Apolipoprotein E-ε2 and Resistance to Atherosclerosis in Midlife: The PESA Observational Study.

BACKGROUND: APOE is a known genetic contributor to cardiovascular disease, but the differential role APOE alleles play in subclinical atherosclerosis remains unclear. METHODS: The PESA (Progression of Early Subclinical Atherosclerosis) is an observational cohort study that recruited 4184 middle-aged asymptomatic individuals to be screened for cardiovascular risk and multiterritorial subclinical atherosclerosis. Participants were APOE-genotyped, and omics data were additionally evaluated. RESULTS: In the PESA study, the frequencies for APOE -ε2, -ε3, and -ε4 alleles were 0.060, 0.844, and 0.096, respectively. This study included a subcohort of 3887 participants (45.8±4.3 years of age; 62% males). As expected, APOE-ε4 carriers were at the highest risk for cardiovascular disease and had significantly greater odds of having subclinical atherosclerosis compared with ε3/ε3 carriers, which was mainly explained by their higher levels of low-density lipoprotein (LDL)-cholesterol. In turn, APOE-ε2 carriers were at the lowest risk for cardiovascular disease and had significantly lower odds of having subclinical atherosclerosis in several vascular territories (carotids: 0.62 [95% CI, 0.47-0.81]; P=0.00043; femorals: 0.60 [0.47-0.78]; P=9.96×10-5; coronaries: 0.53 [0.39-0.74]; P=0.00013; and increased PESA score: 0.58 [0.48-0.71]; P=3.16×10-8). This APOE-ε2 atheroprotective effect was mostly independent of the associated lower LDL-cholesterol levels and other cardiovascular risk factors. The protection conferred by the ε2 allele was greater with age (50-54 years: 0.49 [95% CI, 0.32-0.73]; P=0.00045), and normal (<150 mg/dL) levels of triglycerides (0.54 [0.44-0.66]; P=4.70×10-9 versus 0.90 [0.57-1.43]; P=0.67 if ≥150 mg/dL). Omics analysis revealed an enrichment of several canonical pathways associated with anti-inflammatory mechanisms together with the modulation of erythrocyte homeostasis, coagulation, and complement activation in ε2 carriers that might play a relevant role in the ε2's atheroprotective effect. CONCLUSIONS: This work sheds light on the role of APOE in cardiovascular disease development with important therapeutic and prevention implications on cardiovascular health, especially in early midlife. REGISTRATION: URL: https://www.clinicaltrials.gov: NCT01410318.

Capillary pruning couples tissue perfusion and oxygenation with cardiomyocyte maturation in the postnatal mouse heart.

Introduction: Removal of poorly perfused capillaries by pruning contributes to remodeling the microvasculature to optimize oxygen and nutrient delivery. Blood flow drives this process by promoting the intravascular migration of endothelial cells in developing networks, such as in the yolk sac, zebrafish brain or postnatal mouse retina. Methods: In this study, we have implemented innovative tools to recognize capillary pruning in the complex 3D coronary microvasculature of the postnatal mouse heart. We have also experimentally tested the impact of decreasing pruning on the structure and function of this network by altering blood flow with two different vasodilators: losartan and prazosin. Results: Although both drugs reduced capillary pruning, a combination of experiments based on ex vivo imaging, proteomics, electron microscopy and in vivo functional approaches showed that losartan treatment resulted in an inefficient coronary network, reduced myocardial oxygenation and metabolic changes that delayed the arrest of cardiomyocyte proliferation, in contrast to the effects of prazosin, probably due to its concomitant promotion of capillary expansion. Discussion: Our work demonstrates that capillary pruning contributes to proper maturation and function of the heart and that manipulation of blood flow may be a novel strategy to refine the microvasculature and improve tissue perfusion after damage.

Cardiac Insulin Resistance in Subjects With Metabolic Syndrome Traits and Early Subclinical Atherosclerosis.

OBJECTIVE: Experimental evidence suggests that metabolic syndrome (MetS) is associated with changes in cardiac metabolism. Whether this association occurs in humans is unknown. RESEARCH DESIGN AND METHODS: 821 asymptomatic individuals from the Progression of Early Subclinical Atherosclerosis (PESA) study (50.6 [46.9-53.6] years, 83.7% male) underwent two whole-body 18F-fluorodeoxyglucose positron emission tomography-magnetic resonance (18F-FDG PET-MR) 4.8 ± 0.6 years apart. Presence of myocardial 18F-FDG uptake was evaluated qualitatively and quantitatively. No myocardial uptake was grade 0, while positive uptake was classified in grades 1-3 according to target-to-background ratio tertiles. RESULTS: One hundred fifty-six participants (19.0%) showed no myocardial 18F-FDG uptake, and this was significantly associated with higher prevalence of MetS (29.0% vs. 13.9%, P < 0.001), hypertension (29.0% vs. 18.0%, P = 0.002), and diabetes (11.0% vs. 3.2%, P < 0.001), and with higher insulin resistance index (HOMA-IR, 1.64% vs. 1.23%, P < 0.001). Absence of myocardial uptake was associated with higher prevalence of early atherosclerosis (i.e., arterial 18F-FDG uptake, P = 0.004). On follow-up, the associations between myocardial 18F-FDG uptake and risk factors were replicated, and MetS was more frequent in the group without myocardial uptake. The increase in HOMA-IR was associated with a progressive decrease in myocardial uptake (P < 0.001). In 82% of subjects, the categorization according to presence/absence of myocardial 18F-FDG uptake did not change between baseline and follow-up. MetS regression on follow-up was associated with a significant (P < 0.001) increase in myocardial uptake. CONCLUSIONS: Apparently healthy individuals without cardiac 18F-FDG uptake have higher HOMA-IR and higher prevalence of MetS traits, cardiovascular risk factors, and early atherosclerosis. An improvement in cardiometabolic profile is associated with the recovery of myocardial 18F-FDG uptake at follow-up.

Subclinical atherosclerosis and accelerated epigenetic age mediated by inflammation: a multi-omics study.

AIMS: Epigenetic age is emerging as a personalized and accurate predictor of biological age. The aim of this article is to assess the association of subclinical atherosclerosis with accelerated epigenetic age and to investigate the underlying mechanisms mediating this association. METHODS AND RESULTS: Whole blood methylomics, transcriptomics, and plasma proteomics were obtained for 391 participants of the Progression of Early Subclinical Atherosclerosis study. Epigenetic age was calculated from methylomics data for each participant. Its divergence from chronological age is termed epigenetic age acceleration. Subclinical atherosclerosis burden was estimated by multi-territory 2D/3D vascular ultrasound and by coronary artery calcification. In healthy individuals, the presence, extension, and progression of subclinical atherosclerosis were associated with a significant acceleration of the Grim epigenetic age, a predictor of health and lifespan, regardless of traditional cardiovascular risk factors. Individuals with an accelerated Grim epigenetic age were characterized by an increased systemic inflammation and associated with a score of low-grade, chronic inflammation. Mediation analysis using transcriptomics and proteomics data revealed key pro-inflammatory pathways (IL6, Inflammasome, and IL10) and genes (IL1B, OSM, TLR5, and CD14) mediating the association between subclinical atherosclerosis and epigenetic age acceleration. CONCLUSION: The presence, extension, and progression of subclinical atherosclerosis in middle-aged asymptomatic individuals are associated with an acceleration in the Grim epigenetic age. Mediation analysis using transcriptomics and proteomics data suggests a key role of systemic inflammation in this association, reinforcing the relevance of interventions on inflammation to prevent cardiovascular disease.

Genome Editing and Inherited Cardiac Arrhythmias.

Inherited arrhythmic disorders are a group of heterogeneous diseases predisposing to life-threatening arrhythmias and sudden cardiac death. Their diagnosis is not always simple due to incomplete penetrance and genetic heterogeneity. Furthermore, the available treatments are usually invasive and merely preventive. Genome editing and especially CRISPR/Cas9 technologies have the potential to correct the genetic arrhythmogenic substrate, thereby offering a cure for these fatal diseases. To date, genome editing has allowed reproducing cardiac arrhythmias in vitro, providing a robust platform for variant pathogenicity, mechanistic, and drug-testing studies. However, in vivo approaches still need profound research regarding safety, specificity, and efficiency of the methods.

A novel data-driven method for the analysis and reconstruction of cardiac cine MRI.

Cardiac cine magnetic resonance imaging (MRI) can be considered the optimal criterion for measuring cardiac function. This imaging technique can provide us with detailed information about cardiac structure, tissue composition and even blood flow, which makes it highly used in medical science. But due to the image time acquisition and several other factors the MRI sequences can easily get corrupted, causing radiologists to misdiagnose 40 million people worldwide each and every single year. Hence, the urge to decrease these numbers, researchers from different fields have been introducing novel tools and methods in the medical field. Aiming to the same target, we consider in this work the application of the higher order dynamic mode decomposition (HODMD) technique. The HODMD algorithm is a linear method, which was originally introduced in the fluid dynamics domain, for the analysis of complex systems. Nevertheless, the proposed method has extended its applicability to numerous domains, including medicine. In this work, HODMD in used to analyze sets of MR images of a heart, with the ultimate goal of identifying the main patterns and frequencies driving the heart dynamics. Furthermore, a novel interpolation algorithm based on singular value decomposition combined with HODMD is introduced, providing a three-dimensional reconstruction of the heart. This algorithm is applied (i) to reconstruct corrupted or missing images, and (ii) to build a reduced order model of the heart dynamics.

Non-invasive assessment of HFpEF in mouse models: current gaps and future directions.

BACKGROUND: Heart failure (HF) with preserved ejection fraction (HFpEF) prevalence is increasing, and large clinical trials have failed to reduce mortality. A major reason for this outcome is the failure to translate results from basic research to the clinics. Evaluation of HFpEF in mouse models requires assessing three major key features defining this complex syndrome: the presence of a preserved left ventricular ejection fraction (LVEF), diastolic dysfunction, and the development of HF. In addition, HFpEF is associated with multiple comorbidities such as systemic arterial hypertension, chronic obstructive pulmonary disease, sleep apnea, diabetes, and obesity; thus, non-cardiac disorders assessment is crucial for a complete phenotype characterization. Non-invasive procedures present unquestionable advantages to maintain animal welfare and enable longitudinal analyses. However, unequivocally determining the presence of HFpEF using these methods remains challenging. MAIN TEXT: Transthoracic echocardiography (TTE) represents an invaluable tool in HFpEF diagnosis, allowing evaluation of LVEF, diastolic dysfunction, and lung congestion in mice. Since conventional parameters used to evaluate an abnormal diastole like E/A ratio, isovolumic relaxation time, and E/e' may pose limitations in mice, including advanced TTE techniques to characterize cardiac motion, including an assessment under stress, will improve diagnosis. Patients with HFpEF also show electrical cardiac remodelling and therefore electrocardiography may add valuable information in mouse models to assess chronotropic incompetence and sinoatrial node dysfunction, which are major contributors to exercise intolerance. To complete the non-invasive diagnosis of HF, low aerobic exercise capacity and fatigue using exercise tests, impaired oxygen exchange using metabolic cages, and determination of blood biomarkers can be determined. Finally, since HFpEF patients commonly present non-cardiac pathological conditions, acquisition of systemic and pulmonary arterial pressures, blood glucose levels, and performing glucose tolerance and insulin resistance tests are required for a complete phenotyping. CONCLUSION: Identification of reliable models of HFpEF in mice by using proper diagnosis tools is necessary to translate basic research results to the clinics. Determining the presence of several HFpEF indicators and a higher number of abnormal parameters will lead to more reliable evidence of HFpEF.

Frequency of hereditary transthyretin amyloidosis among elderly patients with transthyretin cardiomyopathy.

AIMS: Transthyretin amyloid cardiomyopathy (ATTR-CM) is increasingly recognized as a cause of heart failure in the elderly. Although wild-type transthyretin amyloidosis is the most frequent form of ATTR-CM found in the elderly, hereditary transthyretin amyloidosis (ATTRv) can also occur. We sought to determine the prevalence of ATTRv among elderly ATTR-CM patients, identify predictors of ATTRv and evaluate the clinical consequences of positive genetic testing in this population. METHODS AND RESULTS: Prevalence of ATTRv in elderly ATTR-CM patients (≥70 years) was assessed in a cohort of 300 consecutive ATTR-CM patients (median age 78 years at diagnosis, 82% ≥70 years, 16% female, 99% Caucasian). ATTRv was diagnosed in 35 (12%; 95% confidence interval [CI] 3.1-8.8) and 13 (5.3%; 95% CI 5.6-26.7) patients in the overall cohort and in those ≥70 years, respectively. Prevalence of ATTRv among elderly female patients with ATTR-CM was 13% (95% CI 2.1-23.5). Univariate analysis identified female sex (odds ratio [OR] 3.66; 95% CI 1.13-11.85; p = 0.03), black ancestry (OR 46.31; 95% CI 3.52-Inf; p = 0.005), eye symptoms (OR 6.64; 95% CI 1.20-36.73; p = 0.03) and polyneuropathy (OR 10.05; 95% CI 3.09-32.64; p < 0.001) as the only factors associated with ATTRv in this population. Diagnosis of ATTRv in elderly ATTR-CM patients allowed initiation of transthyretin-specific drug treatment in 5 individuals, genetic screening in 33 relatives from 13 families, and identification of 9 ATTRv asymptomatic carriers. CONCLUSIONS: Hereditary transthyretin amyloidosis is present in a substantial number of ATTR-CM patients aged ≥70 years. Identification of ATTRv in elderly patients with ATTR-CM has clinical meaningful therapeutic and diagnostic implications. These results support routine genetic testing in patients with ATTR-CM regardless of age.

Higher order dynamic mode decomposition: From fluid dynamics to heart disease analysis.

In this work, we study in detail the performance of Higher Order Dynamic Mode Decomposition (HODMD) technique when applied to echocardiography images. HODMD is a data-driven method generally used in fluid dynamics and in the analysis of complex non-linear dynamical systems modeling several complex industrial applications. In this paper we apply HODMD, for the first time to the authors knowledge, for patterns recognition in echocardiography, specifically, echocardiography data taken from several mice, either in healthy conditions or afflicted by different cardiac diseases. We exploit the HODMD advantageous properties in dynamics identification and noise cleaning to identify the relevant frequencies and coherent patterns for each one of the diseases. The echocardiography datasets consist of video loops taken with respect to a long axis view (LAX) and a short axis view (SAX), where each video loop covers at least three cardiac cycles, formed by (at most) 300 frames each (called snapshots). The proposed algorithm, using only a maximum quantity of 200 snapshots, was able to capture two branches of frequencies, representing the heart rate and respiratory rate. Additionally, the algorithm provided a number of modes, which represent the dominant features and patterns in the different echocardiography images, also related to the heart and the lung. Six datasets were analyzed: one echocardiography taken from a healthy subject and five different sets of echocardiography taken from subjects with either Diabetic Cardiomyopathy, Obesity, SFSR4 Hypertrophy, TAC Hypertrophy or Myocardial Infarction. The results show that HODMD is robust and a suitable tool to identify characteristic patterns able to classify the different pathologies studied.

Functional Impact and Regulation of Alternative Splicing in Mouse Heart Development and Disease.

Alternative splicing (AS) plays a major role in the generation of transcript diversity. In the heart, roles have been described for some AS variants, but the global impact and regulation of AS patterns are poorly understood. Here, we studied the AS profiles in heart disease, their relationship with heart development, and the regulatory mechanisms controlling AS dynamics in the mouse heart. We found that AS profiles characterized the different groups and that AS and gene expression changes affected independent genes and biological functions. Moreover, AS changes, specifically in heart disease, were associated with potential protein-protein interaction changes. While developmental transitions were mainly driven by the upregulation of MBNL1, AS changes in disease were driven by a complex regulatory network, where PTBP1 played a central role. Indeed, PTBP1 over-expression was sufficient to induce cardiac hypertrophy and diastolic dysfunction, potentially by perturbing AS patterns.

Unbiased plasma proteomics discovery of biomarkers for improved detection of subclinical atherosclerosis.

BACKGROUND: Imaging of subclinical atherosclerosis improves cardiovascular risk prediction on top of traditional risk factors. However, cardiovascular imaging is not universally available. This work aims to identify circulating proteins that could predict subclinical atherosclerosis. METHODS: Hypothesis-free proteomics was used to analyze plasma from 444 subjects from PESA cohort study (222 with extensive atherosclerosis on imaging, and 222 matched controls) at two timepoints (three years apart) for discovery, and from 350 subjects from AWHS cohort study (175 subjects with extensive atherosclerosis on imaging and 175 matched controls) for external validation. A selected three-protein panel was further validated by immunoturbidimetry in the AWHS population and in 2999 subjects from ILERVAS cohort study. FINDINGS: PIGR, IGHA2, APOA, HPT and HEP2 were associated with subclinical atherosclerosis independently from traditional risk factors at both timepoints in the discovery and validation cohorts. Multivariate analysis rendered a potential three-protein biomarker panel, including IGHA2, APOA and HPT. Immunoturbidimetry confirmed the independent associations of these three proteins with subclinical atherosclerosis in AWHS and ILERVAS. A machine-learning model with these three proteins was able to predict subclinical atherosclerosis in ILERVAS (AUC [95%CI]:0.73 [0.70-0.74], p < 1 × 10-99), and also in the subpopulation of individuals with low cardiovascular risk according to FHS 10-year score (0.71 [0.69-0.73], p < 1 × 10-69). INTERPRETATION: Plasma levels of IGHA2, APOA and HPT are associated with subclinical atherosclerosis independently of traditional risk factors and offers potential to predict this disease. The panel could improve primary prevention strategies in areas where imaging is not available. FUNDING: This study was supported by competitive grants from the Spanish Ministry of Science, Innovation and Universities (BIO2015-67580-P, PGC2018-097019-B-I00, PID2019-106814RB-I00 and SAF2016-80843-R), through the Carlos III Institute of Health-Fondo de Investigacion Sanitaria grant PRB3 (IPT17/0019 - ISCIII-SGEFI / ERDF, ProteoRed), CIBERCV and CIBERDEM, the Fundacio MaratoTV3 (grant 122/C/2015) and "la Caixa" Banking Foundation (project HR17-00247). The PESA study is co-funded equally by the Centro Nacional de Investigaciones Cardiovasculares (CNIC), Madrid, Spain, and Banco Santander, Madrid, Spain. The ILERVAS study was funded by the Diputacio de Lleida. The study also receives funding from the Instituto de Salud Carlos III (PI15/02019; PI18/00610; RD16/0009) and the FEDER funds. The CNIC is supported by the Instituto de Salud Carlos III (ISCIII), the Ministerio de Ciencia, Innovacion y Universidades (MCNU) and the Pro CNIC Foundation.

Assessment of myocardial viscoelasticity with Brillouin spectroscopy in myocardial infarction and aortic stenosis models.

Heart diseases are associated with changes in the biomechanical properties of the myocardial wall. However, there is no modality available to assess myocardial stiffness directly. Brillouin microspectroscopy (mBS) is a consolidated mechanical characterization technique, applied to the study of the viscoelastic and elastic behavior of biological samples and may be a valuable tool for assessing the viscoelastic properties of the cardiac tissue. In this work, viscosity and elasticity were assessed using mBS in heart samples obtained from healthy and unhealthy mice (n = 6 per group). Speckle-tracking echocardiography (STE) was performed to evaluate heart deformation. We found that mBS was able to detect changes in stiffness in the ventricles in healthy myocardium. The right ventricle showed reduced stiffness, in agreement with its increased compliance. mBS measurements correlated strongly with STE data, highlighting the association between displacement and stiffness in myocardial regions. This correlation was lost in pathological conditions studied. The scar region in the infarcted heart presented changes in stiffness when compared to the rest of the heart, and the hypertrophied left ventricle showed increased stiffness following aortic stenosis, compared to the right ventricle. We demonstrate that mBS can be applied to determine myocardial stiffness, that measurements correlate with functional parameters and that they change with disease.