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(Stroke. 2000;31:852.)
© 2000 American Heart Association, Inc.
Original Contributions |
Nitric Oxide–Related Brain Damage in Acute Ischemic Stroke
From the Department of Neurology, Complejo Hospitalario Universitario, Santiago de Compostela (J.C.); Department of Physiology, Universidad de Barcelona, Barcelona (R.R.); and the Section of Neurology, Hospital Universitari Doctor Josep Trueta, Girona (A.D.), Spain.
Correspondence to Dr Antoni Dávalos, Section of Neurology, Hospital Universitari Doctor Josep Trueta, E-17007 Girona, Spain. E-mail adavalose{at}meditex.es
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Abstract |
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Background and Purpose—The neurotoxic and neuroprotective role of nitric oxide (NO) in experimental cerebral ischemia has generated considerable debate. The aim of this study was to analyze the relationship between NO metabolite (NO-m) concentrations in cerebrospinal fluid (CSF) and clinical and neuroimaging parameters of brain injury in patients with acute ischemic stroke.
Methods—We studied 102 patients and 24 control subjects who were included in a larger previous study conducted to analyze risk factors of progressing stroke. NO generation was calculated by quantifying nitrates and nitrites with a colorimetric assay in CSF samples obtained within the first 24 hours from symptoms onset. Early neurological deterioration was defined as a fall of 1 or more points in Canadian Stroke Scale score between admission and 48 hours after inclusion. Infarct volume was measured on days 4 to 7 by cranial CT.
Results—Median NO-m concentrations [quartiles] were 2.1 [1.0, 4.5] µmol/mL in patients and 1.0 [1.0, 1.0] µmol/mL in control subjects (P<0.0001). In 45 patients with subsequent early neurological deterioration, NO-m levels in CSF were significantly higher than in those with stable stroke (4.0 [1.7, 7.8] versus in 1.6 [1.0, 2.5] µmol/mL, P<0.0001). There was a moderate correlation between NO-m and infarct volume (coefficient 0.39, P<0.001). NO-m concentrations >5.0 µmol/mL were significantly associated with early neurological worsening (OR 5.7, 95% CI 1.2 to 27.4; P=0.030) independent of other important factors related to progressing stroke, such as CSF glutamate levels.
Conclusions—Our clinical findings suggest an important role of NO generation in acute ischemic stroke. Increased NO-m in CSF are associated with a greater brain injury and early neurological deterioration.
Key Words: excitotoxicity • nitric oxide • stroke outcome • stroke, acute • tomography, x-ray computed
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Introduction |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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Nitric oxide (NO) is an inorganic gas that plays a part in the control of cerebral blood flow, thrombogenesis, and modulation of neuronal activity.1 2 NO is produced in the endothelial cells, neurons, glia, and macrophages by 3 different isoforms of the enzyme nitric oxide synthase (NOS).3 4 NOS mediates the conversion of L-arginine and oxygen to NO and citrulline.5 Under cerebral ischemia, high concentrations of NO generated by the calcium-dependent activation of the constitutive neuronal NOS (nNOS) and by the activation of the inducible form of NOS (iNOS) in macrophages and other cell types intervene in inflammatory and cytotoxic actions that lead to neuronal death.1 6 7 8 9 In contrast, NO generated by the activation of the constitutive endothelial NOS (eNOS) may have protective effects that decrease the ability of platelets to aggregate, prevent leukocyte endothelial adhesion, and increase vascular dilation and cerebral blood flow.2 3 10 This dual role of NO in cerebral ischemia, neurotoxic and neuroprotective, has generated a considerable scientific debate and conflicting findings in experimental stroke models.11 12
Because NO has an extremely short half-life, its detection in the brain is difficult. Current assays in vivo include fluorometric measurements of its stable metabolite nitrite and indirect methods such as nitrate concentrations and citrulline concentrations as markers of NOS activity.13 14 The determination of NO metabolites could be useful to clarify whether NO generation is predominantly associated with increased or reduced brain injury in human stroke, and consequently to support the use, and to evaluate the effect, of future treatments with specific NOS antagonists in clinical trials.15 In this study we analyzed the relationship between NO metabolite concentrations in the cerebrospinal fluid (CSF) of patients during the acute phase of cerebral ischemia and clinical and neuroimaging parameters of brain injury.
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Subjects and Methods |
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For the purpose of this investigation, we studied 102 patients (63% males, mean age 67.3±10.9 years), from a larger series of 249 patients admitted consecutively with a first episode of hemispheric ischemic stroke within 24 hours after the onset of symptoms, and 24 individuals (46% males, mean age 54.3±15.9 years) taken from a group of 50 control subjects. The total number of patients and controls were included in a prospective clinical investigation conducted between October 1992 and December 1996 to analyze clinical, biochemical, and radiological factors related to progressing stroke. The protocol was approved by the Ethics Committee of Hospital Xeral de Galicia, and informed consent was given by the control subjects, the patients themselves, or by relatives in cases of patients’ inability to understand our information. Only patients and controls from whom we had obtained and stored (frozen) CSF samples for possible future laboratory determinations were evaluated in the present study. Availability of samples relied exclusively on the amount of CSF obtained at the time of inclusion and not on other clinical or biochemical factors. Therefore, this selection was performed without knowledge of stroke outcome and excitatory amino acid concentrations and before determination of NO metabolites.
The characteristics of the control group and the inclusion criteria for the whole series of patients have already been published.16 17 In summary, all patients had a persistent focal neurological deficit and absence of mass effect or cerebral hemorrhage on the cranial CT carried out before inclusion. Type of stroke was classified accordingly to the TOAST criteria.18 On admission, blood pressure and body temperature were recorded and blood chemistry studies, chest x-ray, ECG, and nonenhanced cranial CT were performed. Stroke severity was assessed by a neurologist immediately after these tests with the Canadian Stroke Scale (CSS). The time (in hours) from symptoms onset until the neurological attention was 7.7±5.2 (range, 1.5 to 23 hours); this time interval was considered the inclusion delay. A lumbar puncture was performed if there were no signs of mass effect on CT; the interval between inclusion and taking the CSF sample was 2.6±3.7 hours.
All the CT examinations were carried out with a CT System 3000 plus (GEC) scanner with a 512x512 display. Early signs of infarction were carefully evaluated during the first examination; these signs included focal hypodensity consistent with the clinical picture, and the indirect signs of cerebral infarction like obscuration of the lenticular nucleus, obscuration of the cortex, hyperdensity of the middle cerebral artery and mass effect on the cortical sulci and structures of the median line. A second scan was performed between the fourth and seventh day after inclusion to determine the size of the lesion according to the formula 0.5xaxbxc (where a and b are the largest perpendicular diameters measured on CT and c is the slice thickness). Infarct topography was classified as cortical when the hypodensity area on the second CT affected predominantly the cortical regions, or as deep when the infarct was limited to the internal capsule, corona radiata, centrum semiovale, caudate, putamen, globus pallidus, or thalamus. All CT evaluations were performed by the same radiologist, who was blinded to the clinical and biochemical results.
Only patients with systolic blood pressure of 
220 mm Hg
or diastolic blood pressure of 120 mm Hg received
antihypertensive drugs during the first 48 hours. Subcutaneous low-dose
heparin was given as prophylaxis against pulmonary
thromboembolism. Antiplatelet drugs were prescribed during
hospitalization in atherothrombotic and lacunar infarcts.
Anticoagulants were administered only to patients with suspected
cardioembolism when cranial CT and clinical examination
excluded a large cerebral infarct. Glucose infusions,
corticosteroids, nimodipine, hemodilution, and
thrombolytics were not allowed. A potential infectious
focus was examined in all patients with hyperthermia (axillary
temperature of >37.5°C) during the first week of hospitalization
following the method described in a previous work.19
Two clinical outcome measures were evaluated: early neurological deterioration, as a potential sign of enlarging brain injury, and functional capacity at 3 months. Following previously published criteria,20 early neurological deterioration was diagnosed when the CSS score dropped 1 or more points between the evaluation performed at inclusion and an assessment repeated at 48 hours of hospitalization by the same physician. Patients whose symptoms worsened exclusively in terms of orientation, or who remained stable or improved in the same 48-hour period, were classified as having nonprogressing cerebral infarction. We assessed patients’ functional condition at 3 months with the Barthel Index (BI). Poor outcome was defined as death or BI score of <85. This score corresponds to a state in which patients report needing help in performing activities of daily living, with a sensitivity of 95% and a specificity of >80%.21 22
Laboratory Determinations
CSF samples were prepared by centrifugation
(2000g for 10 minutes) and immediately stored at -80°C.
Quantification of glutamate and L-arginine was
performed by HPLC following the method described
elsewhere.16 Amino acid determinations were performed
with blinding to clinical and neuroimaging findings and to stroke
outcome.
NO generation was calculated by quantifying the final products of its reactions, nitrates and nitrites, by a colorimetric assay (Cayman Chemical Co).14 This procedure was carried out after the end of the inclusion period in a different laboratory, which had no knowledge of patient or control origin of the CSF sample, stroke characteristics, or amino acid concentrations. The nitrates in the CSF were enzymatically converted into nitrites by incubation with nitrate reductase and NADPH at room temperature for 2 hours. The levels of nitrites were then calculated. Nitrite concentration was measured with use of the Griess reaction by adding 100 µL of Griess reagent (1% sulfanilamide and 0.1% naphthylethylenediamide in 5% phosphoric acid) and mixed in 96-well plates and shaken gently for 20 minutes at room temperature. The addition of the Griess reagent results in a colorimetric product measured at 540 nm.23 The measurement was taken in a microplate read off a standard curve generated by dilutions of nitrate. The assay is sensitive to nitrite concentrations of approximately 1 µmol/L. Interassay variability was 6%, and intra-assay variability was 2.9%. The final product of NO metabolites (NO-m) was used in this clinical investigation.
Statistical Analyses
NO-m concentrations are expressed as median [quartiles],
because they were not normally distributed. Comparison of NO-m between
2 groups was performed with the Mann-Whitney test and between >2
groups with the Kruskal-Wallis test. Spearman analysis was used
for bivariate correlations between NO-m and CSS score, inclusion delay,
L-arginine, glutamate concentrations, and infarct
volume.
The independent relationship between NO-m and infarct volume was evaluated by multiple linear regression analysis. A log transformation of infarct volume was performed to achieve a normal distribution of the dependent variable. The importance of NO-m concentrations for subsequent early neurological deterioration (0=no, 1=yes) and poor outcome (0=no, 1=yes) was assessed by logistic regression analysis based on the maximum likelihood ratio. Model 1 was fitted with a stepwise procedure, adjusting for age, inclusion delay, body temperature, serum glucose, and CSS score on admission; early CT signs of cerebral infarct; infections detected within the first week after stroke; and ultimate infarct volume on the second CT. These variables were related to neurological deterioration and poor outcome in our previous investigations conducted in the same series of patients (P<0.1).17 24 This model was further adjusted for glutamate concentration on admission, which has been the most important factor related to neurological worsening17 (model 2). NO-m, glutamate, and ultimate infarct volume were included as categorical variables because risk did not change linearly with increasing values of these variables. Cutoff values were obtained following the method described by Robert et al.25
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Results |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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NO-m concentrations in CSF were significantly higher in patients than in control subjects (2.1 [1.0, 4.5] versus 1.0 [1.0, 1.0] µmol/mL, P<0.0001). Forty-three patients (42.1%) were diagnosed as having a large-artery atherosclerotic cerebral infarct, 33 (32.4%) a cardioembolic infarct, 13 (12.7%) a lacunar infarct, and 13 (12.7%) a cerebral infarct of unknown origin. No significant differences were observed in NO-m concentrations between stroke subtypes (P=0.71).
Clinical Outcome
NO-m concentrations correlated to CSS score at admission
(coefficient -0.54, P<0.001; Figure 1
). A subsequent early neurological
deterioration was observed in 45 patients (44%), 21 with cortical
infarcts, 10 with deep infarcts, 12 with cortical and deep infarcts,
and 2 with normal CT. Median NO-m concentration was 4.0 [1.7, 7.8]
µmol/mL in the group with early neurological worsening and 1.6 [1.0,
2.5] µmol/mL in the group with nonprogressing cerebral infarct
(P<0.0001). The significant difference between progressing
and nonprogressing strokes was observed in patients with cortical
infarcts (4.5 [1.8, 14.2] versus 1.6 [1.0, 3.0] µmol/mL,
P=0.002), deep infarcts (4.5 [1.6, 12.1] versus 1.6 [1.0,
2.5] µmol/mL, P=0.019), and with infarcts involving
cortical and subcortical regions (3.5 [1.6, 7.7] versus 1.0 [1.0,
2.1] µmol/mL, P=0.029).
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Death and poor outcome at 3 months was observed in 9 (9%) and 44 (43%) patients, respectively. NO-m concentrations on admission were significantly higher in patients with poor outcome (4.2 [2.0, 9.8] µmol/mL) than in those who survived with good functional capacity at 3 months (1.6 [1.0, 2.8] µmol/mL) (P<0.0001).
Neuroimaging Findings
NO-m concentrations were significantly higher in the 65 patients
with early signs of cerebral infarct on the initial CT scan than in the
37 patients without early CT signs (3.0 [1.6, 5.7] versus 1.6 [1.0,
2.4] µmol/mL) (P=0.001). There was a significant
correlation between NO-m and ultimate infarct volume on days 4 to 7
(coefficient 0.39, P<0.001). No differences were observed
on NO-m depending on infarct topography in the 97 patients who showed a
corresponding lesion on the second CT: CSF NO-m concentrations were 2.2
[1.0, 4.6] µmol/mL in 46 cortical infarcts, 1.6 [1.4, 4.4]
µmol/mL in 34 deep infarcts, and 2.2 [1.3, 5.2] µmol/mL in 17
infarcts that involved the cortical and subcortical regions
(P=0.84).
Multivariate Analyses for Brain
Injury
NO-m concentrations were not independently related to the
log-transformed infarct volume in a multiple linear regression model,
after adjustment for age, delay to inclusion, CSS score, body
temperature, and serum glucose on admission (Table 1). Table 2
shows the result of logistic regression analyses for early
neurological deterioration as a dependent variable. NO-m
concentrations in CSF >5.0 µmol/mL were independently and
significantly associated with early neurological worsening (OR 5.3,
95%CI 1.5 to 18.5; model 1). These findings did not change after
adjustment for glutamate concentrations in CSF (model 2). NO-m over the
5.0 µmol/mL cut point did not predict death or dependency at 3
months. Age, stroke severity on admission, inclusion delay, and body
temperature were the only factors selected by the logistic model.
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Biochemical Studies
In the total series of patients, there was a positive correlation
between NO-m concentrations in CSF and the inclusion delay (coefficient
0.41, P<0.001). In patients with stable stroke, NO-m values
did not change significantly with time, although a slight increase was
observed in those studied after the first 12 hours from stroke onset
(P=0.063). In contrast, NO-m concentrations were
significantly higher in CSF samples obtained after the first 6 hours
from stroke onset in the group of patients who had a subsequent
neurological deterioration (P=0.005). Furthermore, in
patients who deteriorated, the median values determined at any time
interval during the first 24 hours from onset were higher than those
obtained in the stable stroke and control groups (Figure 2).
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)
ischemic stroke. Numbers indicate the number of patients
studied. Dotted line indicates the median values and quartiles of NO-m
in control subjects. Note that patients with early neurological
deterioration had higher levels at all time intervals during the first
24 hours from stroke onset, which were particularly increased after the
first 6 hours from stroke onset.
In patients, but not in the control group, NO-m concentrations
correlated negatively to L-arginine concentrations in CSF
(coefficient -0.85, P<0.0001). The levels of NO-m were
particularly high when L-arginine concentrations
were <6 µmol/L (Figure 3). As occurred
with NO-m, L-arginine concentrations correlated
with the inclusion delay. This fact was observed in patients with
progressing stroke (coefficient -0.53, P<0.001), and in
those with stable stroke (coefficient -0.35, P=0.008). A
moderate positive correlation was found between NO-m and glutamate
concentrations in CSF (coefficient 0.24, P=0.01).
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Discussion |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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To our knowledge, this is the first study supporting at the clinical setting an important role of NO generation in acute ischemic stroke. We have found a significant increase of NO-m in the CSF of patients studied within the first 24 hours from stroke onset in comparison to controls. Furthermore, we have demonstrated a significant relationship between NO-m concentrations and clinical and neuroimaging indicators of brain injury. In fact, high NO-m concentrations were associated with deleterious effects in cerebral ischemia, that is, higher stroke severity at admission, early neurological deterioration, larger infarct volumes, and poor outcome at 3 months. Although these findings suggest predominant cytotoxic effects as a result of NO generation, we cannot rule out the possibility that the high levels of NO-m obtained in patients with greater brain injury result in part from an increased endothelial NO generation with neuroprotective, but failed, purposes.
NOS activity and NO release are greatly increased in the acutely human ischemic brain,26 but opposite effects, neurotoxic and neuroprotective, have been demonstrated in experimental stroke models. The effect of selective antagonists of NOS isoforms on acute cerebral ischemia11 12 27 28 29 30 and the effect of cerebral ischemia on mouse models in which a single NOS isoform gene is not expressed (knockout mice)31 32 33 have clarified the protective role of eNOS activity and the neurotoxic role of nNOS and iNOS activity. The former is associated with smaller cerebral infarcts and the latter with increased infarct volume. A further example of this dual action is the time-dependent opposite effects of L-arginine, a precursor amino acid of NO that is the principal natural substrate for each of the 3 NOS isoforms. In animal models, L-arginine has neuroprotective effects when it is administered up to 2 hours after onset of cerebral ischemia, but it is associated with increased infarct volume when its administration is delayed by 24 hours.34 The deleterious consequence of the late administration is thought to result from delayed neuronal injury due to the appearance of inducible NOS in the ischemic penumbra, after a time lag of 6 to 12 hours.35 36 37
The method we used in this clinical study did not allow determination
of which portion of the NO-m concentration derived from
endothelial NO, from neuronal NO, or from NO generated
in other cells by the action of iNOS. However, we believe that the
nitrites and nitrates measured in our patients were metabolites of the
NO, because they showed an extremely high negative correlation to
L-arginine concentration in CSF. This finding is in
agreement with the higher consumption of L-arginine
reported in patients with greater cerebral damage by our
group.38 We hypothesize that the exponential negative
relationship between NO-m and L-arginine values found in
our study (Figure 3
) may be explained by the fact that the
synthesis of small quantities of NO may be supplied by the consumption
of intracellular L-arginine, which would not affect levels
in the CSF. However, the activation of NOS isoforms during cerebral
ischemia would increase the demand for L-arginine
and would rapidly reduce extracellular levels.
In our opinion, some findings in this study support the hypothesis that high levels of NO-m derived mainly from a delayed inducible NOS activation, a process that is not mediated by glutamate release and calcium influx.35 First, we found only a moderate correlation between glutamate and NO-m concentrations in CSF, and the effect of NO-m on stroke worsening was independent of the glutamate effect. Second, in patients with neurological deterioration, NO-m levels were higher in CSF samples obtained after 12 hours from symptom onset than in those obtained earlier, which suggests a delayed generation of NO by the iNOS. Although the time course of iNOS expression in the human brain has not yet been defined, iNOS synthesis in rodents begins at 12 hours after permanent middle cerebral artery occlusion.36 Furthermore, although the cerebral dynamics of NO-m are unknown, the delayed appearance of NO-m in CSF differs from the early increase of CSF glutamate reported by our group in patients with progressing stroke.39 Third, the late increase of NO-m was consistent with a delayed consumption of L-arginine, which suggests the activation of iNOS. Finally, NO-m concentrations were equally high in patients with cortical and subcortical cerebral infarcts in the whole group and in patients with early neurological deterioration. This argues against a glutamate-calcium–dependent generation of NO, because glutamate release is significantly higher in cortical than in subcortical infarcts, probably owing to a higher number of glutamatergic neurons in the gray matter.40
The involvement of NO in the progression of cerebral infarction is an appealing hypothesis. The present study shows that NO plays a part in early neurological deterioration, a fact that has been attributed to the expansion of the ischemic area.41 42 43 As occurred with glutamate,17 this relationship was independent to other important predictors and associated factors with progressing stroke, such as hyperthermia, high serum glucose concentrations, early infarct signs on CT, infections detected within the first week of stroke, and ultimate infarct volume. These findings suggest a direct neurotoxic effect of NO on the propagation of ischemic penumbra and neuronal death, a process that is supported by some experimental findings. It has been shown that immediately after ischemia, NOS is much more active in the core than in the surrounding area, which gives rise to an increasing concentration of NO that gradually extends from the core to the vulnerable neighboring neurons in the penumbra.44 This phenomenon may explain in part the progressive expansion of the brain injury in the absence of relevant changes in cerebral blood flow. Although the neurotoxic mechanisms mediated by NO have been only partially elucidated, free radical damage by formation of peroxynitrite may have an important role.45 Peroxynitrite decomposes to other reactive oxygen species, such as radical hydroxyl and nitrogen dioxide, which cause lipid peroxidation and thus destroy cell membranes. This mechanism may be particularly important in the peri-infarct region, because oxygen delivery during reperfusion facilitates a delayed generation of NO and a higher production of peroxynitrite due to the reduction of NO by the superoxide anion.46 47
In conclusion, although current experimental studies show that NO can play a dual role in cerebral ischemia, our clinical findings suggest that increased NO generation is predominantly associated with cytotoxic effects. High NO metabolites in CSF within the first hours of acute stroke predict subsequent early neurological deterioration regardless other biochemical mechanisms mediated by glutamate. The possibility of establishing markers for NOS isoforms48 would provide the opportunity to determine precisely the mechanism for neuronal lesion and how it can be treated.
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Acknowledgments |
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This study was supported by a grant from the Fondo de Investigaciones Sanitarias, Investigation Project FISS 96/1513, and from the Programa de Promoción Xeral de Investigación do Plan Galego de IDT da Xunta de Galicia (PGDIT99PXI20803B).
Received November 19, 1999; revision received January 21, 2000; accepted January 21, 2000.
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References |
|---|
|
TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
|---|
1. Moncada S, Palmer RMJ, Higgs EA. Nitric oxide: physiology, pathophysiology, and pharmacology. Pharmacol Rev. 1991;43:109–142.[Medline] [Order article via Infotrieve]
2. Faraci FM, Brian JE Jr. Nitric oxide and the cerebral circulation. Stroke. 1994;25:692–703.[Abstract]
3. Iadecola C, Pelligrino DA, Moscowitz MA, Lassen NA. Nitric oxide synthase inhibition and cerebrovascular regulation. J Cereb Blood Flow Metab. 1994;14:175–192.[Medline] [Order article via Infotrieve]
4. Dalkara T, Moskowitz MA. The role of nitric oxide in cerebral ischemia. In: Welch KMA, Caplan LR, Reis DJ, Siesjö BK, Weir B, eds. Primer on Cerebrovascular Diseases. San Diego, Calif: Academic Press; 1997:207–208.
5.
Moncada S, Higgs A. The
L-arginine-nitric oxide pathway. N Engl J
Med.. 1993;329:2002–2012.
6.
Bredt DS, Synder SH. Isolation of nitric oxide
synthetase, a calmodulin-requiring enzyme. Proc Natl
Acad Sci U S A. 1990;87:682–685.
7. Dawson VL, Dawson TM. Nitric oxide neurotoxicity. J Chem Neuroanat. 1996;10:179–190.[Medline] [Order article via Infotrieve]
8. Baumgartner WA, Redmond M, Brock M, Tseng E, Blue ME, Troncoso JC, Johnston MV. Pathophysiology of cerebral injury and future management. J Card Surg. 1997;12(suppl 2):300–310.
9. Forman LJ, Liu P, Nagele RG, Yin K, Wong PY. Augmentation of nitric oxide, superoxide, and peroxynitrite production during cerebral ischemia and reperfusion in the rat. Neurochem Res. 1998;23:141–148.[Medline] [Order article via Infotrieve]
10. Stagliano NE, Zhao W, Prado R, Dewanjee MK, Ginsberg MD, Dietrich WD. The effect of nitric oxide synthase inhibition on acute platelet accumulation and hemodynamic depression in a rat model of thromboembolic stroke. J Cereb Blood Flow Metab. 1997;17:1182–1190.[Medline] [Order article via Infotrieve]
11. Dalkara T, Yoshida T, Irikura K, Moskowitz MA. Dual role of nitric oxide in focal cerebral ischemia. Neuropharmacology. 1994;33:1447–1452.[Medline] [Order article via Infotrieve]
12. Iadecola C. Bright and dark sides of nitric oxide in ischemic brain injury. Trends Neurosci. 1997;20:132–139.[Medline] [Order article via Infotrieve]
13.
Karder A, Frazzini VI, Solomon RA, Trifiletti RR.
Nitric oxide production during focal cerebral ischemia
in rats. Stroke. 1993;24:1709–1716.
14. Archer S. Measurement of nitric oxide in biological models. FASEB J. 1993;7:349–360.[Abstract]
15.
Chabrier PE, Auguet M, Spinnewyn B, Auvin S,
Cornet S, Demerle-Pallardy C, Guilmard-Favre C, Marin JG, Pignol B,
Gillard-Roubert V, Roussillot-Charnet C, Schulz J, Viossat I, Bigg D,
Moncada S. BN 80933, a dual inhibitor of neuronal nitric
oxide synthase and lipid peroxidation: a promising neuroprotective
strategy. Proc Natl Acad Sci U S A. 1999;96:10824–10829.
16.
Castillo J, Dávalos A, Naveiro J, Noya M.
Neuroexcitatory amino acids and their relation to infarct size and
neurological deficit in ischemic stroke. Stroke. 1996;27:1060–1065.
17. Castillo J, Dávalos A, Noya M. Progression of ischaemic stroke and excitotoxic amino acids. Lancet. 1997;349:79–83.[Medline] [Order article via Infotrieve]
18.
Adams HP, Bendixen BH, Kappelle LJ, Biller J,
Love BB, Gordon DL, Marsh EE III, and the TOAST Investigators.
Classification of subtype of acute ischemic stroke: definitions
for use in a multicenter clinical trial. Stroke. 1993;24:35–41.
19.
Castillo J, Dávalos A, Marrugat J, Noya M.
Timing for fever-related brain damage in acute ischemic stroke.
Stroke. 1998;29:2455–2460.
20.
Dávalos A, Cendra E, Teruel J,
Martínez M, Genis D. Deteriorating ischemic stroke:
risk factors and prognosis. Neurology. 1990;40:1865–1869.
21.
Kay R, Wong KS, Perez G, Woo J. Dichotomizing
stroke outcomes based on self-reported dependency.
Neurology. 1997;49:1694–1696.
22.
Sluter G, Steen C, De Keyser J. Use of Barthel
index and modified Rankin scale in acute stroke trials.
Stroke. 1999;30:1538–1541.
23. Gilliam MB, Sherman MP, Griscavage JM, Ignarro LJ. A spectrophotometric assay for nitrate using NADPH oxidation by aspergillus nitrate reductase. Anal Biochem. 1993;212:359–365.[Medline] [Order article via Infotrieve]
24. Dávalos A, Castillo J, Pumar JM, Noya M. Body temperature and fibrinogen are related to early neurological deterioration in acute ischemic stroke. Cerebrovasc Dis. 1997;7:64–69.
25. Robert C, Vermont J, Bosson JL. Formulae for threshold computations. Comput Biomed Res. 1991;24:514–529.[Medline] [Order article via Infotrieve]
26. Foster C, Clark HB, Ross ME, Iadecola C. Inducible nitric oxide synthase expression in human cerebral infarcts. Acta Neuropathol (Berl). 1999;97:215–220.[Medline] [Order article via Infotrieve]
27.
Zhang F, Casey RM, Ross ME, Iadecola C.
Aminoguanidine ameliorates and L-arginine worsens brain
damage from intraluminal middle cerebral artery occlusion.
Stroke. 1996;27:317–323.
28.
Cockroft KM, Meistrell M, Zimmerman GA, Risucci
D, Bloom O, Cerami A, Tracey KJ. Cerebroprotective effects of
aminoguanidine in a rodent model of stroke. Stroke. 1996;27:1393–1398.
29.
Iadecola C, Zhang F, Xu X. Inhibition of
inducible nitric oxide synthase ameliorates cerebral ischemic
damage. Am J Physiol. 1995;268:R286–R292.
30. Nagayama M, Zhang F, Iadecola C. Delayed treatment with aminoguanidine decreases focal cerebral ischemic damage and enhances neurologic recovery in rats. J Cereb Blood Flow Metab. 1998;18:1107–1113.[Medline] [Order article via Infotrieve]
31.
Huang Z, Huang PL, Panahian N, Dalkara T, Fishman
MC, Moskowitz MA. Effects of cerebral ischemia in mice
deficient in neuronal nitric oxide synthase. Science. 1994;265:1883–1885.
32. Huang Z, Huang PL, Ma J, Meng W, Ayata C, Fishman MC, Moskowitz MA. Enlarged infarcts in endothelial nitric oxide synthase knockout mice are attenuated by nitro-L-arginine. J Cereb Blood Flow Metab. 1996;16:981–987.[Medline] [Order article via Infotrieve]
33.
Iadecola C, Zhang F, Casey R, Nagayama M, Ross
ME. Delayed reduction of ischemic brain injury and neurological
deficits in mice lacking the inducible nitric oxide synthase gene.
J Neurosci. 1997;17:9157–9164.
34. Zhang F, Xu S, Iadecola C. Time dependence of effect of nitric oxide synthase inhibition on cerebral ischemic damage. J Cereb Blood Flow Metab. 1995;15:595–601.[Medline] [Order article via Infotrieve]
35. Iadecola C, Xu X, Zhang F, el-Fakahany EE, Ross ME. Marked induction of calcium-independent nitric oxide synthase activity after focal cerebral ischemia. J Cereb Blood Flow Metab. 1995;15:52–59.[Medline] [Order article via Infotrieve]
36. Iadecola C, Zhang F, Xu S, Casey R, Ross ME. Inducible nitric oxide synthase gene expression in brain following cerebral ischemia. J Cereb Blood Flow Metab. 1995;15:378–384.[Medline] [Order article via Infotrieve]
37.
Iadecola C, Zhang F, Casey R, Clark HB, Ross ME.
Inducible nitric oxide synthase gene expression in vascular cells after
transient focal cerebral ischemia. Stroke. 1996;27:1373–1380.
38. Castillo J, Dávalos A, Naveiro J, Armengou A, Noya M. A reduction in L-arginine levels during acute stroke is associated with greater cerebral damage. Cerebrovasc Dis. 1997;7(suppl 4):82. Abstract.
39.
Dávalos A, Castillo J, Serena J, Noya M.
Duration of glutamate release after acute ischemic stroke.
Stroke. 1997;28:708–710.
40. Castillo J, Dávalos A, Lema M, Serena J, Noya M. Glutamate is a marker for cerebral ischemia in cortical but not deep infarcts. Cerebrovasc Dis. 1997;7:245–250.
41. Dávalos A, Castillo J. Progressing stroke: potential mechanisms of worsening. Cerebrovasc Dis. 1997;8(suppl 5):19–24.
42. Dávalos A, Castillo J. Progressing stroke. In: Fisher M, Bogousslavsky J, eds. Current Review of Cerebrovascular Disease. Boston, Mass: Butterworth-Heinemann; 1999:149–160.
43. Castillo J. Deteriorating stroke: diagnostic criteria, predictors, mechanisms and treatment. Cerebrovasc Dis. 1999 (suppl 3):1–8.
44.
Ashwal S, Tone B, Tian HR, Cole DJ, Pearce WJ.
Core and penumbral nitric oxide synthase activity during cerebral
ischemia and reperfusion. Stroke. 1998;29:1037–1046.
45. Beckman JS. Peroxynitrite versus hydroxyl radical: the role of nitric oxide in superoxide-dependent cerebral injury. Ann N Y Acad Sci. 1994;738:69–75.[Medline] [Order article via Infotrieve]
46. Malinski T, Bailey F, Zhang ZG, Chopp M. Nitric oxide measured by a porphyrinic microsensor in rat brain after transient middle cerebral artery occlusion. J Cereb Blood Flow Metab. 1993;13:355–358.[Medline] [Order article via Infotrieve]
47. Fukuyama N, Takizawa S, Ishida H, Hoshiai K, Shinohara Y, Nakazawa H. Peroxynitrite formation in focal cerebral ischemia-reperfusion in rats occurs predominantly in the peri-infarct region. J Cereb Blood Flow Metab. 1998;18:123–129.[Medline] [Order article via Infotrieve]
48. Kuprinski J, Vodorotz Y, Li C, Slowik A, Beevers D, Flanders KC, Lip G, Kumar P, Szczudlik A. Inducible nitric oxide production and expression of transforming growth factor-beta 1 in serum and CSF after cerebral ischaemic stroke in man. Nitric Oxide. 1998;2:442–453.[Medline] [Order article via Infotrieve]
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