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Unrecognized myocardial infarction (MI) is an MI that remains undetected in the acute phase and is associated with an unfavourable prognosis. With this systematic review and meta-analysis, we evaluated the burden of cardiovascular risk factors in individuals with unrecognized MI. We searched general population-based cohort studies diagnosing unrecognized MI by electrocardiogram or myocardial imaging up to 24 November 2023. Pooled mean differences (MDs) or risk ratios (RRs) with 95% confidence intervals (CIs) were determined, and random-effects meta-analyses were performed. Fourteen cohort studies were included involving 200 450 individuals (mean age 62.8 ± 9.9 years, 56.0% women), among which 4322 (2.2%) experienced unrecognized MI (mean age 66.3 ± 8.2 years, 47.8% women) and 4653 (2.1%) recognized MI (mean age 68.5 ± 7.3 years, 33.8% women). Compared to individuals without MI, those with unrecognized MI had higher body mass index (MD 0.27, 95% CI 0.16-0.39) and systolic blood pressure (MD 4.48, 95% CI 2.81-6.15) levels, and higher prevalence of hypertension (RR 1.27, 95% CI 1.06-1.51) and diabetes mellitus (RR 1.67, 95% CI 1.36-2.06). Furthermore, individuals with unrecognized MI had lower prevalence of hypertension (RR 0.92, 95% CI 0.88-0.97) and diabetes mellitus (RR 0.80, 95% CI 0.70-0.92). Individuals with unrecognized MI are characterized by a substantial burden of metabolic risk factors. Our findings suggest insufficient recognition and management of cardiovascular risk factors among individuals with unrecognized MI., Competing Interests: Conflict of interest: J.R.v.L. received a research grant from Novartis paid to the institution. This funding source had no involvement in the study design, data collection, analysis, interpretation, writing, and submission. The other authors have no conflict of interest to declare., (© The Author(s) 2024. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Background: Cardiovascular disease is the leading cause of death in women worldwide. Yet, women are often underrepresented in cardiovascular clinical trials. Trial characteristics may influence the participation of women. For instance, trials are often entitled with an acronym, which might be perceived as gendered. We aimed to investigate if the perceived gender of the acronym and other trial characteristics affect the representation of female patients in cardiovascular trials., Methods: We searched ClinicalTrials.gov for randomized controlled trials in cardiovascular disease named with an acronym. Cardiovascular patients (n = 148) scored the perceived gender of the acronym of 148 identified trials. Prevalence ratios (PR) were calculated with Poisson regression to link trial characteristics to representation of female patients in the trials., Results: In 62 % of trials, female patients were underrepresented relative to the disease population. There was no improvement over time in proportion of trials with adequate representation. A third of acronyms was classified as gendered. The perceived gender did not affect representation of female patients (PR 1.01; 95% CI 0.95 - 1.08; P = 0.68). A woman as first and/or last author (PR 1.22; 95% CI 1.07 - 1.38; P = 0.002) and recruitment in an outpatient setting (PR 1.15; 95% CI 1.02 - 1.29; P = 0.01) were associated with a higher prevalence of adequate representation of female patients., Conclusions: Representation of female patients in cardiovascular trials does not depend on the perceived gender of the trial acronym but is improved in trials under female leadership in out-patient settings. Our findings may direct efforts towards increasing representation of female patients in cardiovascular trials., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors. Published by Elsevier B.V.)

Background: Left ventricular ejection fraction (EF) is used to categorize heart failure (HF) into phenotypes but this information is often missing in electronic health records or non-HF registries., Methods: We tested the applicability of a simplified version of a multivariable algorithm, that was developed on data of the Swedish Heart Failure Registry to predict EF in patients with HF. We used data from 4,868 patients with HF from the Cardiology Centers of the Netherlands database, an organization of 13 cardiac outpatient clinics that operate between the general practitioner and the hospital cardiologist. The algorithm included 17 demographical and clinical variables. We tested model discrimination, model performance and calculated model sensitivity, specificity, positive and negative predictive values for EF ≥ vs. <50% and EF ≥ vs. <40%. We additionally performed a multivariable multinomial analysis for all three separate HF phenotypes (with reduced, mildly reduced and preserved EF) HFrEF vs. HFmrEF vs. HFpEF. Finally, we internally validated the model by using temporal validation., Results: Mean age was 66 ±12 years, 44% of patients were women, 68% had HFpEF, 17% had HFrEF, and 15% had HFmrEF. The C-statistic was of 0.71 for EF ≥/< 50% (95% CI: 0.69-0.72) and of 0.74 (95% CI: 0.73-0.75) for EF ≥/< 40%. The model had the highest sensitivities for EF ≥50% (0.72, 95% CI: 0.63-0.75) and for EF ≥40% (0.70, 95% CI: 0.65-0.71). Similar results were achieved by the multinomial model, but the C-statistics for predicting HFpEF vs HFrEF was lower (0.61, 95% CI 0.58-0.63). The internal validation confirmed good discriminative ability., Conclusions: A simple algorithm based on routine clinical characteristics can help discern HF phenotypes in non-cardiology datasets and research settings such as research on primary care data, where measurements of EF is often not available., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Canto et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Complex health challenges require professionals to operate across disciplines and to better connect with society. Here, we showcase a community-engaged and challenge-based educational model in which undergraduate students conduct transdisciplinary research on authentic complex biomedical problems. This concept reinforces translational medicine, human capital, and exemplifies synergy between education, research, healthcare, and society., (© 2024 Schakelaar et al.)

Background and Aims: Tobacco smoking is a known risk factor for atherosclerotic disease, with more elevated risks in women compared to men. We hypothesized that atherosclerotic plaques from smokers show different gene expression patterns compared to non-smokers, in a sex-specific manner., Methods: Gene expression data of 625 carotid plaques (151 females and 474 males) were analyzed for differential gene expression between current smokers (n = 226) and non-smokers (n = 399). All analyses were stratified by sex and by molecular plaque characteristics. Finally, we projected the activity of gene regulatory networks and utilized single-cell transcriptomics from 38 plaques (26 males and 12 females) to interpret the sex- and plaque-type specific signals., Results: We observed higher expression levels of CRLF1 gene in atherosclerotic plaques from smokers compared to non-smokers (log2FC = 0.48, FDR = 0.012). CRLF1 upregulation was interacting with sex (p = 0.01) and was more pronounced in females (log2FC = 0.93, p = 1.53E-05) compared to males (log2FC = 0.35, p = 0.0018). Through single-cell RNA-seq analysis, we identified the highest CRLF1 expression within the transitioning and synthetic smooth muscle cell populations. CRLF1 expression was increased in fibro-inflammatory and fibro-cellular plaque types. Gene annotations pointed to increased expression of CRLF1 in networks with extracellular matrix related genes., Conclusions: Atherosclerotic plaques from current smokers show sex-dependent upregulation of smooth muscle cell gene CRLF1. This may explain the different contributions of smoking to cardiovascular risk in females., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Coronary vascular dysfunction comprises VSA and/or MVA and is more common in women than in men with angina without obstructive coronary artery disease (ANOCA). Invasive coronary function testing is considered the reference test for diagnosis, but its burden on patients is large. We aimed to investigate the potential of electrocardiography (ECG) as noninvasive marker for vasospastic angina (VSA) and microvascular angina (MVA) diagnosis., Methods: We systematically screened Pubmed and EMBASE databases for studies reporting on ECG characteristics in ANOCA patients with (a suspicion of) coronary vascular dysfunction. We assessed study quality using QUADAS-2. We extracted data on diagnostic values of different ECG characteristics and analyzed whether the studies were sex-stratified., Results: Thirty publications met our criteria, 13 reported on VSA and 17 on MVA. The majority addressed repolarization-related ECG parameters. Only 1 of the 13 VSA papers and 4 of the 17 MVA papers showed diagnostic accuracy measures of the ECG characteristics. The presence of early repolarization, T-wave alternans, and inverted U waves showed of predictive value for VSA diagnosis. The QTc interval was predictive for MVA diagnosis in all six studies reporting on QTc interval. Sex-stratified results were reported in only 5 of the 30 studies and 3 of those observed sex-based differences., Conclusions: ECG features are not widely evaluated in diagnostic studies for VSA and MVA. Those features predictive for VSA and MVA diagnosis mostly point to repolarization abnormalities and may contribute to noninvasive risk stratification., (© 2024 The Author(s). Annals of Noninvasive Electrocardiology published by Wiley Periodicals LLC.)

Background: Epigenetic age estimators (clocks) are predictive of human mortality risk. However, it is not yet known whether the epigenetic age of atherosclerotic plaques is predictive for the risk of cardiovascular events., Methods: Whole-genome DNA methylation of human carotid atherosclerotic plaques (n=485) and of blood (n=93) from the Athero-Express endarterectomy cohort was used to calculate epigenetic age acceleration (EAA). EAA was linked to clinical characteristics, plaque histology, and future cardiovascular events (n=136). We studied whole-genome DNA methylation and bulk and single-cell transcriptomics to uncover molecular mechanisms of plaque EAA. We experimentally confirmed our in silico findings using in vitro experiments in primary human coronary endothelial cells., Results: Male and female patients with severe atherosclerosis had a median chronological age of 69 years. The median epigenetic age was 65 years in females (median EAA, -2.2 [interquartile range, -4.3 to 2.2] years) and 68 years in males (median EAA, -0.3 [interquartile range, -2.9 to 3.8] years). Patients with diabetes and a high body mass index had higher plaque EAA. Increased EAA of plaque predicted future events in a 3-year follow-up in a Cox regression model (univariate hazard ratio, 1.7; P =0.0034) and adjusted multivariate model (hazard ratio, 1.56; P =0.02). Plaque EAA predicted outcome independent of blood EAA (hazard ratio, 1.3; P =0.018) and of plaque hemorrhage (hazard ratio, 1.7; P =0.02). Single-cell RNA sequencing in plaque samples from 46 patients in the same cohort revealed smooth muscle and endothelial cells as important cell types in plaque EAA. Endothelial-to-mesenchymal transition was associated with EAA, which was experimentally confirmed by TGFβ -triggered endothelial-to-mesenchymal transition inducing rapid epigenetic aging in coronary endothelial cells., Conclusions: Plaque EAA is a strong and independent marker of poor outcome in patients with severe atherosclerosis. Plaque EAA was linked to mesenchymal endothelial and smooth muscle cells. Endothelial-to-mesenchymal transition was associated with EAA, which was experimentally validated. Epigenetic aging mechanisms may provide new targets for treatments that reduce atherosclerosis complications., Competing Interests: Disclosures None.

Background: Thromboembolic events secondary to rupture or erosion of advanced atherosclerotic lesions is the global leading cause of death. The most common and effective means to reduce these major adverse cardiovascular events, including myocardial infarction and stroke, is aggressive lipid lowering via a combination of drugs and dietary modifications. However, we know little regarding the effects of reducing dietary lipids on the composition and stability of advanced atherosclerotic lesions, the mechanisms that regulate these processes, and what therapeutic approaches might augment the benefits of lipid lowering., Methods: Smooth muscle cell lineage-tracing Apoe -/- mice were fed a high-cholesterol Western diet for 18 weeks and then a zero-cholesterol standard laboratory diet for 12 weeks before treating them with an IL (interleukin)-1β or control antibody for 8 weeks. We assessed lesion size and remodeling indices, as well as the cellular composition of aortic and brachiocephalic artery lesions, indices of plaque stability, overall plaque burden, and phenotypic transitions of smooth muscle cell and other lesion cells by smooth muscle cell lineage tracing combined with single-cell RNA sequencing, cytometry by time-of-flight, and immunostaining plus high-resolution confocal microscopic z-stack analysis., Results: Lipid lowering by switching Apoe -/- mice from a Western diet to a standard laboratory diet reduced LDL cholesterol levels by 70% and resulted in multiple beneficial effects including reduced overall aortic plaque burden, as well as reduced intraplaque hemorrhage and necrotic core area. However, contrary to expectations, IL-1β antibody treatment after diet-induced reductions in lipids resulted in multiple detrimental changes including increased plaque burden and brachiocephalic artery lesion size, as well as increasedintraplaque hemorrhage, necrotic core area, and senescence as compared with IgG control antibody-treated mice. Furthermore, IL-1β antibody treatment upregulated neutrophil degranulation pathways but downregulated smooth muscle cell extracellular matrix pathways likely important for the protective fibrous cap., Conclusions: Taken together, IL-1β appears to be required for the maintenance of standard laboratory diet-induced reductions in plaque burden and increases in multiple indices of plaque stability., Competing Interests: Disclosures None.

Aims: We investigated the differences in prevalence of acute coronary syndrome (ACS) by presence versus absence of diabetes in males and females with chest discomfort who called out-of-hours primary care (OHS-PC)., Methods: A cross-sectional study performed in the Netherlands. Patients who called the OHS-PC in the Utrecht region, the Netherlands between 2014 and 2017 with acute chest discomfort were included. We compared those with diabetes with those without diabetes. Multivariable logistic regression was used to determine the relation between diabetes and (i) high urgency allocation and (ii) ACS., Results: Of the 2,195 callers with acute chest discomfort, 180 (8.2%) reported having diabetes. ACS was present in 15.3% of males (22.0% in those with diabetes) and 8.4% of females (18.8% in those with diabetes). Callers with diabetes did not receive a high urgency more frequently (74.4% vs. 67.8% (OR: 1.38; 95% CI 0.98-1.96). However, such callers had a higher odds for ACS (OR: 2.17; 95% CI 1.47-3.19). These differences were similar for females and males., Conclusions: Diabetes holds promise as diagnostic factor in callers to OHS-PC with chest discomfort. It might help triage in this setting given the increased risk of ACS in those with diabetes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

The rapid advancements in genome-scale (omics) techniques has created significant opportunities to investigate complex disease mechanisms in tissues and cells. Nevertheless, interpreting -omics data can be challenging, and pathway enrichment analysis is a frequently used method to identify candidate molecular pathways that drive gene expression changes. With a growing number of -omics studies dedicated to atherosclerosis, there has been a significant increase in studies and hypotheses relying on enrichment analysis. This brief review discusses the benefits and limitations of pathway enrichment analysis within atherosclerosis research. We highlight the challenges of identifying complex biological processes, such as cell phenotypic switching, within -omics data. Additionally, we emphasize the need for more comprehensive and curated gene sets that reflect the biological complexity of atherosclerosis. Pathway enrichment analysis is a valuable tool for gaining insights into the molecular mechanisms of atherosclerosis. Nevertheless, it is crucial to remain aware of the intrinsic limitations of this approach. By addressing these weaknesses, enrichment analysis in atherosclerosis can lead to breakthroughs in identifying the mechanisms of disease progresses, the identification of key driver genes, and consequently, advance personalized patient care., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: Chest discomfort and shortness of breath (SOB) are key symptoms in patients with acute coronary syndrome (ACS). It is, however, unknown whether SOB is valuable for recognising ACS during telephone triage in the out-of-hours primary care (OHS-PC) setting., Methods: A cross-sectional study performed in the Netherlands. Telephone triage conversations were analysed of callers with chest discomfort who contacted the OHS-PC between 2014 and 2017, comparing patients with SOB with those who did not report SOB. We determine the relation between SOB and (1) High urgency allocation, (2) ACS and (3) ACS or other life-threatening diseases., Results: Of the 2195 callers with chest discomfort, 1096 (49.9%) reported SOB (43.7% men, 56.3% women). In total, 15.3% men (13.2% in those with SOB) and 8.4% women (9.2% in those with SOB) appeared to have ACS. SOB compared with no SOB was associated with high urgency allocation (75.9% vs 60.8%, OR: 2.03; 95% CI 1.69 to 2.44, multivariable OR (mOR): 2.03; 95% CI 1.69 to 2.44), but not with ACS (10.9% vs 12.0%; OR: 0.90; 95% CI 0.69 to 1.17, mOR: 0.91; 95% CI 0.70 to 1.19) or 'ACS or other life-threatening diseases' (15.0% vs 14.1%; OR: 1.07; 95% CI 0.85 to 1.36, mOR: 1.09; 95% CI 0.86 to 1.38). For women the relation with ACS was 9.2% vs 7.5%, OR: 1.25; 95% CI 0.83 to 1.88, and for men 13.2% vs 17.4%, OR: 0.72; 95% CI 0.51 to 1.02. For 'ACS or other life-threatening diseases', this was 13.0% vs 8.5%, OR: 1.60; 95% CI 1.10 to 2.32 for women, and 7.5% vs 20.8%, OR: 0.81; 95% CI 0.59 to 1.12 for men., Conclusions: Men and women with chest discomfort and SOB who contact the OHS-PC more often receive high urgency than those without SOB. This seems to be adequate in women, but not in men when considering the risk of ACS or other life-threatening diseases., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Aims: We aim to investigate the association between kidney dysfunction and left ventricular diastolic dysfunction parameters and heart failure with preserved ejection fraction (HFpEF) and whether this is sex-specific., Methods and Results: We included participants from the HELPFul observational study. Outpatient clinical care data, including echocardiography, and an expert panel judgement on HFpEF was collected. Estimated glomerular filtration rate (eGFR) was calculated by creatinine and cystatin C without race. The association between eGFR with E/e', left ventricular mass index, relative wall thickness, and stage C/D heart failure was tested by multivariable adjusted regression models, stratified by sex, reporting odds ratios and 95% confidence intervals (95% confidence interval). We analysed 880 participants, mean age 62.9 (standard deviation: 9.3) years, 69% female. Four hundred six participants had mild (37.6%) kidney dysfunction (eGFR: 60-89 mL/min/1.73 m 2 ) or moderate (8.5%) kidney dysfunction (eGFR: 30-59 mL/min/1.73 m 2 ). HFpEF was significantly more prevalent in participants with mild and moderate kidney dysfunction (10.3% and 16.0%, respectively) than participants with normal kidney function (3.4%). A lower kidney function was associated with higher E/e' and higher relative wall thickness values. Participants with moderate kidney dysfunction had a higher likelihood of American College of Cardiology/American Heart Association stage C/D HF (odds ratio: 2.07, 95% confidence interval: 1.23, 3.49) than participants with normal kidney functions., Conclusions: Both mild and moderate kidney dysfunction are independently associated with left ventricular diastolic dysfunction parameters and HFpEF. This association is independent of sex and strongest for moderate kidney dysfunction. Considering mild-to-moderate kidney dysfunction as risk factor for HFpEF may help identify high-risk groups benefiting most from early intervention., (© 2023 The Authors. ESC Heart Failure published by John Wiley & Sons Ltd on behalf of European Society of Cardiology.)

Background: Coronary vasomotor dysfunction (CVDys) comprises coronary vasospasm (CVS) and/or coronary microvascular dysfunction (CMD) and is highly prevalent in patients with angina and non-obstructive coronary artery disease (ANOCA). Invasive coronary function testing (CFT) to diagnose CVDys is becoming more common, enabling pathophysiologic research of CVDys. This study aims to explore the electrophysiological characteristics of ANOCA patients with CVDys., Methods: We collected pre-procedural 12-lead electrocardiograms of ANOCA patients with CVS (n = 35), CMD (n = 24), CVS/CMD (n = 26) and patients without CVDys (CFT-, n = 23) who participated in the NL-CFT registry and underwent CFT. Heart axis and conduction times were compared between patients with CVS, CMD or CVS/CMD and patients without CVDys., Results: Heart axis, heart rate, PQ interval and QRS duration were comparable between the groups. A small prolongation of the QT-interval corrected with Bazett (QTcB) and Fridericia (QTcF) was observed in patients with CVDys compared to patients without CVDys (CVS vs CFT-: QTcB = 422 ± 18 vs 414 ± 18 ms (p = 0.14), QTcF = 410 ± 14 vs 406 ± 12 ms (p = 0.21); CMD vs CFT-: QTcB = 426 ± 17 vs 414 ± 18 ms (p = 0.03), QTcF = 413 ± 11 vs 406 ± 12 ms (p = 0.04); CVS/CMD vs CFT-: QTcB = 424 ± 17 vs 414 ± 18 ms (p = 0.05), QTcF = 414 ± 14 vs 406 ± 12 ms (p = 0.04))., Conclusions: Pre-procedural 12-lead electrocardiograms were comparable between patients with and without CVDys undergoing CFT except for a slightly longer QTc interval in patients with CVDys compared to patients without CVDys, suggesting limited cardiac remodeling in patients with CVDys., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

Background Plaque myofibroblasts are critical players in the initiation and advancement of atherosclerotic disease. They are involved in the production of extracellular matrix, the formation of the fibrous cap, and the underlying lipidic core via modulation processes in response to different environmental cues. Despite clear phenotypic differences between myofibroblast cells and healthy vascular smooth muscle cells, smooth muscle cells are still widely used as a cellular model in atherosclerotic research. Methods and Results Here, we present a conditioned outgrowth method to isolate and culture myofibroblast cells from plaques. We obtained these cells from 27 donors (24 carotid and 3 femoral endarterectomies). We show that they keep their proliferative capacity for 8 passages, are transcriptionally stable, retain donor-specific gene expression programs, and express extracellular matrix proteins ( FN1 , COL1A1 , and DCN ) and smooth muscle cell markers ( ACTA2 , MYH11 , and CNN1 ). Single-cell transcriptomics reveals that the cells in culture closely resemble the plaque myofibroblasts. Chromatin immunoprecipitation sequencing shows the presence of histone H3 lysine 4 dimethylation at the MYH11 promoter, pointing to their smooth muscle cell origin. Finally, we demonstrated that plaque myofibroblasts can be efficiently transduced (>97%) and are capable of taking up oxidized low-density lipoprotein and undergoing calcification. Conclusions In conclusion, we present a method to isolate and culture cells that retain plaque myofibroblast phenotypical and functional capabilities, making them a suitable in vitro model for studying selected mechanisms of atherosclerosis.

Sex differences in coronary artery disease (CAD) presentation, risk factors and prognosis have been widely studied. Similarly, studies on atherosclerosis have shown prominent sex differences in plaque biology. Our understanding of the underlying genetic and molecular mechanisms that drive these differences remains fragmented and largely understudied. Through reviewing genetic and epigenetic studies, we identified more than 40 sex-differential candidate genes (13 within known CAD loci) that may explain, at least in part, sex differences in vascular remodeling, lipid metabolism and endothelial dysfunction. Studies with transcriptomic and single-cell RNA sequencing data from atherosclerotic plaques highlight potential sex differences in smooth muscle cell and endothelial cell biology. Especially, phenotypic switching of smooth muscle cells seems to play a crucial role in female atherosclerosis. This matches the known sex differences in atherosclerotic phenotypes, with men being more prone to lipid-rich plaques, while women are more likely to develop fibrous plaques with endothelial dysfunction. To unravel the complex mechanisms that drive sex differences in CAD, increased statistical power and adjustments to study designs and analysis strategies are required. This entails increasing inclusion rates of women, performing well-defined sex-stratified analyses and the integration of multi-omics data., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: Genome-wide association studies for glycemic traits have identified hundreds of loci associated with these biomarkers of glucose homeostasis. Despite this success, the challenge remains to link variant associations to genes, and underlying biological pathways., Methods: To identify coding variant associations which may pinpoint effector genes at both novel and previously established genome-wide association loci, we performed meta-analyses of exome-array studies for four glycemic traits: glycated hemoglobin (HbA1c, up to 144,060 participants), fasting glucose (FG, up to 129,665 participants), fasting insulin (FI, up to 104,140) and 2hr glucose post-oral glucose challenge (2hGlu, up to 57,878). In addition, we performed network and pathway analyses., Results: Single-variant and gene-based association analyses identified coding variant associations at more than 60 genes, which when combined with other datasets may be useful to nominate effector genes. Network and pathway analyses identified pathways related to insulin secretion, zinc transport and fatty acid metabolism. HbA1c associations were strongly enriched in pathways related to blood cell biology., Conclusions: Our results provided novel glycemic trait associations and highlighted pathways implicated in glycemic regulation. Exome-array summary statistic results are being made available to the scientific community to enable further discoveries., Competing Interests: Competing interests: Rebecca S. Fine: Rebecca S. Fine is currently employed by Vertex Pharmaceuticals Incorporated. Audrey Y Chu: Currently employed by GlaxoSmithkline. Dennis O. Mook-Kanamori: Dennis Mook-Kanamori is working as a part-time clinical research consultant for Metabolon, Inc. Paul W. Franks: PWF has been a paid consultant for Eli Lilly and Sanofi Aventis and has received research support from several pharmaceutical companies as part of a European Union Innovative Medicines Initiative (IMI) project. Mike A. Nalls: Dr. Mike A. Nalls is supported by a consulting contract between Data Tecnica International LLC and the National Institute on Aging (NIA), National Institutes of Health (NIH), Bethesda, MD, USA. Dr. Nalls also consults for Illumina Inc., the Michael J. Fox Foundation, and the University of California Healthcare. Mark J. Caulfield: MJC is Chief Scientist for Genomics England, a UK government company. Joel N. Hirschhorn: JHN is on the scientific advisory board of Camp4 Therapeutics. Erik Ingelsson: Erik Ingelsson is now an employee of GlaxoSmithKline. Anubha Mahajan: Anubha Mahajan is an employee of Genentech since January 2020, and a holder of Roche stock. Mark I McCarthy: The views expressed in this article are those of the author(s) and not necessarily those of the NHS, the NIHR, or the Department of Health. MMcC has served on advisory panels for Pfizer, NovoNordisk and Zoe Global, has received honoraria from Merck, Pfizer, Novo Nordisk and Eli Lilly, and research funding from Abbvie, Astra Zeneca, Boehringer Ingelheim, Eli Lilly, Janssen, Merck, NovoNordisk, Pfizer, Roche, Sanofi Aventis, Servier, and Takeda. As of June 2019, MMcC is an employee of Genentech, and a holder of Roche stock. Inês Barroso: IB and spouse declare stock ownership in GlaxoSmithkline and Incyte Ltd. James B. Meigs: JBM serves as an Academic Associate for Quest Diagnostics R&D Bruce M Psaty serves on the Steering Committee of the Yale Open Data Access Project funded by Johnson & Johnson. Dr. Sander W. van der Laan has received Roche funding for unrelated work. Matthias Blüher received honoraria as a consultant and speaker from Amgen, AstraZeneca, Bayer, Boehringer-Ingelheim, Lilly, Novo Nordisk, Novartis, Pfizer and Sanofi. Vinicius Tragante: VT became an employee of deCODE genetics/Amgen Inc. after the conclusion of this work Dr Franco is employed by ErasmusAGE, a center for aging research across the life course funded by Nestlé Nutrition (Nestec Ltd.) and Metagenics., (Copyright: © 2023 Willems SM et al.)

The accumulation of erythrocyte membranes within an atherosclerotic plaque may contribute to the deposition of free cholesterol and thereby the enlargement of the necrotic core. Erythrocyte membranes can be visualized and quantified in the plaque by immunostaining for the erythrocyte marker glycophorin C. Hence, we theorized that the accumulation of erythrocytes quantified by glycophorin C could function as a marker for plaque vulnerability, possibly reflecting intraplaque hemorrhage (IPH), and offering predictive value for pre-procedural neurological symptoms. We employed the CellProfiler-integrated slideToolKit workflow to visualize and quantify glycophorin C, defined as the total plaque area that is positive for glycophorin C, in single slides of culprit lesions obtained from the Athero-Express Biobank of 1819 consecutive asymptomatic and symptomatic patients who underwent carotid endarterectomy. Our assessment included the evaluation of various parameters such as lipid core, calcifications, collagen content, SMC content, and macrophage burden. These parameters were evaluated using a semi-quantitative scoring method, and the resulting data was dichotomized as predefined criteria into categories of no/minor or moderate/heavy staining. In addition, the presence or absence of IPH was also scored. The prevalence of IPH and pre-procedural neurological symptoms were 62.4% and 87.1%, respectively. The amount of glycophorin staining was significantly higher in samples from men compared to samples of women (median 7.15 (IQR:3.37, 13.41) versus median 4.06 (IQR:1.98, 8.32), p < 0.001). Glycophorin C was associated with IPH adjusted for clinical confounders (OR 1.90; 95% CI 1.63, 2.21; p =  < 0.001). Glycophorin C was significantly associated with ipsilateral pre-procedural neurological symptoms (OR:1.27, 95%CI:1.06-1.41, p = 0.005). Sex-stratified analysis, showed that this was also the case for men (OR 1.37; 95%CI 1.12, 1.69; p = 0.003), but not for women (OR 1.15; 95%CI 0.77, 1.73; p = 0.27). Glycophorin C was associated with classical features of a vulnerable plaque, such as a larger lipid core, a higher macrophage burden, less calcifications, a lower collagen and SMC content. There were marked sex differences, in men, glycophorin C was associated with calcifications and collagen while these associations were not found in women. To conclude, the accumulation of erythrocytes in atherosclerotic plaque quantified and visualized by glycophorin C was independently associated with the presence of IPH, preprocedural symptoms in men, and with a more vulnerable plaque composition in both men and women. These results strengthen the notion that the accumulation of erythrocytes quantified by glycophorin C can be used as a marker for plaque vulnerability., (© 2023. Springer Nature Limited.)

Background: Women presenting with coronary artery disease more often present with fibrous atherosclerotic plaques, which are currently understudied. Phenotypically modulated smooth muscle cells (SMCs) contribute to atherosclerosis in women. How these phenotypically modulated SMCs shape female versus male plaques is unknown., Methods: Gene regulatory networks were created using RNAseq gene expression data from human carotid atherosclerotic plaques. The networks were prioritized based on sex bias, relevance for smooth muscle biology, and coronary artery disease genetic enrichment. Network expression was linked to histologically determined plaque phenotypes. In addition, their expression in plaque cell types was studied at single-cell resolution using single-cell RNAseq. Finally, their relevance for disease progression was studied in female and male Apoe -/- mice fed a Western diet for 18 and 30 weeks., Results: Here, we identify multiple sex-stratified gene regulatory networks from human carotid atherosclerotic plaques. Prioritization of the female networks identified 2 main SMC gene regulatory networks in late-stage atherosclerosis. Single-cell RNA sequencing mapped these female networks to 2 SMC phenotypes: a phenotypically modulated myofibroblast-like SMC network and a contractile SMC network. The myofibroblast-like network was mostly expressed in plaques that were vulnerable in women. Finally, the mice ortholog of key driver gene MFGE8 (milk fat globule EGF and factor V/VIII domain containing) showed retained expression in advanced plaques from female mice but was downregulated in male mice during atherosclerosis progression., Conclusions: Female atherosclerosis is characterized by gene regulatory networks that are active in fibrous vulnerable plaques rich in myofibroblast-like SMCs., Competing Interests: Disclosures None.

Coronary artery calcification (CAC), a measure of subclinical atherosclerosis, predicts future symptomatic coronary artery disease (CAD). Identifying genetic risk factors for CAC may point to new therapeutic avenues for prevention. Currently, there are only four known risk loci for CAC identified from genome-wide association studies (GWAS) in the general population. Here we conducted the largest multi-ancestry GWAS meta-analysis of CAC to date, which comprised 26,909 individuals of European ancestry and 8,867 individuals of African ancestry. We identified 11 independent risk loci, of which eight were new for CAC and five had not been reported for CAD. These new CAC loci are related to bone mineralization, phosphate catabolism and hormone metabolic pathways. Several new loci harbor candidate causal genes supported by multiple lines of functional evidence and are regulators of smooth muscle cell-mediated calcification ex vivo and in vitro. Together, these findings help refine the genetic architecture of CAC and extend our understanding of the biological and potential druggable pathways underlying CAC., (© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Background: Microvascular dysfunction plays a crucial role in complications of type 2 diabetes and might contribute to heart failure with preserved ejection fraction (HFpEF), a disease that disproportionally affects women. We aimed to investigate if presence and degree of microvascular dysfunction (MVD) in skin relates to markers of left ventricular diastolic dysfunction (LVDD) and HFpEF risk in adults with type 2 diabetes, and whether sex modifies this association., Methods: We recruited 154 participants (50% women) from the Hoorn Diabetes Care System Cohort, a prospective cohort study, for in vivo evaluation of skin MVD, echocardiography and blood sampling. MVD was assessed by laser speckle contrast analysis combined with iontophoresis of insulin, acetylcholine and sodium nitroprusside (SNP). We performed a cross-sectional analysis of the association between perfusion responses and echocardiographic and clinical markers of LVDD and the H2FPEF score by multivariable linear regression analysis adjusted for confounders. Sex was evaluated as a potential effect modifier and the analysis was stratified., Results: Mean age was 67 ± 6y, mean HbA1c 7.6 ± 1.3%. Women were more frequently obese (54.5 vs. 35.1%), had higher NT-proBNP plasma levels (80, IQR:34-165 vs. 46, 27-117 pg/ml) and E/E'(13.3 ± 4.3 vs. 11.4 ± 3.0) than men. Eleven women and three men were diagnosed with HFpEF, and showed lower perfusion response to insulin than those without HFpEF. A lower perfusion response to insulin and acetylcholine was associated with higher HFpEF risk in women, but not men (10% decreased perfusion response was associated with 5.8% [95%CI: 2.3;9.4%] and 5.9% [1.7;10.1%] increase of the H2FPEF score, respectively). A lower perfusion response to SNP was associated with higher pulmonary arterial systolic pressure in men while a lower perfusion response to acetylcholine associated with higher LV mass index in women and with worse LV longitudinal strain in the total population. No significant associations were found between perfusion responses and conventional LVDD markers., Conclusions: Impaired microvascular responses to insulin and acetylcholine in skin confers a higher risk of HFpEF in women with type 2 diabetes. In vivo measures of systemic MVD could represent novel risk markers for HFpEF, opening new avenues for the prevention of HFpEF in type 2 diabetes., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Hypertension is a key modifiable risk factor for cardiovascular disease. Several observational studies have found a stronger association of blood pressure and cardiovascular disease risk in women compared to men. Since observational studies can be affected by sex-specific residual confounding and reverse causation, it remains unclear whether these differences reflect actual differential effects. Other study designs are needed to uncover the causality of sex differences in the strength of risk factor and treatment effects. Mendelian randomisation (MR) uses genetic variants as instrumental variables to provide evidence about putative causal relations between risk factors and outcomes. By exploiting the random allocation of genes at gamete forming, MR is unaffected by confounding and results in more reliable causal effect estimates. In this review, we discuss why and how sex-specific MR and cis-MR could be used to study sex differences in risk factor and drug target effects. Sex-specific MR can be helpful to strengthen causal inferences in the field of sex differences, where it is often challenging to distinguish nature from nurture. The challenge of sex-specific (drug target) MR lays in leveraging robust genetic instruments from sex-specific GWAS studies which are not commonly available. Knowledge on sex-specific causal effects of hypertension, or other risk factors, could improve clinical practice and health policies by tailoring interventions based on personalised risk. Drug target MR can help to determine the anticipated on-target effects of a drug compound and to identify targets to pursue in drug development., (© 2023. The Author(s).)

Background and Aim: Sex differences in atherosclerosis have been described with female plaques being mostly perceived as stable and fibrous. Sex-specific mechanisms such as mosaic loss of the Y chromosome in men have been linked to cardiovascular health. In women, X-linked mechanisms such as X chromosome inactivation (XCI) skewing is common in several tissues. Yet, information on the role of XCI in female atherosclerotic plaques is lacking. Here, we investigated the presence of XCI skewing in advanced atherosclerotic lesions and its association with cardiovascular risk factors, histological plaque data, and clinical data., Methods: XCI skewing was quantified in 154 atherosclerotic plaque and 55 blood DNA samples of women included in the Athero-Express study. The skewing status was determined performing the HUMARA assay. Then, we studied the relationship of XCI skewing in female plaque and cardiovascular risk factors using regression models. In addition, we studied if plaque XCI predicted plaque composition, and adverse events during 3-years follow-up using Cox proportional hazard models., Results: XCI skewing was detected in 76 of 154 (49.4%) plaques and in 27 of 55 (67%) blood samples. None of the clinical risk factors were associated with plaque skewing. Plaque skewing was more often detected in plaques with a plaque hemorrhage (OR [95% CI]: 1.44 [1.06-1.98], P = 0.02). Moreover, skewed plaques were not associated with a higher incidence of composite and major events but were specifically associated with peripheral artery events during a 3-year follow-up period in a multivariate model (HR [95%CI]: 1.46 [1.09-1.97]; P = 0.007)., Conclusions: XCI skewing is common in carotid plaques of females and is predictive for the occurrence of peripheral artery events within 3 years after carotid endarterectomy., (© 2023. The Author(s).)

Background: Concentric remodeling (cRM) can precede heart failure with preserved ejection fraction (HFpEF), a condition prevalent in women., Methods: Patients (n=60 593, 54.2% women) visiting outpatient clinics of Cardiology Centers of the Netherlands were analyzed for cRM, HFpEF development, and mortality risk. We studied risk factors for relative wall thickness both sex-stratified and in women and men combined. Biomarker profiling was performed (4534 plasma proteins) in a substudy involving 557 patients (65.4% women) to identify pathways involved in cRM., Results: cRM was present in 23.5% of women and 27.6% of men and associated with developing HFpEF (HR, 2.15 [95% CI, 1.51-2.99]) and mortality risk (HR, 1.09 [95% CI, 1.00-1.19]) in both sexes. Age, heart rate, and hypertension were statistically significantly stronger risk factors for relative wall thickness in women than men. Higher circulating levels of IFNA5 (interferon alpha-5) were associated with higher relative wall thickness in women only. Pathway analysis revealed differential pathway activation by sex and increased expression of inflammatory pathways in women., Conclusions: cRM is prevalent in approximately 1 in 4 women and men visiting outpatient cardiology clinics and associated with HFpEF development and mortality risk in both sexes. Known risk factors for cRM were more strongly associated in women than men. Proteomic analysis revealed inflammatory pathway activation in women, with a central role for IFNA5. Differential biologic pathway activation by sex in cRM may contribute to the female predominance of HFpEF and holds promise for identification of new therapeutic avenues for prevention and treatment of HFpEF., Registration: URL: https://www., Clinicaltrials: gov; Unique identifier: NCT001747., Competing Interests: Disclosures None.

Background: Angina without angiographic evidence of obstructive coronary artery disease (ANOCA) is a highly prevalent condition with insufficient pathophysiological knowledge and lack of evidence-based medical therapies. This affects ANOCA patients prognosis, their healthcare utilization and quality of life. In current guidelines, performing a coronary function test (CFT) is recommended to identify a specific vasomotor dysfunction endotype. The NetherLands registry of invasive Coronary vasomotor Function testing (NL-CFT) has been designed to collect data on ANOCA patients undergoing CFT in the Netherlands., Methods: The NL-CFT is a web-based, prospective, observational registry including all consecutive ANOCA patients undergoing clinically indicated CFT in participating centers throughout the Netherlands. Data on medical history, procedural data and (patient reported) outcomes are gathered. The implementation of a common CFT protocol in all participating hospitals promotes an equal diagnostic strategy and ensures representation of the entire ANOCA population. A CFT is performed after ruling out obstructive coronary artery disease. It comprises of both acetylcholine vasoreactivity testing as well as bolus thermodilution assessment of microvascular function. Optionally, continuous thermodilution or Doppler flow measurements can be performed. Participating centers can perform research using own data, or pooled data will be made available upon specific request via a secure digital research environment, after approval of a steering committee., Conclusion: NL-CFT will be an important registry by enabling both observational and registry based (randomized) clinical trials in ANOCA patients undergoing CFT., Competing Interests: Declaration of Competing Interest NL-CFT is supported by research grants from Dutch Cardiovascular Alliance consortium IMPRESS (2020B004), Philips and Abbott. Damman P received research grants, lecture and consultancy fees from Phillips, and research grants and lecture fees from Abbott Vascular. Van Royen N has received research grants from Philips, Biotronik, Medtronic and Abbott and speaker fees from Microport, Rainmed and Abbott. Van ‘t Hof A received research grants and lecture fees from Abbott Vascular, Medtronic, Boehringer Ingelheim, Astra Zeneca and AMGEN not related to this work. Van de Hoef T received institutional research grants and consultancy fees from Philips and Abbott Vascular. Appelman Y received research grant from Dutch Heart Foundation IMPRESS, lecture and consultancy fees from Abbott Vascular and Phillips. Woudstra P received lecture fees from Philips. All other authors have no disclosures., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Aims: Atrial fibrillation (AF) is becoming increasingly common. Traditional cardiovascular risk factors (CVRF) do not explain all AF cases. Blood-based biomarkers reflecting cardiac injury such as high-sensitivity troponin I (hsTnI) may help close this gap., Methods: We investigated the predictive ability of hsTnI for incident AF in 45,298 participants (median age 51.4 years, 45.0% men) across European community cohorts in comparison to CVRF and established biomarkers (C-reactive protein, N-terminal pro B-type natriuretic peptide)., Results: During a median follow-up of 7.7 years, 1734 (3.8%) participants developed AF. Those in the highest hsTnI quarter (≥4.2 ng/L) had a 3.91-fold (95% confidence interval (CI) 3.30, 4.63; p < .01) risk for developing AF compared to the lowest quarter (<1.4 ng/L). In multivariable-adjusted Cox proportional hazards models a statistically significant association was seen between hsTnI and AF (hazard ratio (HR) per 1 standard deviation (SD) increase in log10(hsTnI) 1.08; 95% CI 1.01, 1.16; p = .03). Inclusion of hsTnI did improve model discrimination (C-index CVRF 0.811 vs. C-index CVRF and hsTnI 0.813; p < .01). Higher hsTnI concentrations were associated with heart failure (HR per SD 1.37; 95% CI 1.12, 1.68; p < .01) and overall mortality (HR per SD 1.24; 95% CI 1.09, 1.41; p < .01)., Conclusion: hsTnI as a biomarker of myocardial injury does not improve prediction of AF incidence beyond classical CVRF and NT-proBNP. However, it is associated with the AF-related disease heart failure and mortality likely reflecting underlying subclinical cardiovascular impairment., (© 2023 The Authors. European Journal of Clinical Investigation published by John Wiley & Sons Ltd on behalf of Stichting European Society for Clinical Investigation Journal Foundation.)

Background: Coronary artery spasm (CAS), encompassing epicardial and microvascular spasm, is increasingly recognized as cause of angina in patients with non-obstructive coronary artery disease (ANOCA). However, various spasm provocation testing protocols and diagnostic criteria are used, making diagnosis and characterization of these patients difficult and interpretation of study results cumbersome. This review provides a structured overview of the prevalence, characterization and prognosis of CAS worldwide in men and women., Methods: A systematic review identifying studies describing ANOCA patients with CAS was performed. Multiple outcomes (prevalence, clinical features, and prognosis) were assessed. Data, except for prognosis were pooled and analysed using random effects meta-analysis models., Results: Twenty-five publications ( N  = 14.554) were included (58.2 years; 44.2% women). Percentages of epicardial constriction to define epicardial spasm ranged from >50% to >90%. Epicardial spasm was prevalent in 43% (range 16-73%), with a higher prevalence in Asian vs. Western World population (52% vs. 33%, p  = 0.014). Microvascular spasm was prevalent in 25% (range 7-39%). Men were more likely to have epicardial spasm (61%), women were more likely to have microvascular spasm (64%). Recurrent angina is frequently reported during follow-up ranging from 10 to 53%., Conclusion: CAS is highly prevalent in ANOCA patients, where men more often have epicardial spasm, women more often have microvascular spasm. A higher prevalence of epicardial spasm is demonstrated in the Asian population compared to the Western World. The prevalence of CAS is high, emphasizing the use of unambiguous study protocols and diagnostic criteria and highlights the importance of routine evaluation of CAS in men and women with ANOCA., Systematic Review Registration: https://www.crd.york.ac.uk/prospero/display&#95;record.php?RecordID=272100., Competing Interests: TH received an institutional research grant from Philips. JP is consultant for Philips. US and PO received honoraria from Abbott and Philips for speaking about coronary function testing. MB received an institutional research grant from Actelion B.V. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 Woudstra, Vink, Schipaanboord, Eringa, den Ruijter, Feenstra, Boerhout, Beijk, de Waard, Ong, Seitz, Sechtem, Piek, van de Hoef and Appelman.)

Background: Coronary artery disease (CAD) is the leading cause of death worldwide. Recent meta-analyses of genome-wide association studies have identified over 175 loci associated with CAD. The majority of these loci are in noncoding regions and are predicted to regulate gene expression. Given that vascular smooth muscle cells (SMCs) play critical roles in the development and progression of CAD, we aimed to identify the subset of the CAD loci associated with the regulation of transcription in distinct SMC phenotypes., Methods: We measured gene expression in SMCs isolated from the ascending aortas of 151 heart transplant donors of various genetic ancestries in quiescent or proliferative conditions and calculated the association of their expression and splicing with ~6.3 million imputed single-nucleotide polymorphism markers across the genome., Results: We identified 4910 expression and 4412 splicing quantitative trait loci (sQTLs) representing regions of the genome associated with transcript abundance and splicing. A total of 3660 expression quantitative trait loci (eQTLs) had not been observed in the publicly available Genotype-Tissue Expression dataset. Further, 29 and 880 eQTLs were SMC-specific and sex-biased, respectively. We made these results available for public query on a user-friendly website. To identify the effector transcript(s) regulated by CAD loci, we used 4 distinct colocalization approaches. We identified 84 eQTL and 164 sQTL that colocalized with CAD loci, highlighting the importance of genetic regulation of mRNA splicing as a molecular mechanism for CAD genetic risk. Notably, 20% and 35% of the eQTLs were unique to quiescent or proliferative SMCs, respectively. One CAD locus colocalized with a sex-specific eQTL ( TERF2IP ), and another locus colocalized with SMC-specific eQTL ( ALKBH8 ). The most significantly associated CAD locus, 9p21, was an sQTL for the long noncoding RNA CDKN2B-AS1 , also known as ANRIL , in proliferative SMCs., Conclusions: Collectively, our results provide evidence for the molecular mechanisms of genetic susceptibility to CAD in distinct SMC phenotypes.

Women have historically been underrepresented in cardiovascular clinical trials, resulting in a lack of sex-specific data. This is especially problematic in two situations, namely those where diseases manifest differently in women and men and those where biological differences between the sexes might affect the efficacy and/or safety of medication. There is therefore a pressing need for datasets with proper representation of women to address questions related to these situations. Clinical care data could fit this bill nicely because of their unique broad scope across both patient groups and clinical measures. This perspective piece presents the potential of clinical care data in sex differences research and discusses current challenges clinical care data-based research faces. It also suggests strategies to reduce the effect of these limitations, and explores whether clinical care data alone will be sufficient to close evidence gaps or whether a more comprehensive approach is needed., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 Bots, Onland-Moret and den Ruijter.)

Histopathological studies have revealed key processes of atherosclerotic plaque thrombosis. However, the diversity and complexity of lesion types highlight the need for improved sub-phenotyping. Here we analyze the gene expression profiles of 654 advanced human carotid plaques. The unsupervised, transcriptome-driven clustering revealed five dominant plaque types. These plaque phenotypes were associated with clinical presentation and showed differences in cellular compositions. Validation in coronary segments showed that the molecular signature of these plaques was linked to coronary ischemia. One of the plaque types with the most severe clinical symptoms pointed to both inflammatory and fibrotic cell lineages. Further, we did a preliminary analysis of potential circulating biomarkers that mark the different plaques phenotypes. In conclusion, the definition of the plaque at risk for a thrombotic event can be fine-tuned by in-depth transcriptomic-based phenotyping. These differential plaque phenotypes prove clinically relevant for both carotid and coronary artery plaques and point to distinct underlying biology of symptomatic lesions., Competing Interests: CLM has received funding support from AstraZeneca for work unrelated to this study. GP received funding support from Roche to partly cover the generation of biomarker data. The remaining authors declare no competing interests.

Objective: To assess the prognostic value of absolute and sex-specific, age-specific and race/ethnicity-specific (Multi-Ethnic Study of Atherosclerosis, MESA) percentiles of coronary artery calcification in symptomatic women and men., Methods: The study population consisted of 4985 symptomatic patients (2793 women, 56%) visiting a diagnostic outpatient cardiology clinic between 2009 and 2018 who were referred for cardiac CT to determine Coronary Artery Calcium Score (CACS). Regular care data were used and these data were linked to the databases of Statistics Netherlands for all-cause mortality data. Kaplan-Meier curves, multivariate Cox proportional hazards regression and concordance statistics were used to evaluate the prognostic value of CACS and MESA percentiles. Women were older compared with men (60 vs 59 years)., Results: Median CACS was 0 (IQR: 0-54) in women and 42 (IQR: 0-54) in men. After a median follow-up of 4.4 years (IQR: 3.1-6.3), 116 (2.3%; 53 women and 63 men) patients died. MESA percentiles did not perform better compared with absolute CACS (C-statistic 0.65, 95% CI 0.57 to 0.73, vs 0.66, 95% CI 0.58 to 0.74, in women and 0.59, 95% CI 0.51 to 0.67, vs 0.62, 95% CI 0.55 to 0.69, in men, for the percentiles and absolute CACS, respectively)., Conclusions: In symptomatic individuals absolute CACS predicts mortality with a moderately good performance. MESA percentiles did not perform better compared with absolute CACS, thus there is no need to use them. Including degree of stenosis in the model might slightly improve mortality risk prediction in women, but not in men., Competing Interests: Competing interests: LH, IIT and GAS are employed by Cardiology Centers of the Netherlands. All other authors declare no competing interests., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Aims: Microvascular inflammation plays an important role in the pathogenesis of diastolic dysfunction (DD) and metabolic heart disease. NOX1 is expressed in vascular and immune cells and has been implicated in the vascular pathology of metabolic disease. However, its contribution to metabolic heart disease is less understood., Methods and Results: NOX1-deficient mice (KO) and male wild-type (WT) littermates were fed a high-fat high-sucrose diet (HFHS) and injected streptozotocin (75 mg/kg i.p.) or control diet (CTD) and sodium citrate. Despite similar weight gain and increase in fasting blood glucose and insulin, only WT-HFHS but not KO-HFHS mice developed concentric cardiac hypertrophy and elevated left ventricular filling pressure. This was associated with increased endothelial adhesion molecule expression, accumulation of Mac-2-, IL-1β-, and NLRP3-positive cells and nitrosative stress in WT-HFHS but not KO-HFHS hearts. Nox1 mRNA was solidly expressed in CD45+ immune cells isolated from healthy mouse hearts but was negligible in cardiac CD31+ endothelial cells. However, in vitro, Nox1 expression increased in response to lipopolysaccharide (LPS) in endothelial cells and contributed to LPS-induced upregulation of Icam-1. Nox1 was also upregulated in mouse bone marrow-derived macrophages in response to LPS. In peripheral monocytes from age- and sex-matched symptomatic patients with and without DD, NOX1 was significantly higher in patients with DD compared to those without DD., Conclusions: NOX1 mediates endothelial activation and contributes to myocardial inflammation and remodelling in metabolic disease in mice. Given its high expression in monocytes of humans with DD, NOX1 may represent a potential target to mitigate heart disease associated with DD., Competing Interests: Conflict of interest: none declared., (© The Author(s) 2021. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Objective: To develop and validate a symptom-based prediction rule for early recognition of acute coronary syndrome (ACS) in patients with acute chest discomfort who call out-of-hours services for primary care (OHS-PC)., Design: Cross-sectional study. A diagnostic prediction rule was developed with multivariable regression analyses. All models were validated with internal-external cross validation within seven OHS-PC locations. Both age and sex were analysed as statistical interaction terms, applying for age non-linear effects., Setting: Seven OHS-PC in the Netherlands., Participants: 2192 patients who called OHS-PC for acute chest discomfort (pain, pressure, tightness or discomfort) between 2014 and 2017. Backed up recordings of telephone triage conversations were analysed., Primary and Secondary Outcomes Measures: Diagnosis of ACS retrieved from the patient's medical records in general practice, including hospital specialists discharge letters. Performance of the prediction rules was calculated with the c-statistic and the final model was chosen based on net benefit analyses., Results: Among the 2192 patients who called the OHS-PC with acute chest discomfort, 8.3% females and 15.3% males had an ACS. The final diagnostic model included seven predictors (sex, age, acute onset of chest pain lasting less than 12 hours, a pressing/heavy character of the pain, radiation of the pain, sweating and calling at night). It had an adjusted c-statistic of 0.77 (95% CI 0.74 to 0.79) with good calibration., Conclusion: The final prediction model for ACS has good discrimination and calibration and shows promise for replacing the existing telephone triage rules for patients with acute chest discomfort in general practice and OHS-PC., Trial Registration Number: NTR7331., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Objectives: To investigate the impact of a CT-first strategy on all-cause and cardiovascular mortality in patients presenting with chest pain in outpatient cardiology clinics., Methods: Patients with a first presentation of suspected angina pectoris were identified and their data linked to the registrations of Statistics Netherlands for information on mortality. The linked database consisted of 33 068 patients. CT-first patients were defined as patients with a CT calcium score and coronary CT angiography, within 6 weeks after their initial visit. Propensity score matching (1:5) was used to match patients with and without a CT-first strategy. After matching, 12 545 patients were included of which 2308 CT-first patients and 10 237 patients that underwent usual care., Results: Mean age was 57 years, 56.3% were women and median follow-up was 4.9 years. All-cause mortality was significantly lower in CT-first patients (n=43, 1.9%) compared with patients without CT (n=363, 3.5%) (HR: 0.51, 95% CI 0.37 to 0.70). Furthermore, CT-first patients were more likely to receive cardiovascular preventative and antianginal medication (aspirin: 44.9% vs 27.1%, statins: 48.7% vs 30.3%, beta-blockers: 37.8% vs 25.5%, in CT-first and without CT-first patients, respectively) and to undergo downstream diagnostics and interventions (coronary interventions: 8.5% vs 5.7%, coronary angiography: 16.2% vs 10.6% in CT-first and without CT-first patients, respectively)., Conclusions: In a real-world regular care database, a CT-first strategy in patients suspected of angina pectoris was associated with a lowering of all-cause mortality., Competing Interests: Competing interests: LH, IIT and GAS are employed by Cardiology Centres of the Netherlands., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: The New York Heart Association (NYHA) functional class has extensively been used for risk stratification in patients suspected of heart failure, although its prognostic value differs between sexes and disease entities. Functional exercise capacity might explain the association between NYHA functional class and survival, and can serve as an objective proxy for the subjective nature of the NYHA classification. Therefore, we assessed whether sex-differences in exercise capacity explain the association between NYHA functional class and survival in patients suspected of cardiovascular disease., Methods: Electronic health record data from 7259 patients with cardiovascular symptoms, a documented NYHA functional class and cardiac stress electrocardiogram (ECG), was analysed. Follow-up for all-cause mortality was obtained through linkage with Statistics Netherlands. A sex-stratified mediation analysis was performed to assess to what extent the proportional heart rate and -workload during ECG stress testing explain the association between NYHA functional class and survival., Results: In men, increments in NYHA functional class were related to higher all-cause mortality in a dose-response manner (NYHA II vs III/IV: hazard ratio [HR] 1.59 vs 3.64, referenced to NYHA I), whilst in women those classified as NYHA functional class II and III/IV had a similar higher mortality risk (HR 1.49 vs 1.41). Sex-stratified mediation analysis showed that the association between NYHA and survival was mostly explained by proportional workload during stress ECG (men vs women: 22.9%, 95% CI: 18.9%-27.3% vs 40.3%, 95% CI: 28.5%-68.6%) and less so by proportional heart rate (men vs women: 2.5%, 95% CI: 1.3%-4.3% vs 8.0%, 95% CI: 4.1%-18.1%). Post-hoc analysis showed that NYHA classification explained a minor proportion of the association between proportional workload and all-cause mortality (men vs women: 15.1%, 95% CI: 12.0%-18.3% vs 4.4%, 95% CI: 1.5%-7.4%)., Conclusions: This study showed a significant mediation in both sexes on the association between NYHA functional class and all-cause mortality by proportional workload, but the effect explained by NYHA classification on the association between survival and proportional workload is small. This implies that NYHA classification is not a sole representation of a patient's functional capacity, but might also incude other aspects of the patient's overall health status., Competing Interests: LH, IIT and GAS are employed by Cardiology Centers of the Netherlands. HMdR declares to be a Health Holland committee member, a supervisory member of Stichting LEEFH and vice-president of the Young Academy. YA declares to be in the steering committee of two clinical trials (iCorMiCa and ILIAS-ANOCA). FG, KRS, NCOM and NAS declare to have no conflict of interest., (Copyright: © 2022 The Author(s). Published by IMR Press.)

Aims: The HFA-PEFF score was developed to optimize diagnosis and to aid in early recognition of heart failure (HF) with preserved ejection fraction (HFpEF) in patients who present with HF-like symptoms. Recognizing early-HFpEF phenogroups is essential to better understand progression towards overt HFpEF and pave the way for early intervention and treatment. Whether the HFA-PEFF domain scores can identify 'early-HFpEF' phenogroups remains unknown. The aims of this pilot study are to (i) identify distinct phenogroups by cluster analysis of HFA-PEFF domain scores in subjects that present with HF-like symptoms and (ii) study whether these phenogroups may be associated with distinct blood proteome profiles., Methods and Results: Subjects referred to the Cardiology Centers of the Netherlands, location Utrecht, with non-acute possibly cardiac-related symptoms (such as dyspnoea or fatigue) were prospectively enrolled in the HELPFul cohort (N = 507) and were included in the current analysis. Inclusion criteria for this study were (i) age ≥ 45 years and (ii) a left ventricular ejection fraction (LVEF) ≥ 50%, in the absence of a history of HF, coronary artery disease, congenital heart disease, or any previous cardiac interventions. Multinominal-based clustering with latent class model using the HFA-PEFF domain scores (functional, structural, and biomarker scores) as input was used to detect distinct phenotypic clusters. For each bootstrapping run, the 92 Olink proteins were analysed for their association with the identified phenogroups. Four distinct phenogroups were identified in the current analysis (validated by bootstrapping 1000×): (i) no left ventricular diastolic dysfunction (no LVDD, N = 102); (ii) LVDD with functional left ventricular (LV) abnormalities (N = 204); (iii) LVDD with functional and structural LV abnormalities (N = 204); and (iv) LVDD with functional and structural LV abnormalities and elevated BNP (N = 107). The HFA-PEFF total score risk categories significantly differed between the phenogroups (P < 0.001), with an increase of the HFA-PEFF score from Phenogroup 1 to 4 (low/intermediate/high HFA-PEFF risk score: Phenogroup 1: 88%/12%/0%; Phenogroup 2: 9%/91%/0%; Phenogroup 3: 0%/92%/8%; Phenogroup 4: 5%/83%/12%). Thirty-two out of the 92 Olink protein biomarkers significantly differed among the phenogroups. The top eight biomarkers-N-terminal prohormone brain natriuretic peptide, growth differentiation factor-15, matrix metalloproteinase-2, osteoprotegerin, tissue inhibitor of metalloproteinase-4, chitinase-3-like protein 1, insulin-like growth factor-binding protein 2, and insulin-like growth factor-binding protein 7-are mainly involved in inflammation and extracellular matrix remodelling, which are currently proposed key processes in HFpEF pathophysiology., Conclusions: This study identified distinct phenogroups by using the HFA-PEFF domain scores in ambulant subjects referred for HF-like symptoms. The newly identified phenogroups accompanied by their circulating biomarkers profile might aid in a better understanding of the pathophysiological processes involved during the early stages of the HFpEF syndrome., (© 2022 The Authors. ESC Heart Failure published by John Wiley & Sons Ltd on behalf of European Society of Cardiology.)

At least one-half of the growing heart failure population consists of heart failure with preserved ejection fraction (HFpEF). The limited therapeutic options, the complexity of the syndrome, and many related comorbidities emphasize the need for adequate experimental animal models to study the etiology of HFpEF, as well as its comorbidities and pathophysiological changes. The strengths and weaknesses of available animal models have been reviewed extensively with the general consensus that a "1-size-fits-all" model does not exist, because no uniform HFpEF patient exists. In fact, HFpEF patients have been categorized into HFpEF phenogroups based on comorbidities and symptoms. In this review, we therefore study which animal model is best suited to study the different phenogroups-to improve model selection and refinement of animal research. Based on the published data, we extrapolated human HFpEF phenogroups into 3 animal phenogroups (containing small and large animals) based on reports and definitions of the authors: animal models with high (cardiac) age (phenogroup aging); animal models focusing on hypertension and kidney dysfunction (phenogroup hypertension/kidney failure); and models with hypertension, obesity, and type 2 diabetes mellitus (phenogroup cardiometabolic syndrome). We subsequently evaluated characteristics of HFpEF, such as left ventricular diastolic dysfunction parameters, systemic inflammation, cardiac fibrosis, and sex-specificity in the different models. Finally, we scored these parameters concluded how to best apply these models. Based on our findings, we propose an easy-to-use classification for future animal research based on clinical phenogroups of interest., Competing Interests: This work was supported by Netherlands Cardiovascular Research Initiative, with the support of the Dutch Heart Foundation, the Netherlands (CVON2018-30 PREDICT2 to Drs van Ham and van Veen, Senior Clinical Scientist grant 2020T058 to Dr Handoko; EARLY-HFpEF Young Talent Grant 2015-10 to Dr Kessler, RECONNEXT 2020B008 and IMPRESS 2020B004 to Drs Kessler, Handoko, den Ruijter, and de Jager). The authors have reported that they have no relationships relevant to the contents of this paper to disclose., (© 2022 The Authors.)

Background: Fundamental aspects of human identity may play a role in the presentation of stroke symptoms and, consequently, stroke recognition. Strokes must be recognized and treated expeditiously, as delays result in poorer outcomes. It is known that sex plays a role in the presentation of symptoms, such that non-traditional symptoms are more commonly observed among women. However, factors such as geographical location and race/ethnicity, and the interactions between these various factors, need to be considered. This will provide an intersectional approach., Methods: A systematic review and meta-analysis of the literature was conducted to investigate differences in the presentation of stroke symptoms between sexes. Using PubMed and Embase, a search involving the components sex, symptoms and stroke was completed and yielded 26 full-text manuscripts., Results: Our findings indicate that there is substantial overlap in stroke symptom presentation in men and women. Nonetheless, some differences in the clinical manifestations of stroke were observed. In addition, it was discovered that only three studies were conducted outside of North America and Europe. Furthermore, only two studies reported symptoms based on both sex and racial/ethnic group., Conclusion: These findings indicate a research gap and call for increased research in order to uncover the possible interactions between sex and race/ethnicity in an intersectional approach. Resultantly, stroke recognition could be improved and greater equity in healthcare can be achieved., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Hosman, Engels, den Ruijter and Exalto.)

Objectives: Women have a worse prognosis after coronary artery bypass grafting (CABG) surgery compared to men. We sought to quantify to what extent this difference in post-CABG survival could be attributed to sex itself, or whether this was mediated by difference between men and women at the time of intervention. Additionally, we explored to what extent these effects were homogenous across patient subgroups., Methods: Time to all-cause mortality was available for 102,263 CABG patients, including 20,988 (21%) women, sourced through an individual participant data meta-analysis of five cohort studies. Difference between men and women in survival duration was assessed using Kaplan-Meier estimates, and Cox's proportional hazards model., Results: During a median follow-up of 5 years, 13,598 (13%) patients died, with women more likely to die than men: female HR 1.20 (95%CI 1.16; 1.25). We found that differences in patient characteristics at the time of CABG procedure mediated this sex effect, and accounting for these resulted in a neutral female HR 0.98 (95%CI 0.94; 1.02). Next we performed a priori defined subgroup analyses of the five most prominent mediators: age, creatinine, peripheral vascular disease, type 2 diabetes, and heart failure. We found that women without peripheral vascular disease (PVD) or women aged 70+, survived longer than men (interaction p -values 0.04 and 6 × 10 -5 , respectively), with an effect reversal in younger women., Conclusion: Sex differences in post-CABG survival were readily explained by difference in patient characteristics and comorbidities. Pre-planned analyses revealed patient subgroups (aged 70+, or without PVD) of women that survived longer than men, and a subgroup of younger women with comparatively poorer survival., Competing Interests: MH has received honoraria from Idorsia not related to this study. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Schmidt, Haitjema, Sartipy, Holzmann, Malenka, Ross, van Gilst, Rouleau, Meeder, Baker, Shiomi, Kimura, Tran, Smith, Reid, Asselbergs and den Ruijter.)

Objectives: Uncertainty about the benefit of (high-intensity) statins for women remains due to under-representation of women in primary prevention trials and scarcity of sex-stratified data. This study evaluates the sex-specific relation between statin treatment and survival and the additional benefit of high-intensity statins., Methods: Electronic health record data from 47 801 patients (17 008 statin users and 30 793 non-users) without prior cardiovascular disease were extracted from thirteen Dutch outpatient cardiology clinics. Patients prescribed statins at baseline were propensity-score matched to those eligible for statin therapy (low-density lipoprotein >2.5 mmol/L) without a statin prescription. Statins were divided into low-intensity and high-intensity according to Dutch guidelines. Mortality data were obtained via linkage to the national mortality registry. Cox regression was used to evaluate the relationship between statin prescription and intensity and all-cause and cardiovascular mortality., Results: Propensity score matching created a cohort of 8631 statin users and 8631 non-users. 35% of women and 28% of men received a low-intensity statin. The beneficial effect of statins on both all-cause and cardiovascular mortality was stronger in women (HR 0.66, 95% CI 0.58 to 0.74 and HR 0.55, 95% CI 0.39 to 0.71, respectively) than in men (HR 0.89, 95% CI 0.81 to 0.95 and HR 0.93, 95% CI 0.77 to 1.08, respectively). High-intensity statins conferred modest protection against all-cause mortality (HR 0.94, 95% CI 0.88 to 1.00) and cardiovascular mortality (HR 0.86, 95% CI 0.74 to 0.98) in both sexes., Conclusions: The protective effect of primary prevention statins was stronger in women than men for both all-cause and cardiovascular mortality. High-intensity statins conferred a modest additional benefit in both sexes. Statins seem to be effective regardless of treatment intensity, especially in women., Competing Interests: Competing interests: LH, IIT and GAS are employed by Cardiology Centers of the Netherlands. All other authors declare no competing interests., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Atherosclerosis is a complex disease characterized by the formation of arterial plaques with a broad diversity of morphological phenotypic presentations. Researchers often apply one description of the vulnerable plaque as a gold standard in preclinical and clinical research that could be applied as a surrogate measure of a successful therapeutic intervention, despite the variability in lesion characteristics that may underly a thrombotic occlusion. The complex mechanistic interplay underlying progression of atherosclerotic disease is a consequence of the broad range of determinants such as sex, risk factors, hemodynamics, medications, and the genetic landscape. Currently, we are facing an overwhelming amount of data based on genetic, transcriptomic, proteomic, and metabolomic studies that all point to heterogeneous molecular profiles of atherosclerotic lesions that lead to a myocardial infarction or stroke. The observed molecular diversity implies that one unifying model cannot fully recapitulate the natural history of atherosclerosis. Despite emerging data obtained from -omics studies, a description of a natural history of atherosclerotic disease in which cell-specific expression of proteins or genes are included is still lacking. This also applies to the insights provided by genome-wide association studies. This review will critically discuss the dogma that the progression of atherosclerotic disease can be captured in one unifying natural history model of atherosclerosis.

Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Aims: Incorporation of sex in study design can lead to discoveries in medical research. Deep neural networks (DNNs) accurately predict sex based on the electrocardiogram (ECG) and we hypothesized that misclassification of sex is an important predictor for mortality. Therefore, we first developed and validated a DNN that classified sex based on the ECG and investigated the outcome. Second, we studied ECG drivers of DNN-classified sex and mortality., Methods and Results: A DNN was trained to classify sex based on 131 673 normal ECGs. The algorithm was validated on internal (68 500 ECGs) and external data sets (3303 and 4457 ECGs). The survival of sex (mis)classified groups was investigated using time-to-event analysis and sex-stratified mediation analysis of ECG features. The DNN successfully distinguished female from male ECGs {internal validation: area under the curve (AUC) 0.96 [95% confidence interval (CI): 0.96, 0.97]; external validations: AUC 0.89 (95% CI: 0.88, 0.90), 0.94 (95% CI: 0.93, 0.94)}. Sex-misclassified individuals (11%) had a 1.4 times higher mortality risk compared with correctly classified peers. The ventricular rate was the strongest mediating ECG variable (41%, 95% CI: 31%, 56%) in males, while the maximum amplitude of the ST segment was strongest in females (18%, 95% CI: 11%, 39%). Short QRS duration was associated with higher mortality risk., Conclusion: Deep neural networks accurately classify sex based on ECGs. While the proportion of ECG-based sex misclassifications is low, it is an interesting biomarker. Investigation of the causal pathway between misclassification and mortality uncovered new ECG features that might be associated with mortality. Increased emphasis on sex as a biological variable in artificial intelligence is warranted., (© The Author(s) 2022. Published by Oxford University Press on behalf of European Society of Cardiology.)

Aims: This study aimed to assess the sex-specific distribution of heart failure (HF) with preserved, mid-range, and reduced ejection fraction across three health care settings., Methods and Results: In this descriptive observational study, we retrieved the distribution of HF types [with reduced ejection fraction (HFrEF), mid-range ejection fraction (HFmrEF), and preserved ejection fraction (HFpEF)] for men and women between 65 and 79 years of age in three health care settings from a single country: (i) patients with screening-detected HF in the high-risk community (i.e. those with shortness of breath, frailty, diabetes mellitus, and chronic obstructive pulmonary disease) from four screening studies, (ii) patients with confirmed HF from primary care derived from a single observational study, and (iii) patients with confirmed HF from outpatient cardiology clinics participating in a registry. Among 1407 patients from the high-risk community, 288 had screen-detected HF (15% HFrEF, 12% HFmrEF, 74% HFpEF), and 51% of the screen-detected HF patients were women. In both women (82%) and men (65%), HFpEF was the most prevalent HF type. In the routine general practice population (30 practices, 70 000 individuals), among the 160 confirmed HF cases, 35% had HFrEF, 23% HFmrEF, and 43% HFpEF, and in total, 43% were women. In women, HFpEF was the most prevalent HF type (52%), while in men, this was HFrEF (41%). In outpatient cardiology clinics (n = 34), of the 4742 HF patients (66% HFrEF, 15% HFmrEF, 20% HFpEF), 36% were women. In both women (56%) and men (71%), HFrEF was the most prevalent HF type., Conclusions: Both HF types and sex distribution vary considerably in HF patients of 65-79 years of age among health care settings. From the high-risk community through to general practice to the cardiology outpatient setting, there is a shift in HF type from HFpEF to HFrEF and a decrease in the proportion of HF patients that are women., (© 2021 The Authors. ESC Heart Failure published by John Wiley & Sons Ltd on behalf of European Society of Cardiology.)

Background: Knowledge about adverse drug reactions (ADRs) in the population is limited because of underreporting, which hampers surveillance and assessment of drug safety. Therefore, gathering accurate information that can be retrieved from clinical notes about the incidence of ADRs is of great relevance. However, manual labeling of these notes is time-consuming, and automatization can improve the use of free-text clinical notes for the identification of ADRs. Furthermore, tools for language processing in languages other than English are not widely available., Objective: The aim of this study is to design and evaluate a method for automatic extraction of medication and Adverse Drug Reaction Identification in Clinical Notes (ADRIN)., Methods: Dutch free-text clinical notes (N=277,398) and medication registrations (N=499,435) from the Cardiology Centers of the Netherlands database were used. All clinical notes were used to develop word embedding models. Vector representations of word embedding models and string matching with a medical dictionary (Medical Dictionary for Regulatory Activities [MedDRA]) were used for identification of ADRs and medication in a test set of clinical notes that were manually labeled. Several settings, including search area and punctuation, could be adjusted in the prototype to evaluate the optimal version of the prototype., Results: The ADRIN method was evaluated using a test set of 988 clinical notes written on the stop date of a drug. Multiple versions of the prototype were evaluated for a variety of tasks. Binary classification of ADR presence achieved the highest accuracy of 0.84. Reduced search area and inclusion of punctuation improved performance, whereas incorporation of the MedDRA did not improve the performance of the pipeline., Conclusions: The ADRIN method and prototype are effective in recognizing ADRs in Dutch clinical notes from cardiac diagnostic screening centers. Surprisingly, incorporation of the MedDRA did not result in improved identification on top of word embedding models. The implementation of the ADRIN tool may help increase the identification of ADRs, resulting in better care and saving substantial health care costs., (©Klaske R Siegersma, Maxime Evers, Sophie H Bots, Floor Groepenhoff, Yolande Appelman, Leonard Hofstra, Igor I Tulevski, G Aernout Somsen, Hester M den Ruijter, Marco Spruit, N Charlotte Onland-Moret. Originally published in JMIR Medical Informatics (https://medinform.jmir.org), 25.01.2022.)