Rare Missense Functional Variants at COL4A1 and COL4A2 in Sporadic Intracerebral Hemorrhage.

ObjectiveTo test the genetic contribution of rare missense variants in COL4A1 and COL4A2 in which common variants are genetically associated with sporadic intracerebral hemorrhage (ICH), we performed rare variant analysis in multiple sequencing data for the risk for sporadic ICH.MethodsWe performed sequencing across 559Kbp at 13q34 including COL4A1 and COL4A2 among 2,133 individuals (1,055 ICH cases; 1,078 controls) in US-based and 1,492 individuals (192 ICH cases; 1,189 controls) from Scotland-based cohorts, followed by sequence annotation, functional impact prediction, genetic association testing, and in silico thermodynamic modeling.ResultsWe identified 107 rare nonsynonymous variants in sporadic ICH, of which two missense variants, rs138269346 (COL4A1I110T) and rs201716258 (COL4A2H203L), were predicted to be highly functional and occurred in multiple ICH cases but not in controls from the US-based cohort. The minor allele of rs201716258 was also present in Scottish ICH patients, and rs138269346 was observed in two ICH-free controls with a history of hypertension and myocardial infarction. Rs138269346 was nominally associated with non-lobar ICH risk (P=0.05), but not with lobar ICH (P=0.08), while associations between rs201716258 and ICH subtypes were non-significant (P>0.12). Both variants were considered pathogenic based on minor allele frequency (<0.00035 in EUR), predicted functional impact (deleterious or probably damaging), and in silico modeling studies (substantially altered physical length and thermal stability of collagen).ConclusionsWe identified rare missense variants in COL4A1/A2 in association with sporadic ICH. Our annotation and simulation studies suggest that these variants are highly functional and may represent targets for translational follow-up.

Genome-wide association study of cerebral small vessel disease reveals established and novel loci.

Intracerebral haemorrhage and small vessel ischaemic stroke (SVS) are the most acute manifestations of cerebral small vessel disease, with no established preventive approaches beyond hypertension management. Combined genome-wide association study (GWAS) of these two correlated diseases may improve statistical power to detect novel genetic factors for cerebral small vessel disease, elucidating underlying disease mechanisms that may form the basis for future treatments. Because intracerebral haemorrhage location is an adequate surrogate for distinct histopathological variants of cerebral small vessel disease (lobar for cerebral amyloid angiopathy and non-lobar for arteriolosclerosis), we performed GWAS of intracerebral haemorrhage by location in 1813 subjects (755 lobar and 1005 non-lobar) and 1711 stroke-free control subjects. Intracerebral haemorrhage GWAS results by location were meta-analysed with GWAS results for SVS from MEGASTROKE, using 'Multi-Trait Analysis of GWAS' (MTAG) to integrate summary data across traits and generate combined effect estimates. After combining intracerebral haemorrhage and SVS datasets, our sample size included 241 024 participants (6255 intracerebral haemorrhage or SVS cases and 233 058 control subjects). Genome-wide significant associations were observed for non-lobar intracerebral haemorrhage enhanced by SVS with rs2758605 [MTAG P-value (P) = 2.6 × 10-8] at 1q22; rs72932727 (P = 1.7 × 10-8) at 2q33; and rs9515201 (P = 5.3 × 10-10) at 13q34. In the GTEx gene expression library, rs2758605 (1q22), rs72932727 (2q33) and rs9515201 (13q34) are significant cis-eQTLs for PMF1 (P = 1 × 10-4 in tibial nerve), NBEAL1, FAM117B and CARF (P < 2.1 × 10-7 in arteries) and COL4A2 and COL4A1 (P < 0.01 in brain putamen), respectively. Leveraging S-PrediXcan for gene-based association testing with the predicted expression models in tissues related with nerve, artery, and non-lobar brain, we found that experiment-wide significant (P < 8.5 × 10-7) associations at three genes at 2q33 including NBEAL1, FAM117B and WDR12 and genome-wide significant associations at two genes including ICA1L at 2q33 and ZCCHC14 at 16q24. Brain cell-type specific expression profiling libraries reveal that SEMA4A, SLC25A44 and PMF1 at 1q22 and COL4A1 and COL4A2 at 13q34 were mainly expressed in endothelial cells, while the genes at 2q33 (FAM117B, CARF and NBEAL1) were expressed in various cell types including astrocytes, oligodendrocytes and neurons. Our cross-phenotype genetic study of intracerebral haemorrhage and SVS demonstrates novel genome-wide associations for non-lobar intracerebral haemorrhage at 2q33 and 13q34. Our replication of the 1q22 locus previous seen in traditional GWAS of intracerebral haemorrhage, as well as the rediscovery of 13q34, which had previously been reported in candidate gene studies with other cerebral small vessel disease-related traits strengthens the credibility of applying this novel genome-wide approach across intracerebral haemorrhage and SVS.

Association of Apolipoprotein E With Intracerebral Hemorrhage Risk by Race/Ethnicity: A Meta-analysis.

Importance: Genetic studies of intracerebral hemorrhage (ICH) have focused mainly on white participants, but genetic risk may vary or could be concealed by differing nongenetic coexposures in nonwhite populations. Transethnic analysis of risk may clarify the role of genetics in ICH risk across populations. Objective: To evaluate associations between established differences in ICH risk by race/ethnicity and the variability in the risks of apolipoprotein E (APOE) ε4 alleles, the most potent genetic risk factor for ICH. Design, Setting, and Participants: This case-control study of primary ICH meta-analyzed the association of APOE allele status on ICH risk, applying a 2-stage clustering approach based on race/ethnicity and stratified by a contributing study. A propensity score analysis was used to model the association of APOE with the burden of hypertension across race/ethnic groups. Primary ICH cases and controls were collected from 3 hospital- and population-based studies in the United States and 8 in European sites in the International Stroke Genetic Consortium. Participants were enrolled from January 1, 1999, to December 31, 2017. Participants with secondary causes of ICH were excluded from enrollment. Controls were regionally matched within each participating study. Main Outcomes and Measures: Clinical variables were systematically obtained from structured interviews within each site. APOE genotype was centrally determined for all studies. Results: In total, 13 124 participants (7153 [54.5%] male with a median [interquartile range] age of 66 [56-76] years) were included. In white participants, APOE ε2 (odds ratio [OR], 1.49; 95% CI, 1.24-1.80; P < .001) and APOE ε4 (OR, 1.51; 95% CI, 1.23-1.85; P < .001) were associated with lobar ICH risk; however, within self-identified Hispanic and black participants, no associations were found. After propensity score matching for hypertension burden, APOE ε4 was associated with lobar ICH risk among Hispanic (OR, 1.14; 95% CI, 1.03-1.28; P = .01) but not in black (OR, 1.02; 95% CI, 0.98-1.07; P = .25) participants. APOE ε2 and ε4 did not show an association with nonlobar ICH risk in any race/ethnicity. Conclusions and Relevance: APOE ε4 and ε2 alleles appear to affect lobar ICH risk variably by race/ethnicity, associations that are confirmed in white individuals but can be shown in Hispanic individuals only when the excess burden of hypertension is propensity score-matched; further studies are needed to explore the interactions between APOE alleles and environmental exposures that vary by race/ethnicity in representative populations at risk for ICH.

17p12 Influences Hematoma Volume and Outcome in Spontaneous Intracerebral Hemorrhage.

BACKGROUND AND PURPOSE: Hematoma volume is an important determinant of clinical outcome in spontaneous intracerebral hemorrhage (ICH). We performed a genome-wide association study (GWAS) of hematoma volume with the aim of identifying novel biological pathways involved in the pathophysiology of primary brain injury in ICH. METHODS: We conducted a 2-stage (discovery and replication) case-only genome-wide association study in patients with ICH of European ancestry. We utilized the admission head computed tomography to calculate hematoma volume via semiautomated computer-assisted technique. After quality control and imputation, 7 million genetic variants were available for association testing with ICH volume, which was performed separately in lobar and nonlobar ICH cases using linear regression. Signals with P<5×10-8 were pursued in replication and tested for association with admission Glasgow coma scale and 3-month post-ICH dichotomized (0-2 versus 3-6) modified Rankin Scale using ordinal and logistic regression, respectively. RESULTS: The discovery phase included 394 ICH cases (228 lobar and 166 nonlobar) and identified 2 susceptibility loci: a genomic region on 22q13 encompassing PARVB (top single-nucleotide polymorphism rs9614326: ß, 1.84; SE, 0.32; P=4.4×10-8) for lobar ICH volume and an intergenic region overlying numerous copy number variants on 17p12 (top single-nucleotide polymorphism rs11655160: ß, 0.95; SE, 0.17; P=4.3×10-8) for nonlobar ICH volume. The replication included 240 ICH cases (71 lobar and 169 nonlobar) and corroborated the association for 17p12 (P=0.04; meta-analysis P=2.5×10-9; heterogeneity, P=0.16) but not for 22q13 (P=0.49). In multivariable analysis, rs11655160 was also associated with lower admission Glasgow coma scale (odds ratio, 0.17; P=0.004) and increased risk of poor 3-month modified Rankin Scale (odds ratio, 1.94; P=0.045). CONCLUSIONS: We identified 17p12 as a novel susceptibility risk locus for hematoma volume, clinical severity, and functional outcome in nonlobar ICH. Replication in other ethnicities and follow-up translational studies are needed to elucidate the mechanism mediating the observed association.

Diagnosis and Management of Acute Ischemic Stroke.

Acute ischemic stroke (AIS) is among the leading causes of death and long-term disability. Intravenous tissue plasminogen activator has been the mainstay of acute therapy. Recently, several prospective randomized trials documented the value of endovascular revascularization in selected patients with large-vessel occlusion within the anterior circulation. This finding has led to a paradigm shift in the management of AIS, including wide adoption of noninvasive neuroimaging to assess vessel patency and tissue viability, with the supplemental and independent use of intravenous tissue plasminogen activator to improve clinical outcomes. In this article, we review the landmark studies on management of AIS and the current position on the diagnosis and management of AIS. The review also highlights the importance of early stabilization and prompt initiation of therapeutic interventions before, during, and after the diagnosis of AIS within and outside of the hospital.

Multiplanar computed tomography reconstruction to aid in recognition of the middle cerebral artery "Dot" sign: The sagittal string sign.

While non-contrast head computed tomography is effective in detecting blood, it is not sensitive in diagnosing hyperacute ischemic stroke. One neuroradiologic marker for early thromboembolic occlusion of the distal middle cerebral artery is the middle cerebral artery "dot" sign. The "dot" seen on the typical axial plane represents a hyperdensity of the middle cerebral artery in the Sylvian fissure. A review of medical literature was conducted via PubMed utilizing search phrases "MCA," "dot," and "sign." The review was limited to the intravenous tissue-type plasminogen activator era, 1996 and on. Articles were analyzed to determine the use of the sagittal plane of non-contrast head computed tomography to locate the middle cerebral artery "dot" sign. The search terms yielded 11 results which revealed that computed tomography reconstruction and sagittal planes were not used for detection of the middle cerebral artery "dot" signs. Our patient had no known past medical history. The initial non-contrast head computed tomography was read as having a hypodensity in the right insular region and a middle cerebral artery "dot" sign. Multiplanar reconstruction of the computed tomography demonstrated a hyperdense sagittal string-like appearance of the middle cerebral artery along the Sylvian fissure. Computed tomography angiography confirmed the M2 occlusion. This is the first report of using the head computed tomography sagittal plane for diagnosis of the middle cerebral artery "dot" sign. Incorporating multiplanar reconstruction and producing the sagittal plane may lead to a higher sensitivity of the middle cerebral artery "dot" sign. Further studies incorporating a patient cohort will be needed to determine how much the sagittal plane view augments predictive value of the middle cerebral artery "dot" sign.

Genetic variants in CETP increase risk of intracerebral hemorrhage.

OBJECTIVE: In observational epidemiologic studies, higher plasma high-density lipoprotein cholesterol (HDL-C) has been associated with increased risk of intracerebral hemorrhage (ICH). DNA sequence variants that decrease cholesteryl ester transfer protein (CETP) gene activity increase plasma HDL-C; as such, medicines that inhibit CETP and raise HDL-C are in clinical development. Here, we test the hypothesis that CETP DNA sequence variants associated with higher HDL-C also increase risk for ICH. METHODS: We performed 2 candidate-gene analyses of CETP. First, we tested individual CETP variants in a discovery cohort of 1,149 ICH cases and 1,238 controls from 3 studies, followed by replication in 1,625 cases and 1,845 controls from 5 studies. Second, we constructed a genetic risk score comprised of 7 independent variants at the CETP locus and tested this score for association with HDL-C as well as ICH risk. RESULTS: Twelve variants within CETP demonstrated nominal association with ICH, with the strongest association at the rs173539 locus (odds ratio [OR] = 1.25, standard error [SE] = 0.06, p = 6.0 × 10-4 ) with no heterogeneity across studies (I2 = 0%). This association was replicated in patients of European ancestry (p = 0.03). A genetic score of CETP variants found to increase HDL-C by ∼2.85mg/dl in the Global Lipids Genetics Consortium was strongly associated with ICH risk (OR = 1.86, SE = 0.13, p = 1.39 × 10-6 ). INTERPRETATION: Genetic variants in CETP associated with increased HDL-C raise the risk of ICH. Given ongoing therapeutic development in CETP inhibition and other HDL-raising strategies, further exploration of potential adverse cerebrovascular outcomes may be warranted. Ann Neurol 2016;80:730-740.

Rare Coding Variation and Risk of Intracerebral Hemorrhage.

BACKGROUND AND PURPOSE: Intracerebral hemorrhage has a substantial genetic component. We performed a preliminary search for rare coding variants associated with intracerebral hemorrhage. METHODS: A total of 757 cases and 795 controls were genotyped using the Illumina HumanExome Beadchip (Illumina, Inc, San Diego, CA). Meta-analyses of single-variant and gene-based association were computed. RESULTS: No rare coding variants were associated with intracerebral hemorrhage. Three common variants on chromosome 19q13 at an established susceptibility locus, encompassing TOMM40, APOE, and APOC1, met genome-wide significance (P<5e-08). After adjusting for the APOE epsilon alleles, this locus was no longer convincingly associated with intracerebral hemorrhage. No gene reached genome-wide significance level in gene-based association testing. CONCLUSIONS: Although no coding variants of large effect were detected, this study further underscores a major challenge for the study of genetic susceptibility loci; large sample sizes are required for sufficient power except for loci with large effects.

Common variation in COL4A1/COL4A2 is associated with sporadic cerebral small vessel disease.

OBJECTIVES: We hypothesized that common variants in the collagen genes COL4A1/COL4A2 are associated with sporadic forms of cerebral small vessel disease. METHODS: We conducted meta-analyses of existing genotype data among individuals of European ancestry to determine associations of 1,070 common single nucleotide polymorphisms (SNPs) in the COL4A1/COL4A2 genomic region with the following: intracerebral hemorrhage and its subtypes (deep, lobar) (1,545 cases, 1,485 controls); ischemic stroke and its subtypes (cardioembolic, large vessel disease, lacunar) (12,389 cases, 62,004 controls); and white matter hyperintensities (2,733 individuals with ischemic stroke and 9,361 from population-based cohorts with brain MRI data). We calculated a statistical significance threshold that accounted for multiple testing and linkage disequilibrium between SNPs (p < 0.000084). RESULTS: Three intronic SNPs in COL4A2 were significantly associated with deep intracerebral hemorrhage (lead SNP odds ratio [OR] 1.29, 95% confidence interval [CI] 1.14-1.46, p = 0.00003; r(2) > 0.9 between SNPs). Although SNPs associated with deep intracerebral hemorrhage did not reach our significance threshold for association with lacunar ischemic stroke (lead SNP OR 1.10, 95% CI 1.03-1.18, p = 0.0073), and with white matter hyperintensity volume in symptomatic ischemic stroke patients (lead SNP OR 1.07, 95% CI 1.01-1.13, p = 0.016), the direction of association was the same. There was no convincing evidence of association with white matter hyperintensities in population-based studies or with non-small vessel disease cerebrovascular phenotypes. CONCLUSIONS: Our results indicate an association between common variation in the COL4A2 gene and symptomatic small vessel disease, particularly deep intracerebral hemorrhage. These findings merit replication studies, including in ethnic groups of non-European ancestry.

APOE ε variants increase risk of warfarin-related intracerebral hemorrhage.

OBJECTIVE: We aimed to assess the effect of APOE ε variants on warfarin-related intracerebral hemorrhage (wICH), evaluated their predictive power, and tested for interaction with warfarin in causing wICH. METHODS: This was a prospective, 2-stage (discovery and replication), case-control study. wICH was classified as lobar or nonlobar based on the location of the hematoma. Controls were sampled from ambulatory clinics (discovery) and random digit dialing (replication). APOE ε variants were directly genotyped. A case-control design and logistic regression analysis were utilized to test for association between APOE ε and wICH. A case-only design and logistic regression analysis were utilized to test for interaction between APOE ε and warfarin. Receiver operating characteristic curves were implemented to evaluate predictive power. RESULTS: The discovery stage included 319 wICHs (44% lobar) and 355 controls. APOE ε2 was associated with lobar (odds ratio [OR] 2.46; p < 0.001) and nonlobar wICH (OR 1.67; p = 0.04), whereas ε4 was associated with lobar (OR 2.09; p < 0.001) but not nonlobar wICH (p = 0.35). The replication stage (63 wICHs and 1,030 controls) confirmed the association with ε2 (p = 0.03) and ε4 (p = 0.003) for lobar but not for nonlobar wICH (p > 0.20). Genotyping information on APOE ε variants significantly improved case/control discrimination of lobar wICH (C statistic 0.80). No statistical interaction between warfarin and APOE was found (p > 0.20). CONCLUSIONS: APOE ε variants constitute strong risk factors for lobar wICH. APOE exerts its effect independently of warfarin, although power limitations render this absence of interaction preliminary. Evaluation of the predictive ability of APOE in cohort studies is warranted.

Heritability estimates identify a substantial genetic contribution to risk and outcome of intracerebral hemorrhage.

BACKGROUND AND PURPOSE: Previous studies suggest that genetic variation plays a substantial role in occurrence and evolution of intracerebral hemorrhage (ICH). Genetic contribution to disease can be determined by calculating heritability using family-based data, but such an approach is impractical for ICH because of lack of large pedigree-based studies. However, a novel analytic tool based on genome-wide data allows heritability estimation from unrelated subjects. We sought to apply this method to provide heritability estimates for ICH risk, severity, and outcome. METHODS: We analyzed genome-wide genotype data for 791 ICH cases and 876 controls, and determined heritability as the proportion of variation in phenotype attributable to captured genetic variants. Contribution to heritability was separately estimated for the APOE (encoding apolipoprotein E) gene, an established genetic risk factor, and for the rest of the genome. Analyzed phenotypes included ICH risk, admission hematoma volume, and 90-day mortality. RESULTS: ICH risk heritability was estimated at 29% (SE, 11%) for non-APOE loci and at 15% (SE, 10%) for APOE. Heritability for 90-day ICH mortality was 41% for non-APOE loci and 10% (SE, 9%) for APOE. Genetic influence on hematoma volume was also substantial: admission volume heritability was estimated at 60% (SE, 70%) for non-APOEloci and at 12% (SE, 4%) for APOE. CONCLUSIONS: Genetic variation plays a substantial role in ICH risk, outcome, and hematoma volume. Previously reported risk variants account for only a portion of inherited genetic influence on ICH pathophysiology, pointing to additional loci yet to be identified.

Common variants within oxidative phosphorylation genes influence risk of ischemic stroke and intracerebral hemorrhage.

BACKGROUND AND PURPOSE: Previous studies demonstrated association between mitochondrial DNA variants and ischemic stroke (IS). We investigated whether variants within a larger set of oxidative phosphorylation (OXPHOS) genes encoded by both autosomal and mitochondrial DNA were associated with risk of IS and, based on our results, extended our investigation to intracerebral hemorrhage (ICH). METHODS: This association study used a discovery cohort of 1643 individuals, a validation cohort of 2432 individuals for IS, and an extension cohort of 1476 individuals for ICH. Gene-set enrichment analysis was performed on all structural OXPHOS genes, as well as genes contributing to individual respiratory complexes. Gene-sets passing gene-set enrichment analysis were tested by constructing genetic scores using common variants residing within each gene. Associations between each variant and IS that emerged in the discovery cohort were examined in validation and extension cohorts. RESULTS: IS was associated with genetic risk scores in OXPHOS as a whole (odds ratio [OR], 1.17; P=0.008) and complex I (OR, 1.06; P=0.050). Among IS subtypes, small vessel stroke showed association with OXPHOS (OR, 1.16; P=0.007), complex I (OR, 1.13; P=0.027), and complex IV (OR, 1.14; P=0.018). To further explore this small vessel association, we extended our analysis to ICH, revealing association between deep hemispheric ICH and complex IV (OR, 1.08; P=0.008). CONCLUSIONS: This pathway analysis demonstrates association between common genetic variants within OXPHOS genes and stroke. The associations for small vessel stroke and deep ICH suggest that genetic variation in OXPHOS influences small vessel pathobiology. Further studies are needed to identify culprit genetic variants and assess their functional consequences.

Burden of blood pressure-related alleles is associated with larger hematoma volume and worse outcome in intracerebral hemorrhage.

BACKGROUND AND PURPOSE: Intracerebral hemorrhage (ICH) is the acute manifestation of a progressive disease of the cerebral small vessels. The severity of this disease seems to influence not only risk of ICH but also the size of the hematoma. As the burden of high blood pressure-related alleles is associated with both hypertension-related end-organ damage and risk of ICH, we sought to determine whether this burden influences ICH baseline hematoma volume. METHODS: Prospective study in subjects of European descent with supratentorial ICH who underwent genome-wide genotyping. Forty-two single nucleotide polymorphisms associated with high blood pressure were identified from a publicly available database. A genetic risk score was constructed based on these single nucleotide polymorphisms. The score was used as the independent variable in univariate and multivariate regression models for admission ICH volume and poor clinical outcome (modified Rankin Scale, 3-6). RESULTS: A total of 323 ICH cases were enrolled in the study (135 deep and 188 lobar intracranial hematomas). The blood pressure-based genetic risk score was associated with both baseline hematoma volume and poor clinical outcome specifically in deep ICH. In multivariate regression analyses, each additional SD of the score increased mean deep ICH volume by 28% (or 2.7 mL increase; ß=0.28; SE=0.11; P=0.009) and risk of poor clinical outcome by 71% (odds ratio, 1.71; 95% confidence interval, 1.05-2.80; P=0.03). CONCLUSIONS: Increasing numbers of high blood pressure-related alleles are associated with mean baseline hematoma volume and poor clinical outcome in ICH. These findings suggest that the small vessel vasculopathy responsible for the occurrence of the hemorrhage also influences its volume.

Burden of risk alleles for hypertension increases risk of intracerebral hemorrhage.

BACKGROUND AND PURPOSE: Genetic variation influences risk of intracerebral hemorrhage (ICH). Hypertension (HTN) is a potent risk factor for ICH and several common genetic variants (single nucleotide polymorphisms [SNPs]) associated with blood pressure levels have been identified. We sought to determine whether the cumulative burden of blood pressure-related SNPs is associated with risk of ICH and pre-ICH diagnosis of HTN. METHODS: We conducted a prospective multicenter case-control study in 2272 subjects of European ancestry (1025 cases and 1247 control subjects). Thirty-nine SNPs reported to be associated with blood pressure levels were identified from the National Human Genome Research Institute genomewide association study catalog. Single-SNP association analyses were performed for the outcomes ICH and pre-ICH HTN. Subsequently, weighted and unweighted genetic risk scores were constructed using these SNPs and entered as the independent variable in logistic regression models with ICH and pre-ICH HTN as the dependent variables. RESULTS: No single SNP was associated with either ICH or pre-ICH HTN. The blood pressure-based unweighted genetic risk score was associated with risk of ICH (OR, 1.11; 95% CI, 1.02-1.21; P=0.01) and the subset of ICH in deep regions (OR, 1.18; 95% CI, 1.07-1.30; P=0.001), but not with the subset of lobar ICH. The score was associated with a history of HTN among control subjects (OR, 1.17; 95% CI, 1.04-1.31; P=0.009) and ICH cases (OR, 1.15; 95% CI, 1.01-1.31; P=0.04). Similar results were obtained when using a weighted score. CONCLUSIONS: Increasing numbers of high blood pressure-related alleles are associated with increased risk of deep ICH as well as with clinically identified HTN.

APOE genotype and extent of bleeding and outcome in lobar intracerebral haemorrhage: a genetic association study.

BACKGROUND: Carriers of APOE ε2 and ε4 have an increased risk of intracerebral haemorrhage (ICH) in lobar regions, presumably because of the effects of these gene variants on risk of cerebral amyloid angiopathy. We aimed to assess whether these variants also associate with severity of ICH, in terms of haematoma volume at presentation and subsequent outcome. METHODS: We investigated the association of APOE ε2 and ε4 with ICH volume and outcomes in patients with primary ICH in three phases: a discovery phase of 865 individuals of European ancestry from the Genetics of Cerebral Hemorrhage on Anticoagulation study, and replication phases of 946 Europeans (replication 1) and 214 African-Americans (replication 2) from an additional six studies. We also assessed the association of APOE variants with ICH volume and outcomes in meta-analyses of results from all three phases, and the association of APOE ε4 with mortality in a further meta-analysis including data from previous reports. Admission ICH volume was quantified on CT scan. We assessed functional outcome (modified Rankin scale score 3-6) and mortality at 90 days. We used linear regression to establish the effect of genotype on haematoma volume and logistic regression to assess the effect on outcome from ICH. FINDINGS: For patients with lobar ICH, carriers of the APOE ε2 allele had larger ICH volumes than did non-carriers in the discovery phase (p=2·5×10(-5)), in both replication phases (p=0·008 in Europeans and p=0·016 in African-Americans), and in the meta-analysis (p=3·2×10(-8)). In the meta-analysis, each copy of APOE ε2 increased haematoma size by a mean of 5·3 mL (95% CI 4·7-5·9; p=0·004). Carriers of APOE ε2 had increased mortality (odds ratio [OR] 1·50, 95% CI 1·23-1·82; p=2·45×10(-5)) and poorer functional outcomes (modified Rankin scale score 3-6; 1·52, 1·25-1·85; p=1·74×10(-5)) compared with non-carriers after lobar ICH. APOE ε4 was not associated with lobar ICH volume, functional outcome, or mortality in the discovery phase, replication phases, or meta-analysis of these three phases; in our further meta-analysis of 2194 patients, this variant did not increase risk of mortality (1·08, 0·86-1·36; p=0·52). APOE allele variants were not associated with deep ICH volume, functional outcome, or mortality. INTERPRETATION: Vasculopathic changes associated with the APOE ε2 allele might have a role in the severity and clinical course of lobar ICH. Screening of patients who have ICH to identify the ε2 variant might allow identification of those at increased risk of mortality and poor functional outcomes. FUNDING: US National Institutes of Health-National Institute of Neurological Disorders and Stroke, Keane Stroke Genetics Research Fund, Edward and Maybeth Sonn Research Fund, and US National Center for Research Resources.

Variants at APOE influence risk of deep and lobar intracerebral hemorrhage.

OBJECTIVE: Prior studies investigating the association between APOE alleles ε2/ε4 and risk of intracerebral hemorrhage (ICH) have been inconsistent and limited to small sample sizes, and did not account for confounding by population stratification or determine which genetic risk model was best applied. METHODS: We performed a large-scale genetic association study of 2189 ICH cases and 4041 controls from 7 cohorts, which were analyzed using additive models for ε2 and ε4. Results were subsequently meta-analyzed using a random effects model. A proportion of the individuals (322 cases, 357 controls) had available genome-wide data to adjust for population stratification. RESULTS: Alleles ε2 and ε4 were associated with lobar ICH at genome-wide significance levels (odds ratio [OR] = 1.82, 95% confidence interval [CI] = 1.50-2.23, p = 6.6 × 10(-10); and OR = 2.20, 95%CI = 1.85-2.63, p = 2.4 × 10(-11), respectively). Restriction of analysis to definite/probable cerebral amyloid angiopathy ICH uncovered a stronger effect. Allele ε4 was also associated with increased risk for deep ICH (OR = 1.21, 95% CI = 1.08-1.36, p = 2.6 × 10(-4)). Risk prediction evaluation identified the additive model as best for describing the effect of APOE genotypes. INTERPRETATION: APOE ε2 and ε4 are independent risk factors for lobar ICH, consistent with their known associations with amyloid biology. In addition, we present preliminary findings on a novel association between APOE ε4 and deep ICH. Finally, we demonstrate that an additive model for these APOE variants is superior to other forms of genetic risk modeling previously applied.

Proband race/ethnicity affects pedigree completion rate in a genetic study of ischemic stroke.

BACKGROUND: A recent meta-analysis suggested that racial/ethnic status is not a major determinant of willingness to participate in observational studies or treatment trials. However, little is known about the predictors of enrollment in family-based observational genetic studies. We tested the hypothesis that proband race/ethnicity is a significant predictor of enrolling a pedigree. METHODS: Univariable and multivariable logistic regression modeling was used to determine proband characteristics that predict DNA donation from both members of an affected sibling pair. A total of 619 adult male and female probands with first-time or recurrent ischemic stroke and a positive sibling history of stroke enrolled across 53 hospitals and clinics in the United States and Canada into the Siblings with Ischemic Stroke Study, a family-based prospective genomics study. RESULTS: In univariable analysis, probands with siblings who agreed to a blood draw for DNA analyses were more likely to be male and less likely to be nonwhite. In multivariable analysis, only race/ethnicity was significantly associated with likelihood of a proband's having a sibling who agreed to a blood draw. CONCLUSIONS: Contrary to observational studies that are not family based, the willingness of family members to participate in observational genetics studies may be influenced by race/ethnicity. This result reinforces the need for improving methods for recruiting diverse populations into genetic studies of stroke.

Impact of restricting enrollment in stroke genetics research to adults able to provide informed consent.

BACKGROUND AND PURPOSE: The extent of potential consent bias in observational studies elucidating genetic and environmental contributions to ischemic stroke is largely unknown. The purpose of this study was to assess differences in stroke cohort characteristics between those who provided informed consent and those whose enrollment was authorized by surrogate decision makers. METHODS: The Ischemic Stroke Genetics Study (ISGS) is a prospective, 5-center, case-control study of first-ever ischemic stroke. Demographic, clinical, and stroke characteristics were compared between cases enrolled by self versus by surrogate. Data from one site that limits enrollment only to those able to self-consent were also analyzed to compare those who enrolled with those not able to consent. RESULTS: Overall, 10% (54 of 517) were enrolled using surrogate authorization. Self-consented and surrogate-authorized cases did not differ significantly in age, sex, or conventional risk factors. Surrogate-authorized cases had significantly more severe stroke deficits, larger infarcts, and infarcts localizing to left supratentorial regions. Similarly, at the site restricting enrollment, stroke severity and characteristics differed between self-consented individuals and those otherwise eligible but unable to provide consent. CONCLUSIONS: Failure to permit surrogate authorization in genetic studies of ischemic stroke may skew enrollment toward less severe strokes caused by smaller infarcts. This potential consent bias may undermine the ability to identify genetic determinants of risk and severity and suggests that surrogate enrollment in these studies can be ethically justifiable.

Sex differences in stroke evaluations in the Ischemic Stroke Genetics Study.

BACKGROUND: Epidemiologic studies suggest sex differences in evaluation of patients presenting with ischemic stroke. Sex differences in stroke evaluation could lead to sex differences in the validity of diagnosing ischemic stroke subtypes. This study assessed sex differences in the Ischemic Stroke Genetics Study (ISGS). METHODS: The ISGS is a prospective case-control genetic association study of patients with first-ever ischemic stroke at 5 US tertiary stroke centers. The diagnostic tests performed as part of medical care were recorded for each enrolled patient. RESULTS: A total of 505 patients were enrolled; 45% (229 of 505) were women and 55% (276 of 505) were men. Mean age at time of stroke was greater for women (66.6 v 61.9 years; P = .001). Frequency of brain computed tomography was 92% (254 of 276) for men and 90% (206 of 229) for women (P = .42). Magnetic resonance imaging was completed in 84% (232 of 276) of men and 83% (191 of 229) of women (P = .91). Frequency of electrocardiography was 91% (252 of 276) for men and 90% (206 of 229) for women (P = .60). Echocardiography was done in 74% (203 of 276) of men and 79% (180 of 229) of women (P = .19). Cervical arterial imaging occurred in 91% (208 of 229) of women and 86% (237 of 276) of men (P = .09). Intracranial vascular imaging was performed in 75% (207 of 276) of men and 79% (181 of 229) of women (P = .28). CONCLUSIONS: Our findings suggest no significant sex differences in the extent to which major diagnostic tests were performed in patients in ISGS. Dedicated tertiary stroke centers may reduce the sex bias in stroke evaluation that has been identified by previous studies.