Diffusion imaging genomics provides novel insight into early mechanisms of cerebral small vessel disease.

Cerebral small vessel disease (cSVD) is a leading cause of stroke and dementia. Genetic risk loci for white matter hyperintensities (WMH), the most common MRI-marker of cSVD in older age, were recently shown to be significantly associated with white matter (WM) microstructure on diffusion tensor imaging (signal-based) in young adults. To provide new insights into these early changes in WM microstructure and their relation with cSVD, we sought to explore the genetic underpinnings of cutting-edge tissue-based diffusion imaging markers across the adult lifespan. We conducted a genome-wide association study of neurite orientation dispersion and density imaging (NODDI) markers in young adults (i-Share study: N = 1 758, (mean[range]) 22.1[18-35] years), with follow-up in young middle-aged (Rhineland Study: N = 714, 35.2[30-40] years) and late middle-aged to older individuals (UK Biobank: N = 33 224, 64.3[45-82] years). We identified 21 loci associated with NODDI markers across brain regions in young adults. The most robust association, replicated in both follow-up cohorts, was with Neurite Density Index (NDI) at chr5q14.3, a known WMH locus in VCAN. Two additional loci were replicated in UK Biobank, at chr17q21.2 with NDI, and chr19q13.12 with Orientation Dispersion Index (ODI). Transcriptome-wide association studies showed associations of STAT3 expression in arterial and adipose tissue (chr17q21.2) with NDI, and of several genes at chr19q13.12 with ODI. Genetic susceptibility to larger WMH volume, but not to vascular risk factors, was significantly associated with decreased NDI in young adults, especially in regions known to harbor WMH in older age. Individually, seven of 25 known WMH risk loci were associated with NDI in young adults. In conclusion, we identified multiple novel genetic risk loci associated with NODDI markers, particularly NDI, in early adulthood. These point to possible early-life mechanisms underlying cSVD and to processes involving remyelination, neurodevelopment and neurodegeneration, with a potential for novel approaches to prevention.

Skin autofluorescence of advanced glycation end-products relates to new cardiovascular events in type 2 diabetes: A longitudinal observational study.

BACKGROUND: Cardiovascular disease is frequent in type 2 diabetes mellitus (T2DM). We investigated the relationship between skin autofluorescence (SAF) of advanced glycation end-products and later cardiovascular events (CVEs) in patients with T2DM. RESEARCH DESIGN AND METHODS: We conducted a retrospective analysis of 504 patients hospitalized for uncontrolled and/or complicated T2DM between 2009 and 2017. SAF was measured using an AGE-Reader. Participants were followed up from admission to December 2020, for the onset of a CVE (myocardial infarction, stroke, revascularization procedures or cardiovascular death). The relationship between SAF and CVE was analyzed by multivariable Cox regression. Log-rank curves were used to compare CVE-free survival in patients whose SAF at admission was above versus below the whole-population median. The analysis was repeated in subjects without/with macroangiopathy (defined as myocardial infarction, stroke, peripheral revascularization) at baseline. FINDINGS: During 54 months of follow-up, 69 (13.7%) patients had a CVE. Baseline SAF was significantly higher in patients with T2DM who later experienced a CVE (2.89 ± 0.70 arbitrary units versus 2.64 ± 0.62 in others, P = 0.002). This relationship was significant after adjusting for age, sex, conventional risk factors (diabetes duration, HbA1c, arterial hypertension, dyslipidemia, smoking, body mass index), vascular complications, C-reactive protein, and treatments for diabetes. The CVE-free survival curves differed between subjects whose SAF was above the whole-population median (log-rank: P = 0.002) and those whose SAF was above the macroangiopathy-free sub-population median (log-rank: P = 0.016). CONCLUSION: SAF of advanced glycation end-products was related to a higher incidence of later CVE in patients with T2DM.

Systemic Hemodynamics, Cardiac Mechanics, and Signaling Pathways Induced by Extracorporeal Membrane Oxygenation in a Cardiogenic Shock Model.

Peripheral venoarterial extracorporeal membrane oxygenation (VA-ECMO) is increasingly being used in patients suffering from refractory cardiogenic shock (CS). Although considered life-saving, peripheral VA-ECMO may also be responsible for intracardiac hemodynamic changes, including left ventricular overload and dysfunction. Venoarterial extracorporeal membrane oxygenation may also increase myocardial wall stress and stroke work, possibly affecting the cellular cardioprotective and apoptosis signaling pathways, and thus the infarct size. To test this hypothesis, we investigated the effects of increasing the peripheral VA-ECMO blood flow (25-100% of the baseline cardiac output) on systemic and cardiac hemodynamics in a closed-chest CS model. Upon completion of the experiment, the hearts were removed for assessment of infarct size, histology, apoptosis measurements, and phosphorylation statuses of p38 and protein Kinase B (Akt), and extracellular signal-regulated kinase mitogen-activated protein kinases (ERK-MAPK). Peripheral VA-ECMO restored systemic perfusion but induced a significant and blood flow-dependent increase in left ventricular preload and afterload. Venoarterial extracorporeal membrane oxygenation did not affect infarct size but significantly decreased p38-MAPK phosphorylation and cardiac myocyte apoptosis in the border zone.

The skin autofluorescence may help to select patients with Type 2 diabetes candidates for screening to revascularization procedures.

Chen et al. recently related the skin autofluorescence (SAF) of Advanced Glycation End-products to subclinical cardiovascular disease in the 3001 participants from the general population (Rotterdam study), with a particularly close relationship for the 413 subjects with diabetes. Because conventional vascular risk factors do not capture the risk in diabetes very well, this relationship may help to select high-risk individuals for the screening of silent myocardial ischemia, which has yet to prove its benefit in randomized controlled trials. Among 477 patients with uncontrolled and/or complicated Type 2 Diabetes, we measured the SAF ten years ago, and we registered new revascularizations during a 54-months follow-up. The patients with SAF > 2.6 Arbitrary units (AUs), the median population value, experienced more revascularizations of the coronary (17/24) and lower-limb arteries (13/17) than patients with a lower SAF, adjusted for age, sex, diabetes duration, vascular complications, and smoking habits: HR 2.17 (95% CI: 1.05-4.48), p = 0.035. The SAF has already been reported to predict cardiovascular events in three cohorts of people with diabetes. We suggest that its measurement may help to improve the performance of the screening before vascular explorations and revascularizations.

Thrombosis in the Coronary Microvasculature Impairs Cardiac Relaxation and Induces Diastolic Dysfunction.

BACKGROUND: Heart failure with preserved ejection fraction is proposed to be caused by endothelial dysfunction in cardiac microvessels. Our goal was to identify molecular and cellular mechanisms underlying the development of cardiac microvessel disease and diastolic dysfunction in the setting of type 2 diabetes. METHODS: We used Leprdb/db (leptin receptor-deficient) female mice as a model of type 2 diabetes and heart failure with preserved ejection fraction and identified Hhipl1 (hedgehog interacting protein-like 1), which encodes for a decoy receptor for HH (hedgehog) ligands as a gene upregulated in the cardiac vascular fraction of diseased mice. RESULTS: We then used Dhh (desert HH)-deficient mice to investigate the functional consequences of impaired HH signaling in the adult heart. We found that Dhh-deficient mice displayed increased end-diastolic pressure while left ventricular ejection fraction was comparable to that of control mice. This phenotype was associated with a reduced exercise tolerance in the treadmill test, suggesting that Dhh-deficient mice do present heart failure. At molecular and cellular levels, impaired cardiac relaxation in DhhECKO mice was associated with a significantly decreased PLN (phospholamban) phosphorylation on Thr17 (threonine 17) and an alteration of sarcomeric shortening ex vivo. Besides, as expected, Dhh-deficient mice exhibited phenotypic changes in their cardiac microvessels including a prominent prothrombotic phenotype. Importantly, aspirin therapy prevented the occurrence of both diastolic dysfunction and exercise intolerance in these mice. To confirm the critical role of thrombosis in the pathophysiology of diastolic dysfunction, we verified Leprdb/db also displays increased cardiac microvessel thrombosis. Moreover, consistently, with Dhh-deficient mice, we found that aspirin treatment decreased end-diastolic pressure and improved exercise tolerance in Leprdb/db mice. CONCLUSIONS: Altogether, these results demonstrate that microvessel thrombosis may participate in the pathophysiology of heart failure with preserved ejection fraction.

Predictive utility of stress tests in the detection of asymptomatic coronary artery disease in atherosclerotic stroke patients.

INTRODUCTION: Whether and how atherosclerotic ischemic stroke patients should be investigated for asymptomatic coronary artery disease (CAD) is controversial. Our aim was to carry out a prospective observational study to determine the frequency and predictors of functionally significant coronary stenosis in these patients as well as the predictors of major adverse cardiovascular events (MACE) during post-stroke follow-up. MATERIAL AND METHODS: From January 2014 to June 2018, patients with atherosclerotic ischemic stroke were referred from the stroke unit to our cardiovascular department 3+/- 1 months after the acute event where they benefited from evaluation of cardiovascular risk factors, vascular and myocardial disease. Main outcome was defined as the prevalence of myocardial ischemia defined by perfusion stress echography 3 months after stroke. Secondary outcome (MACE) was defined as the incidence of stroke, transient ischemic attack (TIA), acute coronary syndrome, cardiovascular (CV) death or coronary or peripheral revascularization during a 3 year follow-up. RESULTS: Three hundred and twenty five patients (92% of strokes and 8% TIA) were included and median follow-up was 1075 days. At 3 months post-stroke, myocardial ischemia was found in 17 patients (5.2%). During the 3 year follow-up, 11 MACE occurred (3.4%, all in the non-ischemic group) of which 6 were recurrent strokes. In multivariate analysis, myocardial ischemia was significantly associated with the number of atheromatous vascular beds (OR 4.3; 95% CI, 1.7 to 10.6) and ECG signs of necrosis (OR 6.5; 95% CI, 1.9 to 21.9). MACE were also associated with ECG signs of necrosis (OR 3.5; 95% CI, 1.3 to 9.1), and unrelated to myocardial ischemia. CONCLUSION: Myocardial ischemia and CV events were infrequent and both strongly associated with ECG signs of necrosis, suggesting a low yield of stress tests and the potential for a more straightforward algorithm in the choice of patients eligible to coronary angiogram or other coronary imaging in post-stroke setting.

Autophagy protein 5 controls flow-dependent endothelial functions.

Dysregulated autophagy is associated with cardiovascular and metabolic diseases, where impaired flow-mediated endothelial cell responses promote cardiovascular risk. The mechanism by which the autophagy machinery regulates endothelial functions is complex. We applied multi-omics approaches and in vitro and in vivo functional assays to decipher the diverse roles of autophagy in endothelial cells. We demonstrate that autophagy regulates VEGF-dependent VEGFR signaling and VEGFR-mediated and flow-mediated eNOS activation. Endothelial ATG5 deficiency in vivo results in selective loss of flow-induced vasodilation in mesenteric arteries and kidneys and increased cerebral and renal vascular resistance in vivo. We found a crucial pathophysiological role for autophagy in endothelial cells in flow-mediated outward arterial remodeling, prevention of neointima formation following wire injury, and recovery after myocardial infarction. Together, these findings unravel a fundamental role of autophagy in endothelial function, linking cell proteostasis to mechanosensing.

Partial Mural Cell Ablation Disrupts Coronary Vasculature Integrity and Induces Systolic Dysfunction.

Background Although the critical role of pericytes in maintaining vascular integrity has been extensively demonstrated in the brain and the retina, little is known about their role in the heart. We aim to investigate structural and functional consequences of partial pericyte depletion (≈60%) in the heart of adult mice. Methods and Results To deplete pericytes in adult mice, we used platelet-derived growth factor receptor ß-Cre/ERT2; RosaDTA mice and compared their phenotype with that of control mice (RosaDTA) chosen among their littermates. Cardiac function was assessed via echocardiography and left ventricular catheterization 1 month after the first tamoxifen injection. We found mice depleted with pericytes had a reduced left ventricular ejection fraction and an increased end-diastolic pressure, demonstrating both systolic and diastolic dysfunction. Consistently, mice depleted with pericytes presented a decreased left ventricular contractility and an increased left ventricular relaxation time (dP/dtmin). At the tissue level, mice depleted of pericytes displayed increased coronary endothelium leakage and activation, which was associated with increased CD45+ cell infiltration. Consistent with systolic dysfunction, pericyte depletion was associated with an increased expression of myosin heavy chain 7 and decreased expression of ATPase sarcoplasmic/endoplasmic reticulum Ca2+ transporting 2 and connexin 43. More important, coculture assays demonstrated, for the first time, that the decreased expression of connexin 43 is likely attributable to a direct effect of pericytes on cardiomyocytes. Besides, this study reveals that cardiac pericytes may undergo strong remodeling on injury. Conclusions Cardiac pericyte depletion induces both systolic and diastolic dysfunction, suggesting that pericyte dysfunction may contribute to the occurrence of cardiac diseases.

ROR2/PCP a New Pathway Controlling Endothelial Cell Polarity Under Flow Conditions.

BACKGROUND: Endothelial cells (ECs) are sensitive to physical forces created by blood flow, especially to laminar shear stress. Among the cell responses to laminar flow, EC polarization against the flow direction emerges as a key event, particularly during the development and remodeling of the vascular network. EC adopt an elongated planar cell shape with an asymmetrical distribution of intracellular organelles along the axis of blood flow. This study aimed to investigate the involvement of planar cell polarity via the receptor ROR2 (receptor tyrosine kinase-like orphan receptor 2) in endothelial responses to laminar shear stress. METHODS: We generated a genetic mouse model with EC-specific deletion of Ror2, in combination with in vitro approaches involving loss- and gain-of-function experiments. RESULTS: During the first 2 weeks of life, the endothelium of the mouse aorta undergoes a rapid remodeling associated with a loss of EC polarization against the flow direction. Notably, we found a correlation between ROR2 expression and endothelial polarization levels. Our findings demonstrate that deletion of Ror2 in murine ECs impaired their polarization during the postnatal development of the aorta. In vitro experiments further validated the essential role of ROR2 in both EC collective polarization and directed migration under laminar flow conditions. Exposure to laminar shear stress triggered the relocalization of ROR2 to cell-cell junctions where it formed a complex with VE-Cadherin and ß-catenin, thereby regulating adherens junctions remodeling at the rear and front poles of ECs. Finally, we showed that adherens junctions remodeling and cell polarity induced by ROR2 were dependent on the activation of the small GTPase Cdc42. CONCLUSIONS: This study identified ROR2/planar cell polarity pathway as a new mechanism controlling and coordinating collective polarity patterns of EC during shear stress response.

Ephrin-B1 regulates the adult diastolic function through a late postnatal maturation of cardiomyocyte surface crests.

The rod-shaped adult cardiomyocyte (CM) harbors a unique architecture of its lateral surface with periodic crests, relying on the presence of subsarcolemmal mitochondria (SSM) with unknown role. Here, we investigated the development and functional role of CM crests during the postnatal period. We found in rodents that CM crest maturation occurs late between postnatal day 20 (P20) and P60 through both SSM biogenesis, swelling and crest-crest lateral interactions between adjacent CM, promoting tissue compaction. At the functional level, we showed that the P20-P60 period is dedicated to the improvement of relaxation. Interestingly, crest maturation specifically contributes to an atypical CM hypertrophy of its short axis, without myofibril addition, but relying on CM lateral stretching. Mechanistically, using constitutive and conditional CM-specific knock-out mice, we identified ephrin-B1, a lateral membrane stabilizer, as a molecular determinant of P20-P60 crest maturation, governing both the CM lateral stretch and the diastolic function, thus highly suggesting a link between crest maturity and diastole. Remarkably, while young adult CM-specific Efnb1 KO mice essentially exhibit an impairment of the ventricular diastole with preserved ejection fraction and exercise intolerance, they progressively switch toward systolic heart failure with 100% KO mice dying after 13 months, indicative of a critical role of CM-ephrin-B1 in the adult heart function. This study highlights the molecular determinants and the biological implication of a new late P20-P60 postnatal developmental stage of the heart in rodents during which, in part, ephrin-B1 specifically regulates the maturation of the CM surface crests and of the diastolic function.

Praliciguat Promotes Ischemic Leg Reperfusion in Leptin Receptor-Deficient Mice.

BACKGROUND: Lower-limb peripheral artery disease is one of the major complications of diabetes. Peripheral artery disease is associated with poor limb and cardiovascular prognoses, along with a dramatic decrease in life expectancy. Despite major medical advances in the treatment of diabetes, a substantial therapeutic gap remains in the peripheral artery disease population. Praliciguat is an orally available sGC (soluble guanylate cyclase) stimulator that has been reported both preclinically and in early stage clinical trials to have favorable effects in metabolic and hemodynamic outcomes, suggesting that it may have a potential beneficial effect in peripheral artery disease. METHODS: We evaluated the effect of praliciguat on hind limb ischemia recovery in a mouse model of type 2 diabetes. Hind limb ischemia was induced in leptin receptor-deficient (Leprdb/db) mice by ligation and excision of the left femoral artery. Praliciguat (10 mg/kg/day) was administered in the diet starting 3 days before surgery. RESULTS: Twenty-eight days after surgery, ischemic foot perfusion and function parameters were better in praliciguat-treated mice than in vehicle controls. Improved ischemic foot perfusion was not associated with either improved traditional cardiovascular risk factors (ie, weight, glycemia) or increased angiogenesis. However, treatment with praliciguat significantly increased arteriole diameter, decreased ICAM1 (intercellular adhesion molecule 1) expression, and prevented the accumulation of oxidative proangiogenic and proinflammatory muscle fibers. While investigating the mechanism underlying the beneficial effects of praliciguat therapy, we found that praliciguat significantly downregulated Myh2 and Cxcl12 mRNA expression in cultured myoblasts and that conditioned medium form praliciguat-treated myoblast decreased ICAM1 mRNA expression in endothelial cells. These results suggest that praliciguat therapy may decrease ICAM1 expression in endothelial cells by downregulating Cxcl12 in myocytes. CONCLUSIONS: Our results demonstrated that praliciguat promotes blood flow recovery in the ischemic muscle of mice with type 2 diabetes, at least in part by increasing arteriole diameter and by downregulating ICAM1 expression.

Endothelial Dysfunction in Heart Failure With Preserved Ejection Fraction: What are the Experimental Proofs?

Heart failure with preserved ejection fraction (HFpEF) has been recognized as the greatest single unmet need in cardiovascular medicine. Indeed, the morbi-mortality of HFpEF is high and as the population ages and the comorbidities increase, so considerably does the prevalence of HFpEF. However, HFpEF pathophysiology is still poorly understood and therapeutic targets are missing. An unifying, but untested, theory of the pathophysiology of HFpEF, proposed in 2013, suggests that cardiovascular risk factors lead to a systemic inflammation, which triggers endothelial cells (EC) and coronary microvascular dysfunction. This cardiac small vessel disease is proposed to be responsible for cardiac wall stiffening and diastolic dysfunction. This paradigm is based on the fact that microvascular dysfunction is highly prevalent in HFpEF patients. More specifically, HFpEF patients have been shown to have decreased cardiac microvascular density, systemic endothelial dysfunction and a lower mean coronary flow reserve. Importantly, impaired coronary microvascular function has been associated with the severity of HF. This review discusses evidence supporting the causal role of endothelial dysfunction in the pathophysiology of HFpEF in human and experimental models.

Myocardial infarction associated with erenumab: A case report.

BACKGROUND: Monoclonal antibodies acting on the calcitonin gene-related peptide or its receptor (CGRP-mabs) are novel drugs for resistant migraine prophylaxis. As CGRP-mabs cause inhibition of vasodilatation, their use is reserved to patients with no recent history of cardiovascular diseases. We report a case of myocardial infarction associated with erenumab. CASE: A 57-year-old woman with a familial history of coronaropathy was first treated with erenumab 70 mg for 6 months and then increased to 140 mg. Almost 5 months after, the patient presented chest pain, increased troponin, and abnormal electrocardiogram. A myocardial infarction without coronarography abnormality was diagnosed through MRI. CONCLUSION: Further evidence is needed to assess the risk of myocardial infarction in patients treated with a CGRP-mab. In patients over 40 years of age, the risk of coronary or cardiovascular events should be assessed using risk tables or algorithms to take into account cardiovascular risk factors. This may be complemented by appropriate examinations to measure the burden of coronary atherosclerosis, if necessary.

Increased Capillary Permeability in Heart Induces Diastolic Dysfunction Independently of Inflammation, Fibrosis, or Cardiomyocyte Dysfunction.

BACKGROUND: While endothelial dysfunction is suggested to contribute to heart failure with preserved ejection fraction pathophysiology, understanding the importance of the endothelium alone, in the pathogenesis of diastolic abnormalities has not yet been fully elucidated. Here, we investigated the consequences of specific endothelial dysfunction on cardiac function, independently of any comorbidity or risk factor (diabetes or obesity) and their potential effect on cardiomyocyte. METHODS: The ubiquitine ligase Pdzrn3, expressed in endothelial cells (ECs), was shown to destabilize tight junction. A genetic mouse model in which Pdzrn3 is overexpressed in EC (iEC-Pdzrn3) in adults was developed. RESULTS: EC-specific Pdzrn3 expression increased cardiac leakage of IgG and fibrinogen blood-born molecules. The induced edema demonstrated features of diastolic dysfunction, with increased end-diastolic pressure, alteration of dP/dt min, increased natriuretic peptides, in addition to limited exercise capacity, without major signs of cardiac fibrosis and inflammation. Electron microscopic images showed edema with disrupted EC-cardiomyocyte interactions. RNA sequencing analysis of gene expression in cardiac EC demonstrated a decrease in genes coding for endothelial extracellular matrix proteins, which could be related to the fragile blood vessel phenotype. Irregularly shaped capillaries with hemorrhages were found in heart sections of iEC-Pdzrn3 mice. We also found that a high-fat diet was not sufficient to provoke diastolic dysfunction; high-fat diet aggravated cardiac inflammation, associated with an altered cardiac metabolic signature in EC-Pdzrn3 mice, reminiscent of heart failure with preserved ejection fraction features. CONCLUSIONS: An increase of endothelial permeability is responsible for mediating diastolic dysfunction pathophysiology and for aggravating detrimental effects of a high-fat diet on cardiac inflammation and metabolism.

PHACTR-1 (Phosphatase and Actin Regulator 1) Deficiency in Either Endothelial or Smooth Muscle Cells Does Not Predispose Mice to Nonatherosclerotic Arteriopathies in 3 Transgenic Mice.

BACKGROUND: Genome-wide association studies have revealed robust associations of common genetic polymorphisms in an intron of the PHACTR-1 (phosphatase and actin regulator 1) gene (chr6p24), with cervical artery dissection, spontaneous coronary artery dissection, and fibromuscular dysplasia. The aim was to assess its role in the pathogenesis of cervical artery dissection or fibromuscular dysplasia. METHODS: Using various tissue-specific Cre-driver mouse lines, Phactr1 was deleted either in endothelial cells using 2 tissue-specific Cre-driver (PDGFB [platelet-derived growth factor B]-CreERT2 mice and Tie2 [tyrosine kinase with immunoglobulin and EGF homology domains]-Cre) and smooth muscle cells (smooth muscle actin-CreERT2) with a third tissue-specific Cre-driver. RESULTS: To test the efficacy of the Phactr1 deletion after cre-induction, we confirmed first, a decrease in Phactr1 transcription and Phactr1 expression in endothelial cell and smooth muscle cell isolated from Phactr1iPDGFB and Phactr1iSMA mice. Irrespective to the tissue or the duration of the deletion, mice did not spontaneously display pathological phenotype or vascular impairment: mouse survival, growth, blood pressure, large vessel morphology, or actin organization were not different in knockout mice than their comparatives littermates. Challenging vascular function and repair either by angiotensin II-induced hypertension or limb ischemia did not lead to vascular morphology or function impairment in Phactr1-deleted mice. Similarly, there were no more consequences of Phactr1 deletion during embryogenesis in endothelial cells. CONCLUSIONS: Loss of PHACTR-1 function in the cells involved in vascular physiology does not appear to induce a pathological vascular phenotype. The in vivo effect of the intronic variation described in genome-wide association studies is unlikely to involve downregulation in PHACTR-1 expression.

Phenotypic and genotypic characterization of familial hypercholesterolemia in French adult and pediatric populations.

BACKGROUND: Familial hypercholesterolemia (FH) is the most common genetic disorder associated with a high risk for premature atherosclerotic cardiovascular disease attributable to increased levels of LDL-cholesterol (LDL-C) from birth. FH is both underdiagnosed and undertreated. OBJECTIVE: We describe the clinical, biological, and genetic characteristics of 147 patients in France with clinical FH (including a group of 26 subjects aged < 20 years); we explore how best to detect patients with monogenic FH. METHODS: We retrospectively reviewed all available data on patients undergoing genetic tests for FH from 2009 to 2019. FH diagnoses were based on the Dutch Lipid Clinics Network (DLCN) scores of adults, and elevated LDL-C levels in subjects < 20 years of age. We evaluated LDLR, APOB, and PCSK9 status. RESULTS: The mutations of adults (in 25.6% of all adults) were associated with DLCN scores indicating "possible FH," "probable FH, and "definitive FH" at rates of 4%, 16%, and 53%, respectively. The areas under the ROC curves of the DLCN score and the maximum LDL-C level did not differ (p = 0.32). We found that the pediatric group evidenced more monogenic etiologies (77%, increasing to 91% when an elevated LDL-C level was combined with a family history of hypercholesterolemia and/or premature coronary artery disease). CONCLUSION: Diagnosis of monogenic FH may be optimized by screening children in terms of their LDL-C levels, associated with reverse-cascade screening of relatives when the children serve as index cases.