Machine learning approximations to predict epigenetic age acceleration in stroke patients

Age acceleration (Age-A) is a useful tool that is able to predict a broad range of health outcomes. It is necessary to determine DNA methylation levels to estimate it, and it is known that Age-A is influenced by environmental, lifestyle, and vascular risk factors (VRF). The aim of this study is to estimate the contribution of these easily measurable factors to Age-A in patients with cerebrovascular disease (CVD), using different machine learning (ML) approximations, and try to find a more accessible model able to predict Age-A. We studied a CVD cohort of 952 patients with information about VRF, lifestyle habits, and target organ damage. We estimated Age-A using Hannum\u27s epigenetic clock, and trained six different models to predict Age-A: a conventional linear regression model, four ML models (elastic net regression (EN), K-Nearest neighbors, random forest, and support vector machine models), and one deep learning approximation (multilayer perceptron (MLP) model). The best-performing models were EN and MLP; although, the predictive capability was modest (

Single-nucleus RNA-sequencing of autosomal dominant Alzheimer disease and risk variant carriers

Genetic studies of Alzheimer disease (AD) have prioritized variants in genes related to the amyloid cascade, lipid metabolism, and neuroimmune modulation. However, the cell-specific effect of variants in these genes is not fully understood. Here, we perform single-nucleus RNA-sequencing (snRNA-seq) on nearly 300,000 nuclei from the parietal cortex of AD autosomal dominant (APP and PSEN1) and risk-modifying variant (APOE, TREM2 and MS4A) carriers. Within individual cell types, we capture genes commonly dysregulated across variant groups. However, specific transcriptional states are more prevalent within variant carriers. TREM2 oligodendrocytes show a dysregulated autophagy-lysosomal pathway, MS4A microglia have dysregulated complement cascade genes, and APOEε4 inhibitory neurons display signs of ferroptosis. All cell types have enriched states in autosomal dominant carriers. We leverage differential expression and single-nucleus ATAC-seq to map GWAS signals to effector cell types including the NCK2 signal to neurons in addition to the initially proposed microglia. Overall, our results provide insights into the transcriptional diversity resulting from AD genetic architecture and cellular heterogeneity. The data can be explored on the online browser ( http://web.hararilab.org/SNARE/ )

Stroke genetics informs drug discovery and risk prediction across ancestries

Previous genome-wide association studies (GWASs) of stroke — the second leading cause of death worldwide — were conducted predominantly in populations of European ancestry1,2. Here, in cross-ancestry GWAS meta-analyses of 110,182 patients who have had a stroke (five ancestries, 33% non-European) and 1,503,898 control individuals, we identify association signals for stroke and its subtypes at 89 (61 new) independent loci: 60 in primary inverse-variance-weighted analyses and 29 in secondary meta-regression and multitrait analyses. On the basis of internal cross-ancestry validation and an independent follow-up in 89,084 additional cases of stroke (30% non-European) and 1,013,843 control individuals, 87% of the primary stroke risk loci and 60% of the secondary stroke risk loci were replicated (P < 0.05). Effect sizes were highly correlated across ancestries. Cross-ancestry fine-mapping, in silico mutagenesis analysis3, and transcriptome-wide and proteome-wide association analyses revealed putative causal genes (such as SH3PXD2A and FURIN) and variants (such as at GRK5 and NOS3). Using a three-pronged approach4, we provide genetic evidence for putative drug effects, highlighting F11, KLKB1, PROC, GP1BA, LAMC2 and VCAM1 as possible targets, with drugs already under investigation for stroke for F11 and PROC. A polygenic score integrating cross-ancestry and ancestry-specific stroke GWASs with vascular-risk factor GWASs (integrative polygenic scores) strongly predicted ischaemic stroke in populations of European, East Asian and African ancestry5. Stroke genetic risk scores were predictive of ischaemic stroke independent of clinical risk factors in 52,600 clinical-trial participants with cardiometabolic disease. Our results provide insights to inform biology, reveal potential drug targets and derive genetic risk prediction tools across ancestries

The copy number variation and stroke (CaNVAS) risk and outcome study

Background and purpose: The role of copy number variation (CNV) variation in stroke susceptibility and outcome has yet to be explored. The Copy Number Variation and Stroke (CaNVAS) Risk and Outcome study addresses this knowledge gap. Methods: Over 24,500 well-phenotyped IS cases, including IS subtypes, and over 43,500 controls have been identified, all with readily available genotyping on GWAS and exome arrays, with case measures of stroke outcome. To evaluate CNV-associated stroke risk and stroke outcome it is planned to: 1) perform Risk Discovery using several analytic approaches to identify CNVs that are associated with the risk of IS and its subtypes, across the age-, sex- and ethnicity-spectrums; 2) perform Risk Replication and Extension to determine whether the identified stroke-associated CNVs replicate in other ethnically diverse datasets and use biomarker data (e.g. methylation, proteomic, RNA, miRNA, etc.) to evaluate how the identified CNVs exert their effects on stroke risk, and lastly; 3) perform outcome-based Replication and Extension analyses of recent findings demonstrating an inverse relationship between CNV burden and stroke outcome at 3 months (mRS), and then determine the key CNV drivers responsible for these associations using existing biomarker data. Results: The results of an initial CNV evaluation of 50 samples from each participating dataset are presented demonstrating that the existing GWAS and exome chip data are excellent for the planned CNV analyses. Further, some samples will require additional considerations for analysis, however such samples can readily be identified, as demonstrated by a sample demonstrating clonal mosaicism. Conclusion: The CaNVAS study will cost-effectively leverage the numerous advantages of using existing case-control data sets, exploring the relationships between CNV and IS and its subtypes, and outcome at 3 months, in both men and women, in those of African and European-Caucasian descent, this, across the entire adult-age spectrum

Epigenome-wide association study identifies novel genes associated with ischemic stroke

Background: DNA methylation has previously been associated with ischemic stroke, but the specific genes and their functional roles in ischemic stroke remain to be determined. Here we aimed to identify differentially methylated genes that play a functional role in ischemic stroke in a Chinese population. Results: Genome-wide DNA methylation assessed with the Illumina Methylation EPIC Array in a discovery sample including 80 Chinese adults (40 cases vs. 40 controls) found that patients with ischemic stroke were characterized by increased DNA methylation at six CpG loci (individually located at TRIM6, FLRT2, SOX1, SOX17, AGBL4, and FAM84A, respectively) and decreased DNA methylation at one additional locus (located at TLN2). Targeted bisulfite sequencing confirmed six of these differentially methylated probes in an independent Chinese population (853 cases vs. 918 controls), and one probe (located at TRIM6) was further verified in an external European cohort (207 cases vs. 83 controls). Experimental manipulation of DNA methylation in engineered human umbilical vein endothelial cells indicated that the identified differentially methylated probes located at TRIM6, TLN2, and FLRT2 genes may play a role in endothelial cell adhesion and atherosclerosis. Conclusions: Altered DNA methylation of the TRIM6, TLN2, and FLRT2 genes may play a functional role in ischemic stroke in Chinese populations

Sex-related differences in primary intracerebral hemorrhage

OBJECTIVE: Little information is available about sex-related differences in intracerebral hemorrhage (ICH). This is a prospective observational study to describe the sex differences in demographics, vascular risk factors, stroke care, and outcomes in primary ICH. METHODS: BasicMar is a hospital-based registry of all stroke patients admitted to a single public hospital that covers a population of 330,000. From 2005 to 2015, there were 515 consecutive acute primary ICH patients. Outcome data were obtained at 3 months. RESULTS: More men than women had ICH (52.4% vs 47.6%); the women were older and had worse previous functional status than men, who were more likely to drink alcohol and smoke and to have ischemic heart disease and peripheral arterial disease. There were no sex differences in etiology, severity, or hemorrhage volume. ICH score was greater in women than in men (p = 0.018). Women had more lobar ICH than men (odds ratio adjusted by age was 1.75 [95% confidence interval 1.18-2.58], p = 0.005). The quality of stroke care was similar in both sexes. Mortality at 3 months was 44.1% in women and 41.1% in men (p = 0.656), and 3-month poor outcome among survivors (modified Rankin Scale [mRS] score 3-5) 58.4% in women and 45.3% in men (p = 0.027). After adjustment for previous mRS and ICH score, there were no differences in 3-month mortality or poor outcome at 3 months between sexes. CONCLUSIONS: Patients with ICH showed sex-related differences in demographic characteristics, ICH location, and vascular risk factors, but not in stroke care, 3-month mortality, or adjusted poor outcome

Sex-related differences in primary intracerebral hemorrhage

OBJECTIVE: Little information is available about sex-related differences in intracerebral hemorrhage (ICH). This is a prospective observational study to describe the sex differences in demographics, vascular risk factors, stroke care, and outcomes in primary ICH. METHODS: BasicMar is a hospital-based registry of all stroke patients admitted to a single public hospital that covers a population of 330,000. From 2005 to 2015, there were 515 consecutive acute primary ICH patients. Outcome data were obtained at 3 months. RESULTS: More men than women had ICH (52.4% vs 47.6%); the women were older and had worse previous functional status than men, who were more likely to drink alcohol and smoke and to have ischemic heart disease and peripheral arterial disease. There were no sex differences in etiology, severity, or hemorrhage volume. ICH score was greater in women than in men (p = 0.018). Women had more lobar ICH than men (odds ratio adjusted by age was 1.75 [95% confidence interval 1.18-2.58], p = 0.005). The quality of stroke care was similar in both sexes. Mortality at 3 months was 44.1% in women and 41.1% in men (p = 0.656), and 3-month poor outcome among survivors (modified Rankin Scale [mRS] score 3-5) 58.4% in women and 45.3% in men (p = 0.027). After adjustment for previous mRS and ICH score, there were no differences in 3-month mortality or poor outcome at 3 months between sexes. CONCLUSIONS: Patients with ICH showed sex-related differences in demographic characteristics, ICH location, and vascular risk factors, but not in stroke care, 3-month mortality, or adjusted poor outcome

Long-term cardiovascular prognosis after transient ischemic attack: Associated predictors

OBJECTIVE: To determine long-term cardiovascular risk after TIA and to identify the factors associated with increased risk. METHODS: This was a prospective observational registry of TIA patients admitted to the emergency room of our tertiary stroke center from June 2006 to January 2016. New vascular events (NVEs) were recorded from 3 months after TIA onset until June 2017, including both stroke and nonstroke events (coronary and peripheral disease). We registered TIA etiology, age, sex, vascular risk factors, radiologic data, and clinical TIA features and analyzed these variables in relation to NVE long-term risk. RESULTS: In total, 676 patients 71.7 ± 13.7 years of age were included, with a mean follow-up of 48.8 ± 32.7 months. An NVE was detected in 173 patients (25.6%) without significant differences between event types (p = 0.84). Univariate analysis associated NVEs with etiologic subgroup, male sex, diabetes mellitus, hypertension, previous vascular disease, duration and clinical features of TIA, and signs of acute infarction. Multivariable analysis showed an independent association of NVEs with etiologic TIA subgroup, signs of acute infarction, and duration of TIA symptoms. Large artery atherosclerosis and cardioaortic embolism had the highest NVE risk, with a slightly higher percentage of nonstroke events. The small artery disease subgroup had the lowest NVE risk, with a higher percentage of stroke events. CONCLUSIONS: Etiology subgroup was the main factor determining high long-term risk of vascular events in patients with TIA. Large artery atherosclerosis carried the highest vascular risk, both nonstroke and stroke, followed by cardioaortic embolism

Long-term cardiovascular prognosis after transient ischemic attack: Associated predictors

OBJECTIVE: To determine long-term cardiovascular risk after TIA and to identify the factors associated with increased risk. METHODS: This was a prospective observational registry of TIA patients admitted to the emergency room of our tertiary stroke center from June 2006 to January 2016. New vascular events (NVEs) were recorded from 3 months after TIA onset until June 2017, including both stroke and nonstroke events (coronary and peripheral disease). We registered TIA etiology, age, sex, vascular risk factors, radiologic data, and clinical TIA features and analyzed these variables in relation to NVE long-term risk. RESULTS: In total, 676 patients 71.7 ± 13.7 years of age were included, with a mean follow-up of 48.8 ± 32.7 months. An NVE was detected in 173 patients (25.6%) without significant differences between event types (p = 0.84). Univariate analysis associated NVEs with etiologic subgroup, male sex, diabetes mellitus, hypertension, previous vascular disease, duration and clinical features of TIA, and signs of acute infarction. Multivariable analysis showed an independent association of NVEs with etiologic TIA subgroup, signs of acute infarction, and duration of TIA symptoms. Large artery atherosclerosis and cardioaortic embolism had the highest NVE risk, with a slightly higher percentage of nonstroke events. The small artery disease subgroup had the lowest NVE risk, with a higher percentage of stroke events. CONCLUSIONS: Etiology subgroup was the main factor determining high long-term risk of vascular events in patients with TIA. Large artery atherosclerosis carried the highest vascular risk, both nonstroke and stroke, followed by cardioaortic embolism

Genomic and phenotypic insights from an atlas of genetic effects on DNA methylation

Characterizing genetic influences on DNA methylation (DNAm) provides an opportunity to understand mechanisms underpinning gene regulation and disease. In the present study, we describe results of DNAm quantitative trait locus (mQTL) analyses on 32,851 participants, identifying genetic variants associated with DNAm at 420,509 DNAm sites in blood. We present a database of >270,000 independent mQTLs, of which 8.5% comprise long-range (trans) associations. Identified mQTL associations explain 15-17% of the additive genetic variance of DNAm. We show that the genetic architecture of DNAm levels is highly polygenic. Using shared genetic control between distal DNAm sites, we constructed networks, identifying 405 discrete genomic communities enriched for genomic annotations and complex traits. Shared genetic variants are associated with both DNAm levels and complex diseases, but only in a minority of cases do these associations reflect causal relationships from DNAm to trait or vice versa, indicating a more complex genotype-phenotype map than previously anticipated