Transcobalamin 2 variant associated with poststroke homocysteine modifies recurrent stroke risk.

OBJECTIVES: The Vitamin Intervention for Stroke Prevention trial found an association between baseline poststroke homocysteine (Hcy) and recurrent stroke. We investigated genes for enzymes and cofactors in the Hcy metabolic pathway for association with Hcy and determined whether associated single nucleotide polymorphisms (SNPs) influenced recurrent stroke risk. METHODS: Eighty-six SNPs in 9 candidate genes (BHMT1, BHMT2, CBS, CTH, MTHFR, MTR, MTRR, TCN1, and TCN2) were genotyped in 2,206 subjects (83% European American). Associations with Hcy measures were assessed using linear regression models assuming an additive genetic model, adjusting for age, sex, and race and additionally for baseline Hcy when postmethionine load change was assessed. Associations with recurrent stroke were evaluated using survival analyses. RESULTS: Five SNPs in the transcobalamin 2 (TCN2) gene were associated with baseline Hcy (false discovery rate [FDR]-adjusted p = 0.049). TCN2 SNP rs731991 was associated with recurrent stroke risk in the low-dose arm of the trial under a recessive model (log-rank test p = 0.009, hazard ratio 0.34). Associations with change in postmethionine load Hcy levels were found with 5 SNPs in the cystathionine ß-synthase (CBS) gene (FDR-adjusted p < 0.031). CONCLUSIONS: TCN2 variants contribute to poststroke Hcy levels, whereas variants in the CBS gene influence Hcy metabolism. Variation in the TCN2 gene also affects recurrent stroke risk in response to cofactor therapy.

Theoretical simulation of oxygen transport to brain by networks of microvessels: effects of oxygen supply and demand on tissue hypoxia.

OBJECTIVE: Simulations of oxygen delivery by a three-dimensional network of microvessels in rat cerebral cortex were used to examine how the distribution of partial pressure of oxygen (PO2) in tissue depends on blood flow and oxygen consumption rates. METHODS: Network geometry was deduced from previously published scanning electron micrographs of corrosion casts. A nonlinear least-squares method, using images obtained at three different angles, was used to estimate vessel locations. The network consisted of 50 segments in a region 140 microm x 150 microm x 160 microm. A Green's function method was used to predict the PO2 distribution. Effects of varying perfusion and consumption were examined, relative to a control state with consumption 10 cm3O2/100 g per min and perfusion 160 cm3/100 g per min. RESULTS: In the control state, minimum tissue PO2, was 7 mm Hg. A Krogh-type model with the same density of vessels, but with uniform spacing, predicted a minimum tissue PO2 of 23 mm Hg. For perfusion below 60% of control, tissue hypoxia (PO2 <1 mm Hg) was predicted. When perfusion was reduced by 75%, the resulting hypoxia could be eliminated by a 31% reduction in oxygen consumption rate. CONCLUSIONS: The simulations suggest that tissue hypoxia resulting from a severe decrease in brain perfusion, as can occur in stroke, may be avoided by a moderate decrease in oxygen consumption rate.

Leukocyte accumulation and hemodynamic changes in the cerebral microcirculation during early reperfusion after stroke.

BACKGROUND AND PURPOSE: Leukocytes contribute to cerebral ischemia-reperfusion injury. However, few experimental models examine both in vivo behavior of leukocytes and microvascular rheology after stroke. The purpose of the present study was to characterize patterns of leukocyte accumulation in the cerebral microcirculation and to examine the relationship between leukocyte accumulation and microcirculatory hemodynamics after middle cerebral artery occlusion and reperfusion (MCAO-R). METHODS: Male rats (250 to 350 g) were anesthetized and ventilated. Tail catheters were inserted for measurement of arterial blood gases and administration of drugs. Body temperature was maintained at 37 degrees C. Animals were subjected to 2 hours of MCAO by the filament method. A cranial-window preparation was performed, and the brain was superfused with warm, aerated artificial cerebrospinal fluid. Reperfusion was initiated by withdrawing the filament, and the pial microcirculation was observed by use of intravital fluorescence microscopy. Leukocyte accumulation in venules, arterioles, and capillaries; leukocyte rolling in venules; and leukocyte venular shear rate were assessed during 1 hour of reperfusion. RESULTS: We found significant leukocyte adhesion in cerebral venules during 1 hour of reperfusion after 2 hours of MCAO. Leukocyte trapping in capillaries and adhesion to arterioles after MCAO-R tended to increase compared with controls, but the increase was not significant. We also found that shear rate was significantly reduced in venules during early reperfusion after MCAO. CONCLUSIONS: A model using the filament method of stroke and fluorescence microscopy was used to examine white-cell behavior and hemodynamics in the cerebral microcirculation after MCAO-R. We observed a significant increase in leukocyte rolling and adhesion in venules and a significant decrease in blood shear rate in the microcirculation of the brain during early reperfusion. Leukocytes may activate and damage the blood vessels and surrounding brain cells, which contributes to an exaggerated inflammatory component to reperfusion. The model described can be used to examine precisely blood cell-endothelium interactions and hemodynamic changes in the microcirculation during postischemic reperfusion. Information from these and similar experiments may contribute to our understanding of the early inflammatory response in the brain during reperfusion after stroke.

Role of a common mutation in the homocysteine regulatory enzyme methylenetetrahydrofolate reductase in ischemic stroke.

A common mutation in methylenetetrahydrofolate reductase (MTHFR), a homocysteine metabolic pathway enzyme, has been associated with increased homocysteine levels and increased risk for premature cardiovascular disease. The purpose of this study was to assess the association between the prevalence of the MTHFR mutation, hyperhomocysteinemia, and subtypes of ischemic stroke in an elderly population comprised of three age-balanced groups of patients. The presence of the C677T MTHFR mutation was determined by a direct polymerase chain reaction-based assay performed on blood samples from 136 patients with acute ischemic stroke, 95 patients with atherosclerotic risk factors for stroke (including some with a history of previous stroke or transient ischemic attack), and 52 healthy control subjects. The prevalence of the homozygous C677T mutation was not significantly higher in the elderly stroke patients (7%) than in the atherosclerotic risk (8%) or healthy elderly control (2%) groups. Plasma homocysteine levels were higher in the acute stroke patient group (14.5+/-4.5 micromol/L) and atherosclerotic risk patient group (14.6+/-6.2 micromol/L) compared with the control subjects (10.3+/-3.1 micromol/ L, P < 0.03). Homozygotes for the C677T MTHFR mutation did not have significantly higher homocysteine levels than non-homozygotes. Moderate hyperhomocysteinemia, though common in older patients with ischemic cerebrovascular disease, is not attributable, at least in this patient group, to a higher prevalence of the C677T MTHFR mutation.

Microalbuminuria in ischemic stroke.

OBJECTIVES: To determine (1) the incidence of microalbuminuria in patients with recent ischemic stroke, (2) its relationship to risk factors for stroke, (3) its prevalence in the major subtypes of ischemic stroke, and (4) its potential for identifying patients at increased risk for recurrent stroke, myocardial infarction, or vascular death. DESIGN: Prospective case-control study. SETTING: Outpatient clinics at the medical centers affiliated with the Department of Veterans Affairs and Oregon Health Sciences University in Portland, Ore. PATIENTS: A total of 186 older men and women (median age, 65 years) who were enrolled in a prospective study of risk factors for recurrent stroke, including 97 patients with recent (6-8 weeks) ischemic stroke, 51 with similar clinical risk factors for stroke, including 24 with a history of remote stroke or transient ischemic attack, and 38 community-dwelling volunteers. RESULTS: Microalbuminuria was 3 times more prevalent in patients with recent stroke (29%) than in those with clinical risk factors for stroke (10%), and was undetectable in healthy elderly controls (P<.001). The presence of microalbuminuria in recent stroke as well as in the combined recent and remote stroke or transient ischemic attack group (n = 121) was predicted by diabetes (odds ratio [OR], 8.4; 95% confidence interval [CI], 2.6-27.0; P<.001; serum albumin levels (OR, 0.12; 95% CI, 0.03-0.50; P<.005); age (OR, 1.1; 95% CI, 1.0-1.2; P<.01), and ischemic heart disease (OR, 3.0; 95% CI, 1.0-9.1; P<.05). Among patients with recent stroke the prevalence of microalbuminuria did not differ among major ischemic stroke subtypes, ie, atheroembolic, 23%; cardioembolic, 30%; and lacunar, 33%. During a mean +/- SD of 1.5 +/- 0.9 years of follow-up, 20% of patients with recent stroke, 14% with risk factors for stroke, and 0% of healthy elderly volunteers had vascular end points (P<.004), with events being as frequent in patients with microalbuminuria (32%) as in patients with macroalbuminuria (33%). After controlling for major clinical risk factors, microalbuminuria remained an independently significant predictor of future stroke in the combined recent stroke and remote stroke or transient ischemic attack group (Cox proportional hazard ratio, 4.9; 95% CI, 1.4-17.6; P<.01). CONCLUSIONS: Microalbuminuria is a common finding in patients with cerebrovascular disease and is associated with increased risk for stroke even after correction for the presence of confounding clinical risk factors. These data suggest that microalbuminuria merits further examination as a potentially inexpensive and easily measured marker of increased risk for stroke.

Cognitive impairment and intracranial calcification in chronic hypoparathyroidism.

BACKGROUND: Intracranial calcification is associated with chronic hypoparathyroidism. The relationship between intracranial calcification, neurological abnormalities and cognitive deficits in this disorder is unknown. The purpose of this study was to determine whether chronic hypoparathyroidism is associated with cognitive impairment. METHODS: We studied 11 hypoparathyroid patients and compared them with a sex-, age-, and education-matched control group. The hypoparathyroidism was postsurgical in nine and idiopathic in two. All patients underwent nonenhanced head computed tomography, detailed neurological examinations, and a battery of cognitive tests. These tests were performed separately and individual examiners were blinded to the results of the other components of the study. RESULTS: The mean age of the patients was 55 years; the duration of hypoparathyroidism was at least 9 years. Neuropsychological testing revealed cognitive impairment in 65% of hypoparathyroid subjects, and the presence of significant differences between the hypoparathyroid and control groups. Computed tomography showed intracranial calcification in 6 of 10 hypoparathyroid subjects tested, and neurological (motor) examination revealed 5 of 11 with abnormal findings. There were positive correlations between the presence of cognitive deficits and cerebral calcification (r = 0.59, P = 0.07), between abnormal motor findings and cerebral calcification (r = 0.77, P < 0.01) and between abnormal motor findings and the degree of cognitive deficit (r = 0.83, P < 0.01). CONCLUSIONS: We conclude that cognitive and neurological deficits commonly occur in patients with chronic hypoparathyroidism and may be pathophysiologically related to the presence of intracranial calcification.

Persistent inflammatory response in stroke survivors.

OBJECTIVE: Goals were to determine how long acute-phase markers remain elevated after ischemic stroke and how marker levels relate to stroke risk factors, stroke mechanism, and subsequent vascular events. METHODS: Fibrinogen (FIB), C-reactive protein (CRP), leukocytes (WBC), neutrophils (PMN), interleukin-6, and interleukin-1 receptor antagonist were measured at stroke onset and at 6 weeks, 6 months, and 1 year after enrollment, or until a vascular event occurred in 136 acute ischemic stroke patients, 76 patients with comparable risk factors for stroke, and 48 age-balanced healthy subjects. RESULTS: Multivariate logistic analysis showed that prior stroke and FIB level predicted new events in stroke patients (p < 0.04 for both), whereas congestive heart failure (p < 0.02) and creatinine level (p < 0.006) were predictive in at-risk patients. After controlling for infection, FIB, CRP, and PMN levels at baseline were higher in at-risk but not in stroke patients with recurrent events (p < 0.05 for all). At 1 year, FIB levels remained elevated in event-free stroke survivors compared with levels in the risk and control groups (p < 0.001 for both). FIB also remained higher in stroke survivors who had atheroembolism (AE) compared with non-AE stroke survivors (381+/-72 versus 342+/-78 mg/dL, p < 0.02). Peripheral vascular disease was an independent predictor (p < 0.0001) of longitudinal FIB in stroke survivors. Of note, both WBC and PMN levels were chronically elevated in patients with stroke risk factors and in stroke survivors (p < 0.0001 for both) compared with healthy elderly subjects. CONCLUSIONS: Most acute-phase markers decline gradually after stroke, but FIB remains significantly elevated and is associated with increased risk for recurrent vascular events.

The initial acute phase response predicts long-term stroke recovery.

Indicators of an acute phase response (APR) in acute ischemic stroke have been shown to correlate with infarct size and predict stroke recurrence. In this study, we examined how well the APR indicators predicted long-term stroke recovery compared with standard clinical predictors of recovery. Plasma levels of interleukin-6 (IL-6), fibrinogen, white blood cells (WBCs), and serum albumin were measured within 4+/-2 days of onset in 131 stroke patients who were free of apparent infections. Standard clinical predictors included initial National Institutes of Health Stroke Scale (NIHSS), infarct size on computed tomography (CT), and Glasgow scale. The individual correlations with 6-month Glasgow outcome were IL-6, 0.42; fibrinogen, 0.24; WBC, 0.35; albumin, 0.47; NIHSS, 0.53; infarct size, 0.19; and initial Glasgow, 0.57. (all P<.005). Multiple regression analysis yielded an adjusted R(2) of .31 for the APR indicators, compared with .38 for the clinical variables. These results indicate that the initial APR is highly correlated with 6-month stroke recovery and that this correlation approaches that observed with standard clinical predictors.

Randomized trial of Cervene, a kappa receptor-selective opioid antagonist, in acute ischemic stroke.

The purpose of this randomized trial was to confirm drug safety and to obtain preliminary efficacy data on Cervene (nalmefene), an opioid antagonist with relative kappa receptor selectivity, for the treatment of acute ischemic stroke. Patients were treated for 24 hours with either intravenous Cervene (0.05 mg/kg as an initial infusion over 15 minutes and 0.01 mg/kg/h maintenance) or placebo within 6 hours of an ischemic stroke. Efficacy was assessed by comparing the change from baseline to day 7 in the National Institutes of Health stroke scale score (NIHSSS) and the Glasgow Outcome Scale and Barthel Index at 3 months. Forty-four evaluable patients were randomized (3:1) to Cervene (n = 34; treated at 5.0 +/- 0.9 hours after onset) and placebo (n = 10; treated at 4.6 +/- 1.5 hours). No deaths or serious adverse events reasonably attributable to Cervene have been reported. A "major improvement" (NHSSS > 4) was seen at day 7: placebo, 33% (three of nine patients) and Cervene, 66% (19 of 29 patients). Only patients with initial NIHSSS >/= 4 were considered evaluable for this primary endpoint. "Good recovery" at 3 months (Glasgow = 5) was as follows: placebo, 50% (5 of 10 patients) and Cervene, 73% (24 of 33 patients). The death rate at 3 months was placebo, 20% (2 of 10 patients) and Cervene, 9.1% (3 of 33 patients). One patient was lost to follow-up. In conclusion, results from this randomized trial suggest that Cervene is safe, tolerable, and may be beneficial in the treatment of acute stroke patients.

Ischemic stroke in the elderly. Role of the common factor V mutation causing resistance to activated protein C.

BACKGROUND AND PURPOSE: A common missense mutation in coagulation factor V (Arg 506 Gln) creates phenotypic resistance to the anticoagulant effects of activated protein C and predisposes carriers to venous thrombosis. To assess a correlation between this common hypercoagulable state and ischemic cerebrovascular disease, we have compared the prevalence of this mutation in a group of stroke patients with that in several control patient groups. METHODS: The presence of the factor V Arg 506 Gln mutation was determined by a direct polymerase chain reaction-based assay on peripheral blood leukocytes from 161 elderly patients with acute ischemic stroke, 116 elderly patients with stroke risk factors but without acute stroke, 54 healthy elderly control subjects, and 287 younger control individuals (197 blood donors and 90 neonates). RESULTS: The prevalence of the heterozygous Arg 506 Gln factor V mutation was not significantly different in the elderly stroke patients (2.5%) compared with either of the age-matched control groups (2% to 4%). The prevalence of this mutation was significantly higher in each of two younger control groups (approximately 8%) than in the elderly stroke patients (2.5%). CONCLUSIONS: The common factor V Arg 506 Gln mutation predisposing to venous thrombosis is not a significant genetic risk factor for ischemic stroke in the elderly.

Lazaroids. CNS pharmacology and current research.

The 21-aminosteroids (lazaroids) are inhibitors of lipid membrane peroxidation and appear to function as oxygen free radical scavengers. The therapeutic potential of the lazaroid tirilazad mesylate has been extensively studied in several CNS disorders. Tirilazad and related compounds have been found to be highly beneficial in spinal cord trauma. Spinal cord injury studies utilising tirilazad are currently underway to determine the optimal combination of medications. Tirilazad has also been found to be beneficial in experimental head injury models, however current clinical studies have failed to confirm this efficacy, due in part to difficulties in obtaining therapeutic drug concentrations. Clinical studies using tirilazad in subarachnoid haemorrhage have been more promising. It has been shown to be beneficial in terms of reducing vasospasm and cerebral infarction associated with subarachnoid haemorrhage, and has now been approved in several European countries in this indication. Results from US studies are expected shortly. Finally, tirilazad has also been extensively tested in a variety of stroke models. Although it appears to be highly beneficial in experimental models, the clinical studies to date have failed to confirm this efficacy. Again, this failure appears to be due largely to inadequate drug concentrations having so far been tested.

Time course of ICAM-1 expression and leukocyte subset infiltration in rat forebrain ischemia.

The time course of ICAM-1 expression and leukocyte subset infiltration was studied in a model of CNS reperfusion injury in adult rats. Leukocyte adhesion and infiltration, mediated in part by intercellular adhesion molecule-1 (ICAM-1), appears to potentiate CNS reperfusion injury. The timing and relationship between ICAM-1 staining and leukocyte infiltration postglobal CNS ischemia is unknown. Reversible forebrain ischemia was produced in 32 adult Sprague-Dawley rats using the two-vessel occlusion model with histologic analysis performed at specific intervals postischemia: 1, 3, 6, 12, and 24 h, 4 and 7 d, or sham-operated controls (n = 4 each group). Monoclonal antibodies against ICAM-1 (1A29 and TM8), a specific granulocyte (PMN) (HIS48), and a specific monocyte/macrophage (M phi) (ED1) were used. No specific leukocyte and only rare ICAM-1 vessel immunoreactivity was observed in sham controls. ICAM-1: Significant expression in microvessels beginning at 1 h with additional diffuse CA1 pyramidal layer staining beginning at 4 d. Leukocytes: No PMN cells and rare M phi identified at 6 and 12 h. By 24 h: moderate infiltrate in areas of ICAM-1 expression of PMN and M phi. At 4 and 7 d: only M phi accumulation, cellular morphology now similar to microglia. The results of this study indicate that early and persistent ICAM-1 expression occurs following CNS ischemia with associated leukocyte infiltration.

Safety and efficacy of percutaneous transluminal angioplasty for intracranial atherosclerotic stenosis.

BACKGROUND AND PURPOSE: Percutaneous transluminal angioplasty (PCTA) is increasingly used to treat extracerebral arterial stenosis. The present study evaluates the safety and efficacy of PCTA treatment of symptomatic intracranial atherosclerotic stenosis. METHODS: A series of 22 vessels in 17 patients were treated with PCTA. All patients had recurrent neurological symptoms referable to the stenotic vessel despite optimal medical therapy. Critical (> 70%) arterial stenosis was confirmed by angiogram, and angioplasty was performed with a 3.0- to 3.5-mm Stealth balloon. RESULTS: The average preangioplasty stenosis (North American Symptomatic Carotid Endarterectomy Trial criteria) was 72 +/- 8% (mean +/- SD), with a significant improvement seen after angioplasty; the best angiographic stenosis (after healing of intimal injury, if any) was 43 +/- 24% (P < .001). Overall PCTA was successful in 82% of the vessels. There were two strokes during angioplasty for a 30-day morbidity rate of 9.1% per treated vessel and 11.7% per case. The other 15 patients were clinically evaluated at 3 and 6 months; all cases were without further events. Restenosis was evaluated in 8 patients (12 vessels) with an angiogram at 6 months showing further improvement compared with the initial post-PCTA stenosis (51 +/- 10% versus 37 +/- 21% [P = .05]). CONCLUSIONS: PCTA may be a beneficial therapy in selected cases of symptomatic intracranial atherosclerotic stenosis. Further study using a randomized trial is needed.

Interleukin-6 and interleukin-1 receptor antagonist in acute stroke.

Elevated plasma levels of interleukin-6 (IL-6), a key regulator of the acute phase response that includes increased fibrinogen synthesis, have recently been detected in patients with acute stroke. Nevertheless, the role of the acute phase response in stroke has been controversial, with some studies suggesting that preexisting infection accounts for most of the acute phase response. Increased IL-6 could signal the involvement of antiinflammatory activity, since IL-6 stimulates the production of endogenous antiinflammatory mediators such as interleukin-1 receptor antagonist (IL-1RA). To better understand the interaction of pro- and antiinflammatory acute phase processes in brain infarction, plasma levels of IL-1RA, IL-6, and acute phase proteins including fibrinogen and c-reactive protein (CRP) were measured within 4 +/- 2 days of onset in 50 patients with acute ischemic stroke and in 20 age-matched healthy controls. After excluding patients with evidence of infection, both IL-1RA and IL-6 were significantly elevated in stoke patients compared with controls (p < 0.0001). IL-1RA and IL-6 were both significantly correlated with levels of CRP, p < 0.05 and p < 0.001, respectively, but not with each other. Levels of IL-6 and IL-1RA, together with fibrinogen and CRP were higher in patients with infarcts of greater than 3 cm and lowest in patients with lacunar syndromes.(ABSTRACT TRUNCATED AT 250 WORDS)

The influence of antiadhesion therapies on leukocyte subset accumulation in central nervous system ischemia in rats.

Although treatment with agents that block leukocyte function, including anti-ICAM-1 and doxycycline, reduces experimental central nervous system (CNS) ischemic injury, it is not known how leukocyte subset accumulation is affected by these agents. Using the rat two-vessel occlusion model and immunohistochemistry, we investigated granulocyte (PMN) and monocyte/macrophage (M phi) accumulation at 1 and 4 d postischemia. A total of 24 animals were randomized to sham surgery, or to ischemia with saline, anti-ICAM-1, or doxycycline treatments. No leukocytes were observed in sham animals. At 24 h postischemia, there was a moderate infiltration of PMN and M phi in untreated animals that was significantly decreased with either treatment. At 4 d after ischemia no PMN were identified, with extensive M phi accumulation occurring in untreated animals that was only partially reduced with doxycycline treatment. These results confirm that both anti-ICAM-1 and doxycycline treatments reduce PMN and M phi infiltration at 24 h. Delayed M phi accumulation occurs despite treatment, suggesting that some of these cells represent transformed resident microglia.