Published online before print January 8, 2009, doi: 10.1161/STROKEAHA.108.524587
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(Stroke. 2009;40:683.)
© 2009 American Heart Association, Inc.
Original Contributions |
A Meta-Analysis of Candidate Gene Polymorphisms and Ischemic Stroke in 6 Study Populations
Association of Lymphotoxin-Alpha in Nonhypertensive Patients
From the Laboratory of Human Genetics (X.W., Y.S., L.L.), Beijing Hypertension League Institute, Beijing, China.; the Department of Human Genetics (S.C., V.H.B., H.A.E.), Roche Molecular Systems, Inc., Pleasanton, Calif; the Department of Medical and Chemical Laboratory Diagnostics (C.M.) and the University Clinic of Neurology (W.L.), Medical University Vienna, Vienna, Austria; the Institute of Epidemiology and Social Medicine (K.B.) and Department of Neurology (E.B.R.), University of Muenster, Muenster, Germany; S.F. Coordinating Center (W.S.B.), California Pacific Medical Center, Research Institute, San Francisco, Calif; the Department of Neurology (C.K.) and the Institute of Clinical Chemistry and Laboratory Medicine (J.L.), Ernst Moritz Arndt University, Greifswald, Germany; Roche Center for Medical Genomics (K.L.), F. Hoffmann-La Roche, Ltd, Basel, Switzerland; and the Center for Cardiovascular Disease Prevention and Division of Preventive Medicine (P.M.R., R.Y.L.Z., N.R.C.), Brigham and Women’s Hospital, Boston, Mass.
Correspondence to Nancy R. Cook, ScD, Division of Preventive Medicine, Department of Medicine, Brigham and Women’s Hospital, 900 Commonwealth Avenue East, Boston, MA 02215. E-mail ncook{at}rics.bwh.harvard.edu
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Abstract |
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Background and Purpose— Ischemic stroke is a multifactorial disease with a strong genetic component. Pathways, including lipid metabolism, systemic chronic inflammation, coagulation, blood pressure regulation, and cellular adhesion, have been implicated in stroke pathophysiology, and candidate gene polymorphisms in these pathways have been proposed as genetic risk factors.
Methods— We genotyped 105 simple deletions and single nucleotide polymorphisms from 64 candidate genes in 3550 patients and 6560 control subjects from 6 case–control association studies conducted in the United States, Europe, and China. Genotyping was performed using the same immobilized probe typing system and meta-analyses were based on summary logistic regressions for each study. The primary analyses were fixed-effects meta-analyses adjusting for age and sex with additive, dominant, and recessive models of inheritance.
Results— Although 7 polymorphisms showed a nominal additive association, none remained statistically significant after adjustment for multiple comparisons. In contrast, after stratification for hypertension, 2 lymphotoxin-alpha polymorphisms, which are in strong linkage disequilibrium, were significantly associated among nonhypertensive individuals: LTA 252A>G (additive model; OR, 1.41 with 95% CI, 1.20 to 1.65; P=0.00002) and LTA 26Thr>Asn (OR, 1.19 with 95% CI, 1.06 to 1.34; P=0.003). LTA 252A>G remained significant after adjustment for multiple testing using either the false discovery rate or by permutation testing. The 2 single nucleotide polymorphisms showed no association in hypertensive subjects (eg, LTA 252A>G, OR, 0.93; 95% CI, 0.84 to 1.03; P=0.17).
Conclusions— These observations may indicate an important role of LTA-mediated inflammatory processes in the pathogenesis of ischemic stroke.
Key Words: embolic stroke • genetics • hypertension • inflammation
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Introduction |
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Ischemic stroke is a complex multifactorial and polygenic disorder thought to reflect interactions between an individual’s genetic background and various environmental components. Previous studies have established hypertension, smoking, diabetes mellitus, body mass index, and age as reliable stroke risk predictors.1,2 However, these conventional stroke risk factors do not fully account for the overall risk of stroke. Several physiological pathways, including lipid metabolism, blood pressure regulation, coagulation, and cellular adhesion, are thought to play critical roles in stroke pathophysiology.
Among the known risk factors for ischemic stroke, hypertension contributes significantly to the onset of disease. Increased risk of stroke is not, however, limited to those with hypertension and the conventional stroke risk factors do not fully explain the risk among normotensives. Strategies to identify additional risk factors include stratification by hypertension3,4 and using blood pressure as a matching criterion for cases and control subjects.5 A key role for inflammation is suggested by observations that patients with hypertension have elevated circulating levels of markers of inflammation and that some antihypertensive therapies reduce both levels of proinflammatory markers and the risk of ischemic stroke in addition to lowering blood pressure.6
Both systemic and local inflammatory processes are implicated in the etiology of ischemic cerebrovascular disease and in the pathophysiology of cerebral ischemia.7 Viral and bacterial infections are independent risk factors for ischemic stroke8 and increased levels of systemic inflammatory markers such as C-reactive protein, leukocyte count, and fibrinogen are associated with increased risk of ischemic stroke.9 Moreover, many stroke-related diseases such as Alzheimer disease and atherosclerosis are initiated or worsened by systemic inflammation.10,11 Polymorphisms in the C-reactive protein gene have been recently associated with both circulating protein levels and cardiovascular events,12 demonstrating the potential impact of genetic variation. Proinflammatory cytokines are believed to play a pathogenic role in these diseases, and variations in cytokine genes have also been shown to influence both predisposition and penetrance by altering the transcription profile and pattern of proinflammatory cytokine production.13 For example, polymorphism in the lymphotoxin-alpha gene can enhance transcription and susceptibility to myocardial infarction.14 At the local level, migration of inflammatory cells to the vascular wall is associated with vascular changes leading to atherosclerosis, and early atherosclerotic lesions are preceded by inflammatory cell deposition in the subendothelial layer of major cerebral arteries and in small brain vessels.15 Genetic variants influencing inflammatory processes could potentially contribute to the etiology of stroke.
The complex etiology of stroke suggests that individual genetic polymorphisms have modest effects that are difficult to detect, as has been observed to date.16 Large studies are needed to assess these polymorphisms as risk factors. We report a 6-study meta-analysis to investigate the associations of 105 simple deletions and single nucleotide polymorphisms (SNPs) in inflammatory and cardiovascular system-related genes with susceptibility to ischemic stroke. To search for genetic risk factors contributing to ischemic stroke beyond hypertension, we stratified the study cohort on hypertension status.
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Materials and Methods |
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Study Sample Description
As part of the Roche Stroke SNP Consortium, results of 6 independent studies (Table 1) were pooled for this analysis. All 6 study samples were comprised of individuals with proven ischemic stroke status and healthy control subjects. All studies were approved by the local ethics committees and all participants gave informed consent. Briefly, the study subjects were recruited as follows:
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Physician’s Health Study
A nested case–control sample (319 cases, 2092 control subjects) was derived from the Physician’s Health Study (PHS) cohort consisting of 22 071 predominantly white US male physicians initially free of prior myocardial infarction, stroke, transient ischemic attack, and cancer who were enrolled in a placebo-controlled trial of aspirin and β-carotene for the primary prevention of cardiovascular disease and cancer.17 DNA was isolated from baseline blood samples provided by 14 916 (68%) of the participants. Incident cases of ischemic stroke were identified during an average 13-year follow-up and confirmed by medical record review. Control subjects were selected from study participants remaining free of reported cardiovascular disease and matched to cases of any cardiovascular disease by age, smoking, and time since study entry.18
Study of Osteoporotic Fractures
Ambulatory women were recruited from 4 clinical centers in Portland, Ore; Minneapolis, Minn; Baltimore, Md; and the Monongahela Valley, Pa.19 The Study of Osteoporotic Fractures (SOF) cohort consists of 9615 white women of at least 65 years of age who had not had bilateral hip replacement or earlier hip fracture at the time of recruitment. The stroke subgroup included here consists of 247 who had adjudicated ischemic strokes and 559 control subjects who remained free of stroke through the mean follow-up of 5.4 years. Individuals who died during follow-up were included in both cases and control subjects, avoiding survivor bias.
Westphalia, Germany
Cases (n=700) were recruited through the regional Westphalian Stroke Register in northwestern Germany.20 Standardized patient documentation included sociodemographic characteristics, comorbidities, stroke type and severity as well as details regarding the diagnostic and therapeutic procedures and complications; 96.8% had at least one CT or MRI of the brain during hospitalization. Control subjects (n=757) were recruited from the population-based Dortmund Health Study, conducted in the same region.21 Participants in this study were randomly drawn from the city’s registration office within 5-year age groups and stratified by sex. Medical histories were assessed in face-to-face interviews.
Pomerania, Germany
Cases (n=277) were recruited with a standardized patient assessment form; 96.5% had at least one CT or MRI during hospitalization. Control subjects for this region were recruited from the population-based Study of Health in Pomerania.22 Participants in Study of Health in Pomerania were 20 to 79 year olds randomly sampled from registration offices in the area. Face-to-face interviews with each participant included a short stroke symptom questionnaire. A random sample of 702 Study of Health in Pomerania participants who were free of self-reported stroke and within the same age range and sex distribution as the cases formed the control group.
Vienna Stroke Study
In the Vienna Stroke Registry, cases (n=844) consisted of consecutive white patients submitted to one of 9 stroke units within 72 hours of symptom onset of acute ischemic stroke. Patients who died on the way to the hospital or were first admitted to an intensive care unit were not included.23 All patients underwent cranial CT or MRI and were documented according to a standardized protocol, including stroke severity, risk factors, and medical history (with particular reference to vascular diseases). Control subjects (n=979) were voluntary participants in a health care program offered by the city of Vienna, were free of clinically manifest arterial vascular disease, and reported no arterial vascular diseases in first-degree relatives.
Stroke Hypertension Investigation in Genetics
Individuals were recruited from 6 geographical regions within China; 70% came from in and near Beijing. Cases (n=1163) were individuals who had a stroke within the previous 5 years as diagnosed by brain CT/MRI. The original goal was to identify SNPs that predispose to stroke independent of blood pressure; thus, randomly drawn population-based control subjects were initially individually matched to cases by sex, birth year ±3 years, geographic location, and blood pressure category (<140/90, 
140/90 and 
180/105, >180/105 mm Hg). Because some cases could not be matched, additional control subjects were recruited for a total of 1471 control subjects.5
Genotyping
A total of 105 polymorphisms from 64 genes were selected based on reported associations in the literature as well as on evidence of gene product involvement in cardiovascular disease and inflammatory processes. As previously described,24,25 3 separate multilocus polymerase chain reactions were carried out using biotinylated primer pools (Roche Molecular Systems, Inc). The resulting polymerase chain reaction product pools were denatured and hybridized to linear arrays of immobilized, sequence-specific oligonucleotide probes. Hybridized amplicon was detected using a streptavidin–horseradish peroxidase conjugate and a chromogenic substrate. Laboratory technicians were blinded to the case–control status of each sample. Genotype assignments were made either manually and independently by 2 researchers or made by capturing images with a flatbed scanner and then using proprietary software developed by Roche Molecular Systems to resolve probe signals into genotypes for all polymorphisms. Discordant or ambiguous results were resolved by repeat polymerase chain reaction or hybridization. Twenty polymorphisms were not available for PHS and 23 polymorphisms were genotyped in only a subset of the Vienna Stroke participants; for these, genotypes were obtained for >90.5% of the individuals typed. For each of the 62 polymorphisms genotyped for all 10 110 subjects across all 6 studies, the final genotype database was 97.5% complete. For the LTA [MIM 153440] 252A>G and LTA 26Thr>Asn polymorphisms, in particular, the database contained 6090 (not available for PHS) and 10 091 genotypes (99.8% complete), respectively.
Statistical Analysis
Individual-level data were provided from each study site to the Coordinating Center at the Brigham and Women’s Hospital in Boston. Prespecified inclusion criteria for the meta-analyses were age at least 20 years and no history of myocardial infarction. Cases were restricted to those experiencing an ischemic stroke and control subjects had no history of stroke.
Allele and genotype frequencies were estimated by study site among cases and control subjects separately using SAS GENETICS. Tests for Hardy-Weinberg equilibrium (HWE), both large-sample and exact, were conducted among cases and control subjects for each site (supplemental Table I
, available online at http://stroke.ahajournals.org). Results for the effect of each SNP on ischemic stroke were estimated for each study separately using logistic regression. Each analysis controlled for age and sex and assessed genetic effects under 3 modes of inheritance: additive, dominant, and recessive. In addition, analyses were conducted for each site using all 3 genotypes using a 2-degree of freedom test.
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Meta-analyses were conducted based on the summary logistic regression results for each study site.26 The primary analyses were fixed-effects meta-analyses adjusting for age and sex. These meta-analyses were also conducted across whites only (data not shown), because these comprised the majority of participants for 5 of the 6 studies. Effects for each of the 3 modes of inheritance were estimated. PROC MIXED of SAS was used for effect estimation. Tests for heterogeneity of the genetic effect across sites were conducted using the Q-statistic.27 For comparison, random-effects models were estimated that allowed the genetic effect to vary across sites using study-specific effect estimates and PROC MIXED of SAS. To adjust for multiple comparisons, the false discovery rate (FDR)28 was computed and stepdown permutation tests were conducted for selected comparisons.29
Other prespecified analyses adjusted for hypertension as well as age and sex. Across all studies, hypertensives were defined as having current or past antihypertensive medication, systolic blood pressure 140 mm Hg, or diastolic blood pressure 90 mm Hg. Additional smoking-adjusted analyses were limited to 5 studies due to the limited availability of smoking data for the Westphalian study participants. Subgroup analyses were conducted according to age, sex, presence of hypertension, or smoking (ever versus never).
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Results |
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A total of 3550 patients with stroke and 6560 control subjects were genotyped with inflammation and cardiovascular SNP panels in 6 study sites with common methodology and genotyping software. The characteristics of all participants from 6 study sites are listed in Table 1. Two studies were drawn from prospective cohorts; the PHS study followed only male subjects and the SOF study followed only female subjects. The Stroke Hypertension Investigation in Genetics (SHINING) study was comprised of subjects of Han ethnicity, whereas the 5 other study populations were >99% white. There was a greater proportion of hypertension among cases than in control subjects, except in the SHINING study subset, for which blood pressure had been matched between the majority of cases and control subjects.
Table 2 lists the results obtained in the primary fixed-effects meta-analysis for all polymorphic sites under dominant, additive, and recessive genetic modes of inheritance; similar results were observed under a random-effects meta-analysis (data not shown). Nine SNPs were nominally significant (P<0.05) under at least one mode of inheritance: ADRB3 [MIM 109691] Trp64Arg, CETP [MIM 118470] (–629)C>A, GNB3 [MIM 139130] 825C>T, IL4 [MIM 147780] (–590)C>T, LIPC [MIM 151670] (–480)C>T, LPL [MIM 609708] Ser447Ter, NOS3 [MIM 163729](–690)C>T, PON2 [MIM 602447] Ser311Cys, and TGFB1 [MIM 190180] (–509)C>T. To account for multiple hypothesis testing, the FDR or permutation testing was applied and none of these SNPs remained statistically significantly associated with ischemic stroke. Among white participants only, the same GNB3, LPL, NOS3, PON2, and TGFB1 SNPs were nominally significant under at least one mode of inheritance in addition to 8 others (APOB [MIM 107730] 71Ile>Thr, APOC3 [MIM 107720] 3175C>G, CCR5 [MIM 601373] (–2459)G>A, IL6 [MIM 147620] (–174)G>C, IL10 [MIM 124092] (–571)C>A, ITGA3 [MIM 192974] 873G>A, NOS2A [MIM 163730] 231C>T, TNF [MIM 191160] (–376)G>A), but none of the SNPs remained statistically significant after the FDR was applied (data not shown).
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The data were then stratified on age, sex, hypertension, or smoking status. No statistically significant associations were observed in the age- (Supplemental Table IIA) or sex-stratified (Supplemental Table IIB) analyses nor among those with current or past hypertension (Table 3) after adjustment with the FDR. In contrast, a large number of nominally significant associations with ischemic stroke among normotensives were observed (Table 4). The strongest associations under the additive and dominant models were for LTA 252A>G and LTA 26Thr>Asn, 2 SNPs in strong linkage disequilibrium, whereas NOS3 298Glu>Asp had the strongest association under the recessive model. After adjustment with FDR and permutation testing, only the LTA 252A>G SNP showed a significant association among those without hypertension. In the additive mode, the estimated relative risk across the 3 LTA 252 genotypes was 1.41 (P=0.00002) in the fixed-effects analysis and the FDR was 0.002 with P<0.01 in permutation testing. Results for the dominant model were similar (OR, 1.57; FDR, 0.005). In the random-effects meta-analysis (data not shown), the LTA 252A>G association with stroke under the dominant model (OR, 1.56) had an FDR of 0.02. Among whites only, LTA 252A>G was similarly associated with ischemic stroke among those without hypertension under additive and dominant models (OR, 1.28; P=0.016 and OR, 1.39; P=0.019, respectively). Minor allele frequencies among nonhypertensive control subjects are given in Table 4; frequencies among nonhypertensive cases were 0.37, 0.38, 0.31, 0.41, and 0.46 in SOF, Vienna, Westphalia, Pomerania, and SHINING, respectively.
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The point estimates for the OR were somewhat higher for LTA 252A>G, a polymorphism in intron 1, than for the nonsynonymous polymorphism LTA 26Thr>Asn, although the CIs overlapped after adjusting for age and sex (Figure A–D). The FDR values for the Thr>Asn polymorphism were also >0.05. The associations with stroke risk for both LTA SNPs reached statistical significance among normotensives within the individual studies of SHINING and Pomerania, whereas among hypertensives, the OR point estimates were usually just below 1 and were not statistically significant (Figure). This trend for increased stroke risk associated with the LTA SNPs among normotensives relative to hypertensives was observed across the other studies, although none of these individual associations was statistically significant. We note that the PHS cohort was genotyped only for the LTA 26Thr>Asn polymorphism under the expectation that this coding SNP could be functional and would be an effective "tag" for LTA 252 based on the very strong linkage disequilibrium between these 2 polymorphisms; furthermore, in the Vienna and Westphalia studies, some samples had missing genotypes for LTA 252A>G. When the meta-analysis was repeated with only those samples that had been genotyped for both LTA SNPs, the OR estimates were virtually identical (1.572 and 1.565 among normotensives under the dominant model for LTA 252 and LTA 26, respectively; data not shown). In addition, if the LTA 26 result for the PHS was imputed for the missing LTA 252 data, the additive result for LTA 252 would remain highly significant (OR, 1.30; P=0.0001). Alternatively, if a completely null estimate for the PHS was imputed, the overall result would remain significant (OR, 1.27; P=0.0004) and would continue to pass the stringent multiple comparisons testing.
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In the smoking-stratified analyses, no associations remained statistically significant after the FDR was applied, although under the dominant model, CD14 (-260)C>T was suggestively associated (OR, 1.24; P=0.001; FDR, 0.058) with ischemic stroke among never-smokers (Supplemental Table IIC). Both LTA SNPs were associated with a greater risk for ischemic stroke in never-smokers than ever-smokers under the dominant model and although these associations were not statistically significant after accounting for multiple testing, this trend was consistent across 5 studies (data not shown); the Westphalian study was excluded due to limited smoking data.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionAppendixReferences |
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In this meta-analysis, we evaluated the association between 105 polymorphisms in 64 inflammation and cardiovascular-related genes and ischemic stroke in 3550 cases and 6560 control subjects across 6 different studies. Although we could not further define subtypes of ischemic stroke, key strengths of our stroke consortium are that these analyses were not subject to publication bias and all studies used common genotyping reagents. In the primary meta-analysis, modest associations with stroke became nonsignificant after adjustment for multiple testing using the FDR or permutation testing. Stratification on sex or age also revealed no significant associations. Notably, subjects in 2 of our studies were limited to one sex and the consortium encompassed subjects recruited from different regions in Europe, North America, and China. We observed similar results among white participants only, but study population differences resulting in heterogeneity in stroke etiology could have obscured genetic associations.
Stratification on hypertension status did, however, reveal a statistically significant association for LTA 252A>G that remained after adjustment for multiple testing. Across 4 white populations and one Chinese population, the OR for LTA 252G was consistently greater among normotensive than hypertensive subjects. LTA 26Thr>Asn yielded similar results among study participants with genotypes at both sites, as expected, given the strong linkage disequilibrium between these 2 LTA SNPs. Although a recent Japanese study observed no association of these SNPs with any subtype of ischemic stroke,30 a smaller Hungarian study had previously reported LTA 252G as a risk factor for large vessel ischemic stroke31 and an earlier Korean study had identified the LTA 252AA genotype as a risk factor for cerebral infarction.32 We were unable to analyze ischemic stroke subtypes, but there is some evidence that subtypes may differ depending on hypertensive status.33 Although the number of nonhypertensives cases was limited to 1068, stratification by hypertension across our 6 populations may have reduced heterogeneity and thus enabled us to discern the modest risk associated with LTA polymorphism.
A role for LTA in chronic inflammation has been suggested by its ability to induce expression of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 on endothelial cells in vitro.34,35 LTA expression results in a localized infiltrate consisting of T cells, B cells, follicular and interdigitating dendritic cells, and macrophages.36 A recent mouse model study indicated that LTA was expressed in atherosclerotic lesions whose size correlated with concentration. Moreover, loss of the adjacent gene TNF did not affect development of lesions in mice fed an atherogenic diet.37 The A252G site is intronic but has been associated with higher transcriptional activity in a luciferase assay, whereas the variant protein bearing the LTA 26 threonine to asparagine substitution has been observed to induce greater expression of vascular cell adhesion molecule-1 and SELE mRNA in vascular smooth muscle cells. Because these 2 LTA SNPs are in almost complete LD, the variant protein level was estimated to be 1.5-fold higher than the wild-type.10 An increased level of the variant protein may contribute to the increased risk for ischemic stroke through inflammatory processes. Although the mechanism by which LTA polymorphisms influence inflammatory pathways is not clear, the meta-analysis presented here indicated that these LTA variants were associated with ischemic stroke in nonhypertensive patients.
It is believed that subjects with hypertension tend to develop chronic, low-grade systemic inflammation.38–40 Severity of inflammation caused by genetic variation could independently modify predisposition to ischemic stroke. Recent reports on the association of PDE4D variants with ischemic stroke among normotensives3,4 are consistent with the hypothesis that hypertension may obscure or mask the effect of inflammation-related genetic variants and that such genetic effects can be most readily observed in the absence of this major risk factor.
Smoking, like hypertension, can elicit an inflammatory response.41 In our study, the effect of LTA variation on stroke was more discernable among never-smokers than ever-smokers. Whether proinflammatory risks for ischemic stroke caused by hypertension, smoking, or carrying a risk allele are additive remains to be addressed by a carefully designed study.
Summary
Our 6-study analysis surveyed inflammatory and cardiovascular gene polymorphisms in examining the risk for ischemic stroke. Our results indicate that the LTA 252A>G and LTA 26Thr>Asn polymorphisms have significant effects on the risk for ischemic stroke in nonhypertensive subjects. We cannot rule out the possible importance of these polymorphisms in hypertensive subjects, but a much larger cohort may be needed to clarify the interaction of hypertension and inflammation in the etiology of ischemic stroke.
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Appendix |
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RMS Stroke SNP Consortium Investigators
Physicians’ Health Study: Nancy R. Cook, Paul M. Ridker, and Robert Y. L. Zee from the Center for Cardiovascular Disease Prevention and Division of Preventive Medicine, Brigham and Women’s Hospital, Boston, Mass. Study of Osteoporotic Fractures: Warren S. Browner, Steve R. Cummings, and Li-Yung Lui from the S.F. Coordinating Center, California Pacific Medical Center, Research Institute, San Francisco, Calif. Vienna Stroke Study: Georg Endler and Christine Mannhalter from the Department of Medical and Chemical Laboratory Diagnostics, Medical University Vienna, Vienna, Austria; and Stefan Greisenegger and Wolfgang Lalouschek from the University Clinic of Neurology, Medical University Vienna, Vienna, Austria. Pomerania and Westphalia Studies: Klaus Berger, Institute of Epidemiology and Social Medicine, University of Muenster, Muenster, Germany; Harald Funke, Department of Molecular Hemostaseology, University of Jena, Jena, Germany; E. Bernd Ringelstein, Department of Neurology, University of Muenster, Muenster, Germany; Florian Stoegbauer, Department of Neurology, University of Muenster, Muenster, Germany, and Klinikum Osnabrück, Osnabrück, Germany; Jan Luedemann, Institutes of Clinical Chemistry and Laboratory Medicine, University of Greifswald, Greifswald, Germany; Christof Kessler, and Department of Neurology, University of Greifswald, Greifswald, Germany. SHINING: Lisheng Liu, Yu Shi, and Xingyu Wang from the Laboratory of Human Genetics, Beijing Hypertension League Institute, Beijing, China. Roche Molecular Systems: Victoria H. Brophy, Suzanne Cheng, Henry A. Erlich, Andrea M. Johnson, and Brian K. Rhees from the Department of Human Genetics, Roche Molecular Systems, Inc, Pleasanton, Calif.
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Acknowledgments |
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The RMS Stroke SNP Consortium (see Appendix for full list of investigators) thanks the staff and many participants of the PHS, SOF, Vienna Stroke Registry, Westphalian Stroke Register, Dortmund Health Study, Pomeranian Stroke Study, Study of Health in Pomerania, and SHINING for their dedication and cooperation. We thank the RMS CVD project team for their expertise in developing the genotyping reagents used for these studies, the RMS High-Throughput Genotyping Group for their expert technical assistance in genotyping the SOF samples, and Michael Janisiw for assistance in genotyping the Viennese samples.
Sources of Funding
The SOF is supported by National Institutes of Health funding. The following institutes provide support: the National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS) and the National Institute on Aging (NIA) under the following grant numbers: AG05407, AR35582, AG05394, AR35584, AR35583, R01 AG005407, R01 AG027576-22, 2 R01 AG005394-22A1, and 2 R01 AG027574-22A1. Case assessments in the Westphalian and Pomeranian studies are part of the German "Competence Net Stroke," which is supported by the German Federal Ministry of Education and Research (01GI9909/3). Data collection in the Dortmund Health Study was funded by the German Migraine and Headache Society and unrestricted grants of equal share of a consortium of 7 pharmaceutical companies. The Study of Health in Pomerania is funded by grants from the German Federal Ministry of Education and Research (BMBF, 01ZZ96030) and from the Ministry for Education, Research and Cultural Affairs and the Ministry for Social Affairs of the Federal State of Mecklenburg-Vorpommern. The Vienna Stroke Registry is supported by research grants of the Medizinisch-Wissenschaftlicher Fonds des Bürgermeisters der Bundeshauptstadt Wien (project numbers 1540, 1829, 1970), of the Jubiläumsfonds der Oesterreichischen Nationalbank (project numbers 6866, 7115, 8281, 9344), and the Austrian Science Foundation (P13902-MED). The Vienna Stroke Registry is also supported by the Wiener Krankenanstaltenverbund. SHINING was funded through the Beijing Hypertension League Institute, in part through the National Infrastructure Program of Chinese Genetic Resource (2005DKA21300) and an unrestricted educational grant from F. Hoffmann-La Roche. The PHS is supported by grants from the National Heart Lung and Blood Institute (HL-58755, and HL-63293), the Doris Duke Charitable Foundation, the American Heart Association, and the Donald W. Reynolds Foundation, Las Vegas, Nev. Additional funding provided by the Fondation Leducq, Paris, France (P.M.R.).
Disclosures
S.C., V.H.B., and H.A.E. are employees of Roche Molecular Systems, Inc, which provided reagents and support for genotyping to all study sites under research collaborations and partial funding for the meta-analysis. K.L. is an employee of F. Hoffmann-La Roche, Ltd, which provided an unrestricted educational grant to the Beijing Hypertension League Institute.
Received April 30, 2008; revision received July 23, 2008; accepted July 25, 2008.
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