ORIGINAL CONTRIBUTION

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1 ORIGINAL CONTRIBUTION Soluble Vascular Cell Adhesion Molecule 1 and N-terminal Pro B-Type Natriuretic Peptide in Predicting Ischemic Stroke in Patients With Cerebrovascular Disease Duncan J. Campbell, MD, PhD; Mark Woodward, PhD; John P. Chalmers, MD, PhD; Samuel A. Colman, MBios; Alicia J. Jenkins, MD; Bruce E. Kemp, PhD; Bruce C. Neal, MD, PhD; Anushka Patel, MD; Stephen W. MacMahon, PhD Background: Patients with stroke or transient ischemic attack are at high risk of another stroke, and there is need for improved strategies to predict recurrent stroke. Objective: To assess the prognostic value of levels of soluble vascular cell adhesion molecule 1 (svcam-1), N-terminal pro B-type natriuretic peptide (NT-proBNP), C-reactive protein, homocysteine, renin, and lipids and lipoprotein particle concentration and size in patients with previous stroke or transient ischemic attack. Design, Setting, and Participants: A nested casecontrol study of participants of the Perindopril Protection Against Recurrent Stroke Study was performed. The Perindopril Protection Against Recurrent Stroke Study was a placebo-controlled trial of a perindopril erbumine based, blood pressure lowering regimen that reduced ischemic stroke risk by 24% among individuals with previous stroke or transient ischemic attack. Each of 252 patients who experienced ischemic stroke during a mean follow-up of 3.9 years was matched to 1 to 3 control patients. Matching variables were age, sex, treatment allocated, region, and most recent qualifying event at randomization. Main Outcome Measures: Risk of ischemic stroke predicted by baseline levels of svcam-1, NT-proBNP, C-reactive protein, homocysteine, renin, and lipids and lipoprotein particle concentration and size. Results: Levels of svcam-1 and NT-proBNP predicted recurrent ischemic stroke. The odds ratio for patients in the highest, as compared with the lowest, quarter was 2.24 (95% confidence interval, ) for svcam-1 level and 1.62 (95% confidence interval, ) for NT-proBNP level, after adjustment for matching and other risk factors. Patients in the highest quarters for both svcam-1 and NT-proBNP levels had 3.6 times the risk of recurrent ischemic stroke compared with patients in the lowest quarters for both biologic markers. Level of svcam-1 was similarly predictive of ischemic stroke in patients allocated to placebo and perindopril-based therapy. Baseline plasma levels of C-reactive protein, homocysteine, renin, and lipids and lipoprotein particle concentration and size did not predict recurrent ischemic stroke risk. Conclusion: Measurement of svcam-1 and NT-proBNP levels provides prognostic information for recurrent ischemic stroke beyond traditional risk factors. Arch Neurol. 2006;63:60-65 Author Affiliations: St Vincent s Institute of Medical Research, Fitzroy, Australia (Drs Campbell and Kemp); Department of Medicine, University of Melbourne, St Vincent s Hospital, Fitzroy (Drs Campbell, Jenkins, and Kemp); CSIRO Health Sciences and Nutrition, Parkville, Australia (Dr Kemp); The George Institute for International Health, University of Sydney, Camperdown, Australia (Drs Woodward, Chalmers, Neal, Patel, and MacMahon, and Mr Colman). STROKE IS A LEADING CAUSE OF death and disablement, with a risk of 21% for those living beyond 55 years of age. 1 For many individuals, their initial stroke is the first indication of their stroke risk. Among those who survive a stroke or transient ischemic attack (TIA), as many as one third have another stroke within 5 years of the index event. 2-4 Recurrent strokes are associated with higher risk and greater degree of disability than initial strokes, 5,6 and prevention of subsequent stroke in patients with cerebrovascular disease is an important objective of treatment. The objective of this study was to evaluate the prognostic value of a range of potential risk factors for recurrent ischemic stroke by conducting a nested casecontrol study of participants in the Perindopril Protection Against Recurrent Stroke Study (PROGRESS). A multicenter, randomized, double-blind, placebocontrolled study, PROGRESS was designed to determine the effects of active therapy with a perindopril erbumine based, blood pressure lowering regimen on the risk of stroke and other major vascular events among individuals with a stroke or TIA within the previous 5 years. 3 This regimen substantially reduced the risk of total stroke by 28% and ischemic stroke by 24%. 3 The potential risk factors we examined were markers of endothelial dysfunction and inflammation (soluble vas- 60

2 cular cell adhesion molecule 1 [svcam-1] and C-reactive protein [CRP] levels), cardiac dysfunction (N-terminal pro B-type natriuretic peptide [NT-proBNP] level), oxidative stress (homocysteine level), renin-angiotensin system activity (active renin level), and dyslipidemia (lipids levels and lipoprotein particle concentration and size). We also examined whether any of these potential risk factors predicted ischemic stroke in subjects receiving perindopril-based therapy. METHODS STUDY POPULATION The design and major outcomes of PROGRESS are described in detail elsewhere. 3 Briefly, 6105 participants were recruited from 172 collaborating centers in 10 countries from Australasia, Europe, and Asia. Participants were randomized to either placebo (n=3054) or active therapy (n=3051), consisting of a flexible regimen based on the angiotensin-converting enzyme inhibitor perindopril erbumine (4 mg/d) with the addition of the diuretic indapamide at the discretion of treating physicians. The institutional ethics committee of each collaborating center approved the trial, and all participants provided written, informed consent to participate in the study, including the collection of blood samples for laboratory tests and the measurement of factors that may be linked to an increased risk of stroke. Congestive heart failure was a study exclusion criterion. 7 LABORATORY METHODS Collection of blood samples, storage of plasma, and measurement of lipids, renin, NT-proBNP, CRP, and creatinine levels are described elsewhere. 8 Lipoprotein particle concentration and size were measured on first-thaw EDTA plasma by nuclear magnetic resonance by LipoScience, Inc, Raleigh, NC. Homocysteine level was measured by high-performance liquid chromatography with fluorimetric detection, 9,10 and svcam-1 level was measured by immunoassay using reagents from R&D Systems, Inc, Minneapolis, Minn, on second-thaw EDTA plasma. MAIN OUTCOME MEASURES Stroke, either fatal or nonfatal, was defined as an acute disturbance of focal neurological function with symptoms lasting more than 24 hours or resulting in earlier death. 11 Diagnosis of ischemic stroke required a computed tomographic scan (performed within 3 weeks) that either had normal results or showed infarction in the clinically expected area, or autopsy evidence of cerebral infarction. The adjudication committee reviewed source documentation for all deaths and every potential stroke and myocardial infarction recorded during the study follow-up period. STATISTICAL ANALYSIS To minimize the influence of the qualifying event on the biochemical parameters, and also changes in biochemical parameters following the collection of blood before freezing of plasma, the base population for this study was 2673 patients who were enrolled more than 1 month after their most recent qualifying event and whose plasma was frozen at 80 C within 24 hours of blood collection. Each of 252 cases of ischemic stroke during the mean 3.9 years of follow-up was matched with 1 to 3 controls randomly sampled from patients who were alive and did not have a stroke between randomization and the time of case ascertainment; anyone who became a case after the onset of ischemic stroke in the index case was eligible for selection as a match, and controls could be matched for more than 1 case. 12 Matching variables were age (within 5 years), sex, treatment allocated (perindopril-based or placebo, mono or dual therapy), region, and most recent qualifying event at randomization. Baseline variables were compared between cases and controls using 2, parametric, or nonparametric tests, as appropriate. 12 The case-control data were analyzed using 2 different conditional logistic regression models to obtain odds ratios (ORs) for svcam-1, NT-proBNP, and CRP levels, according to equal quarters of the distribution of each plasma marker in the total sample (cases and controls). 12 Model 1 was unadjusted, except for the matching variables. Model 2 was adjusted through matching and also for all baseline predictors and potential predictors of ischemic stroke. These were baseline systolic blood pressure; total cholesterol level; a history of current smoking, diabetes mellitus, and peripheral arterial disease; and antihypertensive medication other than -blockers, calcium channel blockers, or diuretics. In addition, model 2 for svcam-1 level was adjusted for NT-proBNP and CRP levels, model 2 for NT-proBNP level was adjusted for svcam-1 and CRP levels, and model 2 for CRP level was adjusted for svcam-1 and NT-proBNP levels. All P values were 2 tailed, and values.05 were considered to indicate statistical significance. All analyses were performed using SAS version 8.2 (SAS Institute Inc, Cary, NC). RESULTS BASELINE CHARACTERISTICS The baseline clinical and biochemical characteristics of the 252 ischemic stroke cases and 544 controls without ischemic stroke are given in Table 1 and Table 2. More than 90% of cases and controls had previous ischemic stroke and/or TIA. In comparison with controls without ischemic stroke during the period of observation, ischemic stroke cases were more likely to have hypertension on enrollment; to have a history of current smoking, diabetes mellitus, and peripheral arterial disease; and to be taking antihypertensive medication other than -blockers, calcium channel blockers, or diuretics (Table 1). In addition, ischemic stroke cases had higher baseline levels of svcam-1, NT-proBNP, and CRP. There were no differences between cases and controls with respect to levels of plasma lipids, creatinine, homocysteine, and renin and lipoprotein particle concentration and size (Table 2). MAIN OUTCOME MEASURES The OR for ischemic stroke increased with increasing levels of baseline svcam-1, NT-proBNP, and CRP (Table 3) but for none of the other variables listed in Table 2. The OR for patients in the highest, as compared with the lowest, quarter was 2.34 (95% confidence interval [CI], ) for svcam-1 level and 1.90 (95% CI, ) for NT-proBNP level. Whereas the OR was significantly elevated for only patients in the highest quarter of NT-proBNP level, it was similarly elevated for patients in the second, third, and fourth quarters of svcam-1 level. 61

3 Table 1. Baseline Clinical Characteristics of Patients Who Developed Ischemic Stroke During 3.9 Years of Follow-Up and Control Patients Who Did Not Develop Ischemic Stroke Characteristic Cases (n = 252) Controls Without Ischemic Stroke (n = 544) P Value Age, y, mean (SD)* 67 (8) 66 (8).55 Male, No. (%)* 183 (73) 396 (73).96 Asian, No. (%)* 103 (41) 236 (43).51 Randomization to perindopril 107 (42) 241 (44).63 erbumine based therapy, No. (%)* Qualifying event, No. (%) Ischemic stroke 206 (82) 433 (80).48 Cerebral hemorrhage 13 (5) 30 (6).84 Unknown stroke 7 (3) 12 (2).62 TIA or amaurosis fugax 53 (21) 111 (20).84 Time since qualifying event, 8 (3-18) 9 (3-22).13 mo, Other vascular disease history, No. (%) Systolic blood pressure, 150 (19) 148 (18).10 mm Hg, mean (SD) Diastolic blood pressure, 85 (11) 85 (10).63 mm Hg, mean (SD) Hypertension 136 (54) 244 (45).02 Antihypertensive therapy 128 (51) 248 (46).17 History of coronary heart disease 39 (15) 83 (15).94 History of atrial fibrillation 32 (13) 51 (9).15 Valvular heart disease 7 (3) 9 (2).29 Left ventricular hypertrophy on ECG 21 (8) 30 (6).13 Peripheral arterial disease 18 (7) 21 (4).05 Other medical history, No. (%) Current smoker 65 (26) 106 (19).04 History of diabetes mellitus 46 (18) 71 (13).05 Body mass index, mean (SD) 25.7 (3.7) 25.4 (3.7).36 Current medications, No. (%) -Blockers 42 (17) 86 (16).76 Calcium channel blockers 104 (41) 220 (40).83 Diuretics 29 (12) 70 (13).59 Other antihypertensives 37 (15) 44 (8).004 Antiplatelet agents 199 (79) 427 (78).88 Statins 22 (9) 39 (7).44 Oral anticoagulants 25 (10) 54 (10) 7.99 Abbreviations: ECG, electrocardiogram; IQR, interquartile range; TIA, transient ischemic attack. *Controls were matched for these criteria. Participants recruited from People s Republic of China or Japan. Subjects could have more than 1 qualifying event. Baseline systolic blood pressure 160 mm Hg or higher or diastolic blood pressure 90 mm Hg or higher. History of myocardial infarction or coronary revascularization or angina (supported by documented electrocardiographic or angiographic evidence of coronary disease). Calculated as weight in kilograms divided by the square of height in meters. The OR of 1.32 (95% CI, ) for CRP level did not achieve statistical significance (Table 3, CRP level model 1). In multiple-variable analysis (Table 3, model 2), the OR for patients in the highest, as compared with the lowest, quarter was 2.24 (95% CI, ) for svcam-1 level, borderline nonsignificant at 1.62 (95% CI, ) (P=.06) for NT-proBNP level, and nonsignificant for CRP level at 1.00 (95% CI, ). However, the OR for NT-proBNP level increased to 1.79 (95% Table 2. Baseline Biochemical Characteristics of Patients Who Developed Ischemic Stroke During 3.9 Years of Follow-Up and Control Patients Who Did Not Develop Ischemic Stroke Characteristic Cases (n = 252) Controls Without Ischemic Stroke (n = 544) P Value Plasma lipids levels, mean (SD) Total cholesterol, mg/dl 216 (39) 212 (42).61 HDL cholesterol, mg/dl 51 (15) 51 (15).76 LDL cholesterol, mg/dl 126 (32) 124 (36).52 Triglycerides, mg/dl, 177 ( ) 177 ( ).56 Creatinine level, mg/dl, 1.04 (0.24) 1.02 (0.26).32 mean (SD) NT-proBNP level, pmol/l, 16.3 ( ) 12.9 ( ).003 CRP level, mg/l, 2.1 ( ) 1.6 ( ).02 Homocysteine level, mg/l, 2.2 ( ) 2.2 ( ).99 Renin level, pg/ml, 13 (7-22) 13 (7-24).44 svcam-1 level, ng/ml, 560 ( ) 535 ( ).01 NMR, VLDL particles, 81 (60-100) 80 (60-102).98 LDL particles, 1478 ( ) 1474 ( ).99 HDL particles, µmol/l 32 (28-36) 32 (28-37).38 Large VLDL particles, 2.6 ( ) 2.5 ( ).81 Medium VLDL particles, 47 (33-64) 47 (32-65).97 Small VLDL particles, 23 (12-38) 23 (12-36).95 IDL particles, 5 (0-32) 8 (0-35).30 Large LDL particles, 347 ( ) 349 ( ).60 Medium LDL particles, 558 ( ) 520 ( ).35 Small LDL particles, 196 (0-754) 233 (0-723).38 Large HDL particles, 6.2 ( ) 6.2 ( ).66 µmol/l Medium HDL particles, 3.1 ( ) 3.3 ( ).55 µmol/l Small HDL particles, 21.1 ( ) 21.2 ( ).69 µmol/l Mean VLDL diameter, nm 49 (44-58) 49 (44-59).84 Mean LDL diameter, nm 20.8 ( ) 20.8 ( ).48 Mean HDL diameter, nm 9.1 ( ) 9.0 ( ).78 Abbreviations: CRP, C-reactive protein; HDL, high-density lipoprotein; IDL, intermediate-density lipoprotein; IQR, interquartile range; LDL, low-density lipoprotein; NMR, nuclear magnetic resonance; NT-proBNP, N-terminal pro B-type natriuretic peptide; VLDL, very low-density lipoprotein; svcam-1, soluble vascular cell adhesion molecule 1. SI conversion factors: To convert values for total cholesterol, HDL cholesterol, and LDL cholesterol to millimoles per liter, multiply by ; triglycerides to millimoles per liter, multiply by ; creatinine to micromoles per liter, multiply by 88.4; NT-proBNP to nanograms per liter, multiply by 8.4; homocysteine to micromoles per liter, multiply by 7.397; renin to picomoles per liter, multiply by CI, ) (P=.02) when svcam-1 level was removed from model 2. Level of svcam-1 correlated with NT-proBNP level (r=0.13; P<.001) and CRP level (r=0.13; 62

4 Table 3. Odds Ratios for Ischemic Stroke According to Baseline Plasma Levels of svcam-1, NT-proBNP, and CRP* Quarter Variables P Value for Trend svcam-1 level, ng/ml Mean (range) 399 ( ) 497 ( ) 587 ( ) 791 ( ) Model 1, OR (95% CI) ( ) 1.87 ( ) 2.34 ( ).004 Model 2, OR (95% CI) ( ) 1.91 ( ) 2.24 ( ).009 NT-proBNP level, pmol/l Mean (range) 3.9 ( ) 9.6 ( ) 19.1 ( ) 84.6 ( ) Model 1, OR (95% CI) ( ) 1.27 ( ) 1.90 ( ).005 Model 2, OR (95% CI) ( ) 1.08 ( ) 1.62 ( ).13 CRP level, mg/l Mean (range) 0.5 ( ) 1.3 ( ) 2.9 ( ) 10.6 ( ) Model 1, OR (95% CI) ( ) 1.35 ( ) 1.32 ( ).13 Model 2, OR (95% CI) ( ) 1.04 ( ) 1.00 ( ).99 Abbreviations: CI, confidence interval; CRP, C-reactive protein; NT-proBNP, N-terminal pro B-type natriuretic peptide; OR, odds ratio; svcam-1, soluble vascular cell adhesion molecule 1. *Model 1 is not adjusted, except through matching variables. Model 2 is adjusted through matching and also for baseline systolic blood pressure; total cholesterol level; a history of current smoking, diabetes mellitus, and peripheral arterial disease; and antihypertensive medication other than -blockers, calcium channel blockers, or diuretics. In addition, model 2 for svcam-1 level is adjusted for NT-proBNP and CRP levels, model 2 for NT-proBNP level is adjusted for svcam-1 and CRP levels, and model 2 for CRP level is adjusted for svcam-1 and NT-proBNP levels. P.001), and NT-proBNP level correlated with CRP level (r=0.14; P.001). The Hosmer and Lemeshow goodness of fit test gave a 2 value of 12 (P=.15), indicating a satisfactory fit for model 2. The combination of svcam-1 and NT-proBNP levels had a multiplicative effect on the OR for ischemic stroke, such that individuals in the highest quarters for both svcam-1 and NT-proBNP levels had 3.6 times the risk of ischemic stroke compared with patients in the lowest quarters for both biologic markers (Figure). Other variables that were predictors after multiple-variable analysis were a history of current smoking, diabetes mellitus, and peripheral arterial disease and taking antihypertensive medication other than -blockers, calcium channel blockers, or diuretics. The overall 24% reduction in ischemic stroke incidence by active treatment 3 was similar across all quarters of svcam-1 and NT-proBNP levels (data not shown). Baseline svcam-1 level was associated with similarly increased ischemic stroke risk in patients assigned to placebo and active therapy; the OR for patients in the highest, as compared with the lowest, quarter was 2.36 (95% CI, ) for active treatment and 2.18 (95% CI, ) for placebo. Plasma NT-proBNP level also showed a similar association with ischemic stroke risk for patients assigned to placebo and active therapy, although neither association achieved statistical significance (data not shown). COMMENT Levels of svcam-1 and NT-proBNP were independent predictors of ischemic stroke risk in this population of patients with established cerebrovascular disease, more than 90% having had previous ischemic stroke or TIA. Level of NT-proBNP correlated with svcam-1 level, and the OR for prediction of ischemic stroke by NT-proBNP level was borderline nonsignificant when svcam-1 level was included in the multiple-variable model. Odds Ratio NT-proBNP Quarter 2 1 Vascular cell adhesion molecule 1 is mainly expressed by activated endothelial and smooth muscle cells in atherosclerotic plaques, and it has therefore been proposed that the predictive value of svcam-1 level may be limited to individuals with more advanced atherosclerosis at the time of measurement. 13 Our finding that svcam-1 levels in the second, third, and fourth quarters were associated with similarly increased ischemic stroke risk suggests that for most PROGRESS participants their increased risk of recurrent stroke was a consequence of their increased atherosclerotic plaque bur svcam-1 Quarter Figure. Multiple variable adjusted odds ratio for recurrent ischemic stroke according to quarters of soluble vascular cell adhesion molecule 1 (svcam-1) and N-terminal pro B-type natriuretic peptide (NT-proBNP) concentration. The reference group is those subjects in quarter 1 for both svcam-1 and NT-proBNP concentrations. The ranges of svcam-1 and NT-proBNP levels for each quarter are presented in Table 3. Adjustment variables are listed in the Table 3 footnote regarding model

5 den. Angiotensin II induces VCAM-1 expression in vasculature 14 and may therefore contribute to the elevated svcam-1 levels in patients at increased risk of recurrent ischemic stroke. However, renin level did not predict ischemic stroke in this population. Blood pressure lowering with perindopril-based therapy reduced ischemic stroke incidence by 24%, 3 indicating that a proportion of ischemic strokes in PROGRESS participants assigned placebo was attributable to mechanisms responsive to perindopril-based therapy; however, a larger proportion of ischemic strokes was attributable to mechanisms unresponsive to this therapy. Our finding that svcam level predicted recurrent ischemic stroke in patients receiving perindopril-based therapy suggests that svcam-1 level is a marker of mechanisms of ischemic stroke pathogenesis distinct from those responsive to perindoprilbased therapy. Identification of these mechanisms may assist development of novel therapies for the prevention of recurrent ischemic stroke. Our finding that NT-proBNP level predicted ischemic stroke risk is in agreement with previous reports that B- type natriuretic peptide and NT-proBNP levels are associated with an increased risk of death and cardiovascular events, including heart failure, myocardial infarction, atrial fibrillation, and stroke or transient ischemic attack. 7,8,15,16 Whereas previous studies examined initial stroke, to our knowledge, our study is the first to show that B-type natriuretic peptides predict recurrent ischemic stroke. We found that homocysteine level did not predict recurrent ischemic stroke, although other studies reported that homocysteine level predicts recurrent ischemic stroke risk. 17,18 Level of CRP is an established risk factor for cardiovascular events, including initial ischemic stroke, but there are limited data about the association of CRP level with recurrent ischemic stroke risk. 22 The present study was confined to plasma samples frozen within 24 hours of collection, and the OR for recurrent ischemic stroke for patients in the highest, as compared with the lowest, quarter for CRP level was 1.32 (95% CI, ) in an analysis adjusted only for matching. In a separate analysis of nearly twice the number of cases and controls using PROGRESS plasma samples frozen within 48 hours of collection, the OR for recurrent ischemic stroke for patients in the highest, as compared with the lowest, third for CRP level was 1.52 (95% CI, ), 23 and CRP level remained a predictor of recurrent ischemic stroke after adjustment for fibrinogen level. 23 By contrast, the present study showed that CRP level did not predict recurrent ischemic stroke after adjustment for svcam and NT-proBNP levels. Plasma lipids levels have only a weak association with ischemic stroke risk Lipoprotein particle size analysis was proposed to provide better prediction of cardiovascular risk than measurement of plasma lipids levels because it permits specific quantification of the more atherogenic small low-density lipoprotein particles. 29 To our knowledge, this is the first report of the association of lipoprotein particle concentration and size and ischemic stroke risk, and our findings support the lack of association between plasma lipids levels and recurrent ischemic stroke risk. There are several limitations to our study. Our analyses were based on single baseline determinations that may not accurately reflect risk factor levels before enrollment or over the mean 3.9 years of follow-up. In addition, frozen storage may affect lipoprotein analysis, 30 although plasma samples from cases and controls were treated in an identical and blinded fashion. These sources of variability are unlikely to account for the observed associations between svcam-1 and NT-proBNP levels and recurrent ischemic stroke risk because any random misclassification would bias results toward the null hypothesis and systematic differences between cases and controls are unlikely. Another limitation of our study relates to the heterogeneity of ischemic stroke subtypes because these may have different mechanisms of pathogenesis. Ischemic strokes include those due to smallvessel, large-vessel, and cardioembolic causes, and it is possible our grouping of ischemic strokes masked risk factors for specific ischemic stroke subtypes. For example, svcam-1 level may be a marker for smalland large-vessel causes of ischemic stroke, whereas NT-proBNP level may be a marker for cardiac dysfunction that contributes to cardioembolic stroke. Further large studies are required to examine whether these and other potential risk factors are stroke subtype specific. In conclusion, we performed head-to-head evaluation of a large number of potential risk factors for recurrent ischemic stroke. We found svcam-1 and NT-proBNP levels independently predicted ischemic stroke risk in patients with previous stroke or TIA, and svcam-1 level was similarly predictive in patients receiving placebo or perindopril-based, blood pressure lowering treatment. The prognostic information provided by svcam-1 and NT-proBNP levels was in addition to that provided by traditional risk factors for ischemic stroke and may enable more effective targeting of prevention strategies. We also showed levels of CRP, homocysteine, renin, and lipids and lipoprotein particle concentration and size were not associated with recurrent ischemic stroke risk. Characterization of the mechanisms of ischemic stroke pathogenesis associated with increased svcam-1 level may lead to development of therapies that provide benefits additional to those provided by angiotensin-converting enzyme inhibitor based, blood pressure lowering therapies. Accepted for Publication: September 16, Correspondence: Duncan J. Campbell, MD, PhD, St Vincent s Institute of Medical Research, 41 Victoria Parade, Fitzroy, Victoria 3065, Australia (dcampbell@svi.edu.au). Author Contributions: Drs Campbell and Woodward had full access to all data in the study and take full responsibility for the integrity of the data and the accuracy of the data analysis. Study concept and design: Campbell, Woodward, Chalmers, Jenkins, Kemp, Neal, and MacMahon. Acquisition of data: Campbell, Woodward, Jenkins, and Neal. Analysis and interpretation of data: Campbell, Woodward, Colman, Jenkins, Neal, and Patel. Drafting of the manuscript: Campbell and Woodward. Critical revision of the manuscript for important intellectual content: Campbell, Woodward, Chalmers, Colman, Jenkins, Kemp, Neal, Patel, and MacMahon. Statistical analysis: Woodward and Colman. Obtained funding: Campbell, 64

6 Woodward, Chalmers, Jenkins, Kemp, and Neal. Administrative, technical, and material support: Campbell, Jenkins, Kemp, Neal, Patel, and MacMahon. Study supervision: Campbell and Woodward. Financial Disclosure: Drs Chalmers and MacMahon hold research grants from Servier, Neuilly-Sur-Seine CEDEX, France, as chief investigators for the Perindopril Protection Against Recurrent Stroke Study (PROGRESS) and Action in Diabetes and Vascular Disease Preterax and Diamicron MR Controlled Evaluation (ADVANCE), administered by the University of Sydney, Sydney, Australia. Dr Neal is an investigator for ADVANCE, which is sponsored by Servier. Drs Chalmers, Patel, Neal, Woodward, and MacMahon have also received honoraria from Servier for speaking in relation to PROGRESS and/or ADVANCE at scientific meetings. Funding/Support: This study was funded by grant 5 R01 HL from the National Institutes of Health, Bethesda, Md, and grants from the National Health and Medical Research Council of Australia, Canberra, and the National Heart Foundation of Australia, Melbourne. PROGRESS was funded by grants from Servier; the Health Research Council of New Zealand, Auckland; and the National Health and Medical Research Council of Australia. Role of the Sponsor: The National Institutes of Health, the National Health and Medical Research Council of Australia, the National Heart Foundation of Australia, the Health Research Council of New Zealand, and Servier had no role in the design or conduct of the study and no involvement in the management, analysis, and interpretation of the data. Servier had opportunity to review the manuscript but no right to influence the manuscript. Additional Information: Drs Campbell and Neal are recipients of Career Development Awards, and Dr Jenkins is recipient of a Clinical Research Fellowship from the National Heart Foundation of Australia. Dr Kemp is an Australian Research Council Federation Fellow. REFERENCES 1. Hollander M, Koudstaal PJ, Bots ML, Grobbee DE, Hofman A, Breteler MM. Incidence, risk, and case fatality of first ever stroke in the elderly population: the Rotterdam Study. J Neurol Neurosurg Psychiatry. 2003;74: Hankey GJ, Warlow CP. Treatment and secondary prevention of stroke: evidence, costs, and effects on individuals and populations. Lancet. 1999;354: PROGRESS Collaborative Group. Randomised trial of a perindopril-based bloodpressure-lowering regimen among 6,105 individuals with previous stroke or transient ischaemic attack. Lancet. 2001;358: Hardie K, Hankey GJ, Jamrozik K, Broadhurst RJ, Anderson C. Ten-year risk of first recurrent stroke and disability after first-ever stroke in the Perth Community Stroke Study. Stroke. 2004;35: Hankey GJ, Jamrozik K, Broadhurst RJ, Forbes S, Anderson CS. Long-term disability after first-ever stroke and related prognostic factors in the Perth Community Stroke Study, Stroke. 2002;33: Fransen M, Anderson C, Chalmers J, et al. Effects of a perindopril-based blood pressure-lowering regimen on disability and dependency in 6105 patients with cerebrovascular disease: a randomized controlled trial. Stroke. 2003;34: Campbell DJ, Woodward M, Chalmers JP, et al. Prediction of heart failure by aminoterminal-pro-b-type natriuretic peptide and C-reactive protein in subjects with cerebrovascular disease. Hypertension. 2005;45: Campbell DJ, Woodward M, Chalmers JP, et al. Prediction of myocardial infarction by N-terminal-pro-B-type natriuretic peptide, C-reactive protein and renin in subjects with cerebrovascular disease. Circulation. 2005;112: Ubbink JB, Hayward Vermaak WJ, Bissbort S. Rapid high-performance liquid chromatographic assay for total homocysteine levels in human serum. J Chromatogr. 1991;565: Dudman NP, Guo XW, Crooks R, Xie L, Silberberg JS. Assay of plasma homocysteine: light sensitivity of the fluorescent 7-benzo-2-oxa-1, 3-diazole-4- sulfonic acid derivative, and use of appropriate calibrators. Clin Chem. 1996; 42: Chapman N, Huxley R, Anderson C, et al. Effects of a perindopril-based blood pressure-lowering regimen on the risk of recurrent stroke according to stroke subtype and medical history: the PROGRESS Trial. Stroke. 2004;35: Woodward M. Epidemiology: Study Design and Data Analysis. 2nd ed. Boca Raton, Fla: Chapman and Hall/CRC; Blake GJ, Ridker PM. Inflammatory bio-markers and cardiovascular risk prediction. J Intern Med. 2002;252: Tummala PE, Chen XL, Sundell CL, et al. Angiotensin II induces vascular cell adhesion molecule-1 expression in rat vasculature: a potential link between the reninangiotensin system and atherosclerosis. Circulation. 1999;100: Wang TJ, Larson MG, Levy D, et al. Plasma natriuretic peptide levels and the risk of cardiovascular events and death. N Engl J Med. 2004;350: Kistorp C, Raymond I, Pedersen F, Gustafsson F, Faber J, Hildebrandt P. N- terminal pro-brain natriuretic peptide, C-reactive protein, and urinary albumin levels as predictors of mortality and cardiovascular events in older adults. JAMA. 2005;293: Boysen G, Brander T, Christensen H, Gideon R, Truelsen T. Homocysteine and risk of recurrent stroke. Stroke. 2003;34: Toole JF, Malinow MR, Chambless LE, et al. Lowering homocysteine in patients with ischemic stroke to prevent recurrent stroke, myocardial infarction, and death: the Vitamin Intervention for Stroke Prevention (VISP) randomized controlled trial. JAMA. 2004;291: Ridker PM, Cushman M, Stampfer MJ, Tracy RP, Hennekens CH. Inflammation, aspirin, and the risk of cardiovascular disease in apparently healthy men. N Engl J Med. 1997;336: Rost NS, Wolf PA, Kase CS, et al. Plasma concentration of C-reactive protein and risk of ischemic stroke and transient ischemic attack: the Framingham study. Stroke. 2001;32: Curb JD, Abbott RD, Rodriguez BL, et al. C-reactive protein and the future risk of thromboembolic stroke in healthy men. Circulation. 2003;107: Arenillas JF, Alvarez-Sabin J, Molina CA, et al. C-reactive protein predicts further ischemic events in first-ever transient ischemic attack or stroke patients with intracranial large-artery occlusive disease. Stroke. 2003;34: Woodward M, Lowe GDO, Campbell DJ, et al. Associations of inflammatory and hemostatic variables with the risk of recurrent stroke. Stroke. 2005;36: Bowman TS, Sesso HD, Ma J, et al. Cholesterol and the risk of ischemic stroke. Stroke. 2003;34: Asia Pacific Cohort Studies Collaboration. Cholesterol, coronary heart disease, and stroke in the Asia Pacific region. Int J Epidemiol. 2003;32: Curb JD, Abbott RD, Rodriguez BL, et al. High density lipoprotein cholesterol and the risk of stroke in elderly men: the Honolulu Heart Program. Am J Epidemiol. 2004;160: Tirschwell DL, Smith NL, Heckbert SR, Lemaitre RN, Longstreth WT Jr, Psaty BM. Association of cholesterol with stroke risk varies in stroke subtypes and patient subgroups. Neurology. 2004;63: Asia Pacific Cohort Studies Collaboration. Serum triglycerides as a risk factor for cardiovascular diseases in the Asia-Pacific region. Circulation. 2004;110: Blake GJ, Otvos JD, Rifai N, Ridker PM. Low-density lipoprotein particle concentration and size as determined by nuclear magnetic resonance spectroscopy as predictors of cardiovascular disease in women. Circulation. 2002;106: Cohn JS, Rodriguez C, Jacques H, Tremblay M, Davignon J. Storage of human plasma samples leads to alterations in the lipoprotein distribution of apoc-iii and apoe. J Lipid Res. 2004;45: