increased, ADAMTS-13 homeostasis could be altered with possible detrimental physiological consequences.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
26
The Importance of MAP Kinase Pathway in the Upregulation of Contractile ETB Receptors in Rat Cerebral Arteries
H.K.Sandhu
S. Ansar
L. Edvinsson
Department of Clinical Experimental Research, Glostrup Research Institute
DENMARK
Objective: Previous studies have shown that contractile endothelin type B (ETB) receptors are upregulated in cerebral arteries after experimental subarachnoid hemorrhage (SAH) and focal cere-bral ischemia. This upregulation happens via de novo transcription and is associated with enhanced phosphorylation of extracellular signal-regulated protein kinase 1/2 (pERK 1/2). We hypothesize that inhibition of the MAPK pathway alters the ETB receptor upregulation. Method: Rat middle cerebral arteries (MCAs) were incubated for 48 h with or without kinase in-hibitor. The MCAs were mounted in myographs and contractile responses to the ETB specific recep-tor agonist Sarafotoxin (S6c) and the ETA and ETB specific receptor agonist Endothelin-1 (ET-1) were studied. Real Time PCR was used to measure the ETB and ETA receptor mRNA levels. Results: After organ culture S6c induced vasoconstriction. The MEK 1/2 inhibitor U0126 (10-5 M) diminished the contractile response to S6c with 60 %, while the ERK 1/2 inhibitor AG126 (10-6 M) diminished both the S6c and ET-1 contractile response with 60 %. Real time PCR showed that U0126 and AG126 diminished the ETB mRNA level. In addition to U0126 and AG126, additional two MEK 1/2 inhibitors were tested: PD98059 (10-7 M to 10-5.5 M) and SL327 (10-7 M to 10-4.5 M). They did not show any inhibiting effect. Conclusion: Our results show that the MAPK kinase pathway has a role in the upregulation of the contractile ETB receptors in rat cerebral arteries. Since there is similar results in cerebral arteries after experimental SAH and focal cerebral ischemia these results are of pathophysiological impor-tance in cerebrovascular diseases. The in vitro organ culture model can be used to study the path-way leading to upregulation of the contractile ETB receptors.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
27
Gamma-hydroxybutyrate (GHB) accelerates functional recovery after focal cerebral ischemia
B. Gao
E. Kilic
C.R.Baumann
D.M.Hermann
C.L.Bassetti
University Hospital Zürich
SWITZERLAND
Background and purpose: Gamma-hydroxybutyrate, a natural metabolite of GABA and a drug used in humans to promote slow wave sleep and treat narcolepsy, has been suggested to protect against ischemia stroke at high dosages. This study aimed to assess recovery-promoting effects of GHB at a low dosage similar to that used in patients. Methods: Adult mice, subjected to 30 minutes of intraluminal middle cerebral artery occlusion, were intraperitoneally treated with GHB (100mg/kg, twice/day, 8 hours apart) or saline for 10 days. Motor recovery was evaluated by the grip-strength test. Brain lesion was assessed by cresyl violet and NeuN staining 5 weeks after stroke. Expression of neuroplasticity-related genes (GAP43, c-jun, neurocan and ephrinB1) was analyzed by Taqman real-time PCR. Results: GHB-treated mice regained their body weight faster and recovered grip-strength (3 weeks after stroke) more quickly than saline-treated mice. This was noteworthy as GHB did not influence ischemia-induced brain injury, as revealed by cresyl violet and neuronal staining. Taqman PCR assay revealed a decreased expression of c-jun and neurocan in the ischemic striatum of GHB-treated mice in comparison to saline-treated mice. Conclusion: GHB at a low dose accelerates neurological recovery following ischemic stroke. Further studies are necessary to determine the potential relationship between GHB, neuroplasticity, sleep and stroke recovery.
Session:
Poster Session I
Date:
Wednesday 14 May 2008
Time:
12:30 - 14:00
Room:
Agora 3
28
Molecular imaging and quantification of adhesion molecules in rat MCAO model
I.C.Kanter
G. Ladewig
C. Kleinschnitz
M. Reinhardt
P. Hauff
R. Kollmar
S. Schwab
M. Mäurer
Friedrich-Alexander Universität Erlangen-Nürnberg
GERMANY
Background: Cell adhesion molecules show a significant and early upregulation after occlusion of the MCA. These molecules are therefore an attractive target for molecular imaging in order to get insight into the immunological processes in acute stroke. Methods: We used the newly developed ultrasound based quantification technology “sensitive particle acoustic quantification” (SPAQ) for imaging and quantification of specific gas-filled microparticles (MP) targeted against the adhesion molecules ICAM-1. With this approach we aimed to monitor the molecular changes at the blood-brain-barrier in a 2 hour rat MCAO model. Results: In vivo imaging in living anesthetized animals revealed a clear stimulated acoustic emission (SAE) signal in the ischemic hemisphere. However, compared to ICAM1-specifc microparticles unspecific and unlabled microparticles also showed a very prominent SAE signal. This results point to an unspecific entrapment of ultrasound contrast agent in the ischemic hemisphere. However, when we analysed regions of interest outside the ischemic area we could demonstrate specific and time dependent upregulation of ICAM-1. Conclusion: Here we present the first results of SPAQ imaging in an animal model for acute stroke. Our results demonstrate that ultrasound based molecular imaging has the potential to become a valuable tool for monitoring of molecular expression patterns at the blood brain barrier in acute stroke. However, in contrast to inflammatory models unspecific entrapment of microparticles seem to be an obstacle for this method in MCAO rats.
