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Echolucency, a measure of plaque instability associated with increased cardiovascular risk, can be assessed in both the carotid plaque and the plaque-free common carotid intima–media (IM) complex as a gray-scale median (plaque-GSM and IM-GSM, respectively). The impact of specific vascular risk factors on these two phenotypes remains uncertain, including the nature and extent of their influence. This study aims to seek the determinants of plaque-GSM and IM-GSM. Plaque-GSM and IM-GSM were measured in subjects from the IMPROVE study cohort (aged 54–79, 46% men) recruited in five European countries. Plaque-GSM was measured in subjects who had at least one IMTmax ≥ 1.5 mm (n = 2138), whereas IM-GSM was measured in all subjects included in the study (n = 3188). Multiple regression with internal cross-validation was used to find independent predictors of plaque-GSM and IM-GSM. Plaque-GSM determinants were plaque-size (IMTmax), and diastolic blood pressure. IM-GSM determinants were the thickness of plaque-free common carotid intima–media complex (PF CC-IMTmean), height, systolic blood pressure, waist/hip ratio, treatment with fibrates, mean corpuscular volume, treatment with alpha-2 inhibitors (sartans), educational level, and creatinine. Latitude, and pack-yearscode were determinants of both plaque-GSM and IM-GSM. The overall models explain 12.0% of plaque-GSM variability and 19.7% of IM-GSM variability. A significant correlation (r = 0.51) was found between plaque-GSM and IM-GSM. Our results indicate that IM-GSM is a weighty risk marker alternative to plaque-GSM, offering the advantage of being readily measurable in all subjects, including those in the early phases of atherosclerosis where plaque occurrence is relatively infrequent.

Introduction Prevention of cardiovascular disease (CVD) is of key importance in reducing morbidity, disability and mortality worldwide. Observational studies suggest that digital health interventions can be an effective strategy to reduce cardiovascular (CV) risk. However, evidence from large randomised clinical trials is lacking.Methods and analysis The CV-PREVITAL study is a multicentre, prospective, randomised, controlled, open-label interventional trial designed to compare the effectiveness of an educational and motivational mobile health (mHealth) intervention versus usual care in reducing CV risk. The intervention aims at improving diet, physical activity, sleep quality, psycho-behavioural aspects, as well as promoting smoking cessation and adherence to pharmacological treatment for CV risk factors. The trial aims to enrol approximately 80 000 subjects without overt CVDs referring to general practitioners’ offices, community pharmacies or clinics of Scientific Institute for Research, Hospitalization and Health Care (Italian acronym IRCCS) affiliated with the Italian Cardiology Network. All participants are evaluated at baseline and after 12 months to assess the effectiveness of the intervention on short-term endpoints, namely improvement in CV risk score and reduction of major CV risk factors. Beyond the funded life of the study, a long-term (7 years) follow-up is also planned to assess the effectiveness of the intervention on the incidence of major adverse CV events. A series of ancillary studies designed to evaluate the effect of the mHealth intervention on additional risk biomarkers are also performed.Ethics and dissemination This study received ethics approval from the ethics committee of the coordinating centre (Monzino Cardiology Center; R1256/20-CCM 1319) and from all other relevant IRBs and ethics committees. Findings are disseminated through scientific meetings and peer-reviewed journals and via social media. Partners are informed about the study’s course and findings through regular meetings.Trial registration number NCT05339841.

Background The association between common carotid artery intima‐media thickness (CCA‐IMT) and incident carotid plaque has not been characterized fully. We therefore aimed to precisely quantify the relationship between CCA‐IMT and carotid plaque development. Methods and Results We undertook an individual participant data meta‐analysis of 20 prospective studies from the Proof‐ATHERO (Prospective Studies of Atherosclerosis) consortium that recorded baseline CCA‐IMT and incident carotid plaque involving 21 494 individuals without a history of cardiovascular disease and without preexisting carotid plaque at baseline. Mean baseline age was 56 years (SD, 9 years), 55% were women, and mean baseline CCA‐IMT was 0.71 mm (SD, 0.17 mm). Over a median follow‐up of 5.9 years (5th–95th percentile, 1.9–19.0 years), 8278 individuals developed first‐ever carotid plaque. We combined study‐specific odds ratios (ORs) for incident carotid plaque using random‐effects meta‐analysis. Baseline CCA‐IMT was approximately log‐linearly associated with the odds of developing carotid plaque. The age‐, sex‐, and trial arm–adjusted OR for carotid plaque per SD higher baseline CCA‐IMT was 1.40 (95% CI, 1.31–1.50; I2=63.9%). The corresponding OR that was further adjusted for ethnicity, smoking, diabetes, body mass index, systolic blood pressure, low‐ and high‐density lipoprotein cholesterol, and lipid‐lowering and antihypertensive medication was 1.34 (95% CI, 1.24–1.45; I2=59.4%; 14 studies; 16 297 participants; 6381 incident plaques). We observed no significant effect modification across clinically relevant subgroups. Sensitivity analysis restricted to studies defining plaque as focal thickening yielded a comparable OR (1.38 [95% CI, 1.29–1.47]; I2=57.1%; 14 studies; 17 352 participants; 6991 incident plaques). Conclusions Our large‐scale individual participant data meta‐analysis demonstrated that CCA‐IMT is associated with the long‐term risk of developing first‐ever carotid plaque, independent of traditional cardiovascular risk factors.

Abstract: Mutations in the LCAT gene cause familial LCAT deficiency (Online Mendelian Inheritance in Man ID: #245900), a very rare metabolic disorder. LCAT is the only enzyme able to esterify cholesterol in plasma, whereas sterol O-acyltransferases 1 and 2 are the enzymes esterifying cellular cholesterol in cells. Despite the complete lack of LCAT activity, patients with familial LCAT deficiency exhibit circulating cholesteryl esters (CEs) in apoB-containing lipoproteins. To analyze the origin of these CEs, we investigated 24 carriers of LCAT deficiency in this observational study. We found that CE plasma levels were significantly reduced and highly variable among carriers of two mutant LCAT alleles (22.5 [4.0–37.8] mg/dl) and slightly reduced in heterozygotes (218 [153–234] mg/dl). FA distribution in CE (CEFA) was evaluated in whole plasma and VLDL in a subgroup of the enrolled subjects. We found enrichment of C16:0, C18:0, and C18:1 species and a depletion in C18:2 and C20:4 species in the plasma of carriers of two mutant LCAT alleles. No changes were observed in heterozygotes. Furthermore, plasma triglyceride-FA distribution was remarkably similar between carriers of LCAT deficiency and controls. CEFA distribution in VLDL essentially recapitulated that of plasma, being mainly enriched in C16:0 and C18:1, while depleted in C18:2 and C20:4. Finally, after fat loading, chylomicrons of carriers of two mutant LCAT alleles showed CEs containing mainly saturated FAs. This study of CEFA composition in a large cohort of carriers of LCAT deficiency shows that in the absence of LCAT-derived CEs, CEs present in apoB-containing lipoproteins are derived from hepatic and intestinal sterol O-acyltransferase 2.

Abstract The relationship between intake of saturated fats and subclinical atherosclerosis, as well as the possible influence of genetic variants, is poorly understood and investigated. We aimed to investigate this relationship, with a hypothesis that it would be positive, and to explore whether genetics may modulate it, using data from a European cohort including 3,407 participants aged 54–79 at high risk of cardiovascular disease. Subclinical atherosclerosis was assessed by carotid intima-media thickness (C-IMT), measured at baseline and after 30 months. Logistic regression (OR; 95% CI) was employed to assess the association between high intake of food rich in saturated fat (vs. low) and: (1) the mean and the maximum values of C-IMT in the whole carotid artery (C-IMTmean, C-IMTmax), in the bifurcation (Bif-), the common (CC-) and internal (ICA-) carotid arteries at baseline (binary, cut-point ≥ 75th), and (2) C-IMT progression (binary, cut-point > zero). For the genetic-diet interaction analyses, we considered 100,350 genetic variants. We defined interaction as departure from additivity of effects. After age- and sex-adjustment, high intake of saturated fat was associated with increased C-IMTmean (OR:1.27;1.06–1.47), CC-IMTmean (OR:1.22;1.04–1.44) and ICA-IMTmean (OR:1.26;1.07–1.48). However, in multivariate analysis results were no longer significant. No clear associations were observed between high intake of saturated fat and risk of atherosclerotic progression. There was no evidence of interactions between high intake of saturated fat and any of the genetic variants considered, after multiple testing corrections. High intake of saturated fats was not independently associated with subclinical atherosclerosis. Moreover, we did not identify any significant genetic-dietary fat interactions in relation to risk of subclinical atherosclerosis.

Abstract Understanding why individuals with severe mental illness (Schizophrenia, Bipolar Disorder and Major Depressive Disorder) have increased risk of cardiometabolic disease (including obesity, type 2 diabetes and cardiovascular disease), and identifying those at highest risk of cardiometabolic disease are important priority areas for researchers. For individuals with European ancestry we explored whether genetic variation could identify sub-groups with different metabolic profiles. Loci associated with schizophrenia, bipolar disorder and major depressive disorder from previous genome-wide association studies and loci that were also implicated in cardiometabolic processes and diseases were selected. In the IMPROVE study (a high cardiovascular risk sample) and UK Biobank (general population sample) multidimensional scaling was applied to genetic variants implicated in both psychiatric and cardiometabolic disorders. Visual inspection of the resulting plots used to identify distinct clusters. Differences between these clusters were assessed using chi-squared and Kruskall-Wallis tests. In IMPROVE, genetic loci associated with both schizophrenia and cardiometabolic disease (but not bipolar disorder or major depressive disorder) identified three groups of individuals with distinct metabolic profiles. This grouping was replicated within UK Biobank, with somewhat less distinction between metabolic profiles. This work focused on individuals of European ancestry and is unlikely to apply to more genetically diverse populations. Overall, this study provides proof of concept that common biology underlying mental and physical illness may help to stratify subsets of individuals with different cardiometabolic profiles.

Familial LCAT deficiency (FLD) is a rare genetic disorder of HDL metabolism, caused by loss-of-function mutations in the LCAT gene and characterized by a variety of symptoms including corneal opacities and kidney failure. Renal disease represents the leading cause of morbidity and mortality in FLD cases. However, the prognosis is not known and the rate of deterioration of kidney function is variable and unpredictable from patient to patient. In this article, we present data from a follow-up of the large Italian cohort of FLD patients, who have been followed for an average of 12 years. We show that renal failure occurs at the median age of 46 years, with a median time to a second recurrence of 10 years. Additionally, we identify high plasma unesterified cholesterol level as a predicting factor for rapid deterioration of kidney function. In conclusion, this study highlights the severe consequences of FLD, underlines the need of correct early diagnosis and referral of patients to specialized centers, and highlights the urgency for effective treatments to prevent or slow renal disease in patients with LCAT deficiency.

Depression is associated with thrombotic risk and arterial events, its proper management is strongly recommended in coronary artery disease (CAD) patients. We have previously shown that the Brain-Derived Neurotrophic Factor (BDNF)Val66Met polymorphism, related to depression, is associated with arterial thrombosis in mice, and with an increased risk of acute myocardial infarction in humans. Herein, expanding the previous findings on BDNFVal66Met polymorphism, we show that desipramine, a norepinephrine reuptake-inhibitor, rescues behavioral impairments, reduces the arterial thrombosis risk, abolishes pathological coagulation and platelet hyper-reactivity, normalizes leukocyte, platelet, and bone marrow megakaryocyte number and restores physiological norepinephrine levels in homozygous knock-in BDNF Val66Met (BDNFMet/Met) mice. The in vitro data confirm the enhanced procoagulant activity and the alpha2A-adrenergic receptor (α2A-ADR) overexpression found in BDNFMet/Met mice and we provide evidence that, in presence of Met variant, norepinephrine is crucial to up-regulate procoagulant activity and to enhance platelet generation. The α2-ADR antagonist rauwolscine rescues the prothrombotic phenotype in BDNFMet/Met mice and reduces procoagulant activity and platelet generation in cells transfected with BDNFMet plasmid or exposed to pro-BDNFMet peptide. Finally, we show that homozygous BDNFMet/Met CAD patients have hyper-reactive platelets overexpressing abundant α2A-ADR. The great proplatelet release from their megakaryocytes well reflects their higher circulating platelet number compared to BDNFVal/Val patients. These data reveal an unprecedented described role of Met allele in the dysregulation of norepinephrine/α2A-ADR pathway that may explain the predisposition to arterial thrombosis. Overall, the development of α2A-ADR inhibitors might represent a pharmacological treatment for depression-associated thrombotic conditions in this specific subgroup of CAD patients.

Abstract Background High-sensitivity C-reactive protein (hs-CRP) elevation frequently occurs in acute myocardial infarction (AMI) and is associated with adverse outcomes. Since diabetes mellitus (DM) is characterized by an underlying chronic inflammation, hs-CRP may have a different prognostic power in AMI patients with and without DM. Methods We prospectively included 2064 AMI patients; hs-CRP was measured at hospital admission. Patients were grouped according to hs-CRP quartiles and DM status. The primary endpoint was a composite of in-hospital mortality, cardiogenic shock, and acute pulmonary edema. Two-year all-cause mortality was the secondary endpoint. Results Twenty-six percent (n = 548) of patients had DM and they had higher hs-CRP levels than non-DM patients (5.32 vs. 3.24 mg/L; P

Background and Purpose: The Active Connection Matrixes (ACMs) are unsupervised artificial adaptive systems able to extract from digital images features of interest (edges, tissue differentiation, etc.) unnoticeable with conventional systems. In this proof-of-concept study, we assessed the potentiality of ACMs to increase measurement precision of morphological structures (e.g., stenosis and lumen diameter) and to grasp morphological features (arterial walls) from quantitative coronary angiography (QCA), unnoticeable on the original images.Methods: Archive images of QCA and intravascular ultrasound (IVUS) of 10 patients (8 men, age 69.1 ± 9.7 years) who underwent both procedures for clinical reasons were retrospectively analyzed. Arterial features derived from “IVUS images,” “conventional QCA images,” and “ACM-reprocessed QCA images” were measured in 21 coronary segments. Portions of 1-mm length (263 for lumen and 526 for arterial walls) were head-to-head compared to assess quali-quantitative between-methods agreement.Results: When stenosis was calculated on “ACM-reprocessed QCA images,” the bias vs. IVUS (gold standard) did not improve, but the correlation coefficient of the QCA–IVUS relationship increased from 0.47 to 0.83. When IVUS-derived lumen diameters were compared with diameters obtained on ACM-reprocessed QCA images, the bias (−0.25 mm) was significantly smaller (p < 0.01) than that observed with original QCA images (0.58 mm). ACMs were also able to extract arterial wall features from QCA. The bias between the measures of arterial walls obtained with IVUS and ACMs, although significant (p < 0.01), was small [0.09 mm, 95% CI (0.03, 0.14)] and the correlation was fairly good (r = 0.63; p < 0.0001).Conclusions: This study provides proof of concept that ACMs increase the measurement precision of coronary lumen diameter and allow extracting from QCA images hidden features that mirror well the arterial walls derived by IVUS.

Background: Current knowledge regarding the relationship between aortic valve sclerosis (AVSc), cardiovascular risk factors, and mortality in patients with known coronary artery disease (CAD) is still unclear. The present study aimed at investigating the prevalence of AVSc as well as its association with long-term all-cause mortality in high-risk CAD patients that has never been explored in large cohorts thus far.Methods and Results: In this retrospective and observational cohort study we enrolled high-risk CAD patients, hospitalized at Centro Cardiologico Monzino (CCM), Milan, Italy, between January 2006 and December 2016. The morphology and function of the aortic valve were assessed from the recorded echocardiographic images to evaluate the presence of AVSc, defined as a non-uniform thickening of the aortic leaflets with no consequences on hemodynamics. Data on 5-year all-cause mortality was retrieved from a Regional database. Of the 5,489 patients initially screened, 4,938 (mean age 67 ± 11 years, 3,954 [80%] men) were enrolled in the study. In the overall population, AVSc was detected in 2,138 (43%) patients. Multivariable LASSO regression revealed that age, female gender, diabetes mellitus, previous MI, and left ventricular ejection fraction were independently associated with AVSc. All-cause mortality (adjusted hazard ratio: 1.29, 95%CI: 1.05–1.58) was significantly higher in AVSc than in non-AVSc patients.Conclusions: AVSc is frequently detected in high-risk CAD patients and is associated with long-term mortality. Our findings corroborate the hypothesis that AVSc is an underestimated marker of systemic cardiovascular risk. Thus, AVSc detection may be used to improve long-term risk stratification of high-risk CAD patients.

Background The association between elevated serum uric acid (SUA), cardiovascular disease (CVD) risk, and carotid atherosclerosis has long been explored, and contrasting results have been reported. Therefore, the role of SUA as an independent risk factor for vascular events (VEs) and carotid atherosclerosis deserves further attention. We investigated the relationship between SUA, incident VEs, carotid intima‐media thickness (cIMT), and cIMT progression in subjects at moderate‐to‐high CVD risk. Methods and Results In the IMPROVE (IMT‐Progression as Predictors of VEs) study, 3686 participants (median age 64 years; 48% men) with ≥ 3 vascular risk factors, free from VEs at baseline, were grouped according to SUA quartiles (division points: 244–284–328 µmol/L in women, 295–336–385 µmol/L in men). Carotid‐IMT and its 15‐month progression, along with incident VEs, were recorded. A U‐shaped association between SUA and VEs was observed in men, with 2.4‐fold (P = 0.004) and 2.5‐fold (P = 0.002) increased CVD risk in the first and fourth SUA quartiles as compared with the second. Adjusted hazard ratios (HRs) for cerebro‐VEs in men were the highest (first and fourth quartile versus second: HR, 5.3, P = 0.010 and HR, 4.4, P = 0.023, respectively). SUA level was independently associated with cIMT progression in men (β = 0.068, P = 0.014). No significant association between SUA levels, CVD end points, and cIMT progression were found in women. Conclusions Both low and high SUA levels are associated with an increased risk of VEs in men at moderate‐to‐high CVD risk but not in women. Only elevated SUA levels predict cIMT progression and at a lesser but not significant extent in women.

Carotid intima-media thickness (cIMT) and plaque are associated with subclinical atherosclerosis and coronary heart disease (CHD). Here, the authors identify and prioritize genetic loci for cIMT and plaque by GWAS and colocalization approaches and further demonstrate genetic correlation with CHD and stroke.

Background: Takotsubo (TTS) syndrome is an acute cardiac condition characterized by transient and reversible left ventricle dysfunction that mainly affects postmenopausal women. Catecholamine burst is the most accredited mechanism underpinning TTS onset and leading to endothelial dysfunction and platelet activation. Even if the use of low dose acetylsalycilic acid (ASA) in this clinical setting is based on both clinical presentation and unfavorable long-term prognosis, its efficacy has been recently challenged. Aim: This study was designed to assess endothelial function, residual thromboxane formation and platelet aggregation in TTS women on low-dose ASA treatment at long-term follow-up. Methods: Twenty-eight females with previously diagnosis of TTS syndrome were enrolled. Data were compared to those obtained from 23 coronary artery disease (CAD) women with a history of acute myocardial infarction, and 26 control subjects with no TTS or clinically evident CAD. Psychological and clinical profile were assessed in all study groups at the enrollment. Main metabolites involved in L-arginine/nitric oxide pathway, urinary prostacyclin, serum and urine thromboxane metabolites were measured by LCMS/MS methods. Thrombomodulin levels were quantified using an ELISA kit, and platelet aggregation, carried out on platelet rich-plasma, was induced by ADP or by epinephrine (EPI), norepinephrine (NORE) and TRAP-6, alone or in association with ADP and evaluated by Born’s method. Results: In TTS women an endothelial derangement, characterized by reduced citrulline production and increased thrombomodulin concentration, with no perturbation in prostacyclin levels, was evidenced. In addition, despite ASA treatment, TTS displayed a higher residual thromboxane formation, in parallel with an enhanced platelet response to compared to CAD. Conclusions: Our study highlighted the presence of endothelial perturbation in TTS patients even at long-term from the index event. The residual thromboxane production and platelet aggregation still leave open the question about the use of low dose ASA in this clinical setting.

In the last years, a substantial contribution of red blood cells (RBCs) in cardiovascular homeostasis has been evidenced, as these cells are able to regulate cardiovascular function by the export of adenosine triphosphate and nitric oxide as well as to maintain redox balance through a well-developed antioxidant system. Recently a link between high-risk plaque (HRP) features and myocardial ischemia, in the absence of severe lumen stenosis, has been evidenced. Nonobstructive coronary artery disease (nonob CAD) has been associated in fact with a greater 1-year risk of myocardial infarction and all-cause mortality compared with no apparent CAD. This new evidence increases interest in searching new triggers to identify these high-risk patients, in the absence/or on top of traditional hazard markers. In this study, we investigated the existence of any association between RBC morphodynamics and HRP features in individuals with different grades of coronary stenosis detected by coronary computed tomography angiography (CCTA). Ninety-one consecutive individuals who underwent CCTA [33 no CAD; 26 nonobstructive (nonob), and 32 obstructive (ob) CAD] were enrolled. RBC morphodynamic features, i.e., RBC aggregability and deformability, were analyzed by means of Laser Assisted Optical Rotation Cell Analyzer (LoRRca MaxSis). The putative global RBC morphodynamic (RMD) score and the related risk chart, associating the extent of HRP (e.g., the non-calcified plaque volume) with both the RMD score and the max % stenosis were computed. In nonob CAD group only positive correlations between RBC rigidity, osmotic fragility or aggregability and HRP features (plaque necrotic core, fibro-fatty and fibro-fatty plus necrotic core plaque volumes) were highlighted. Interestingly, in this patient cohort three of these RBC morphodynamic features result to be independent predictors of the presence of non-calcified plaque volume in this patients group. The risk chart created shows that only in nonob CAD plaque vulnerability increases according to the score quartile. Findings of this work, by evidencing the association between erythrocyte morphodynamic characteristics assessed by LoRRca and plaque instability in a high-risk cohort of nonob CAD, suggest the use of these blood cell features in the identification of high-risk patients, in the absence of severe coronary stenosis.

Coronary artery bypass graft (CABG) surgery still represents the gold standard for patients with complex multivessel coronary artery disease. However, graft occlusion still occurs in a significant proportion of CABG conduits, and oxidative stress is currently considered to be a potential contributor. Human serum albumin (HSA) represents the main antioxidant in plasma through its reduced amino acid Cys34, which can efficiently scavenge several oxidants. In a nested case–control study including 36 patients with occluded grafts and 38 age- and sex-matched patients without occlusion, we assessed the levels of the native mercaptoalbumin (HSA-SH) and oxidized thiolated form of albumin (Thio-HSA) in relation with graft occlusion within 5 years after CABG. We found that the plasma level of preoperative HSA-SH was significantly lower in patients with occluded graft at 5 years follow-up than in patients with graft patency. Furthermore, low HSA-SH remained independently associated with graft occlusion even after adjusting for preoperative D-dimer, a well-known marker of activated coagulation recently found to be associated with graft occlusion. In conclusion, the preoperative level of HSA-SH is independently associated with graft occlusion in CABG and represents a measurable and potentially druggable predictor.

An in-depth analysis of gathered data collected in several countries on the pre-infectious characteristics of patients who have developed severe forms of COVID-19 disease could be the basis for developing tools to estimate individual risk and tailor protective measures for a safer route through the Phase 2 of the pandemic.

Abstract Background The reduction of RV function after cardiac surgery is a well-known phenomenon. It could persist up-to one year after the operation and often leads to an incomplete recovery at follow-up echocardiographic control. The aim of the present study is to analyze the impact of different modalities of pericardial incision (lateral versus anterior) and of myocardial protection protocols (Buckberg versus Custodiol) onto postoperative RV dynamic by relating two- and three-dimensional echocardiographic parameters in patients undergoing mitral valve repair through minimally invasive or traditional surgery approach. Methods We have analyzed 44 consecutive patients with severe degenerative mitral regurgitation who underwent mitral reparation with different surgical approach and cardioplegia type: Group 1 (17 pts): sternotomy with Buckberg cardioplegia protocol; Group 2 (10 pts): sternotomy with Custodiol cardioplegia; Group 3 (17 pts): mini-invasive surgery with Custodiol cardioplegia. Two-dimensional transthoracic echocardiography was performed pre- and 6 months post-surgery to evaluate RV function by tricuspid annular plane systolic excursion (TAPSE). Results All patients underwent successful and uneventful. A postoperative TAPSE reduction was found in all groups. However, mini-invasive patients experienced a significant reduced variation versus traditional surgery. Conclusions Mini-invasive mitral repair, with lateral incision of pericardium, reduces postoperative TAPSE fall, while cardioplegia protocol fails to have an impact onto longitudinal RV function. In our study, the RV seems to experience a clinically irrelevant geometrical modification too, whose entity appears to be less evident in case of lateral pericardial approach. These results could strengthen the use of minimally invasive approach also to preserve RV function.

(1) Background and purpose: circulating proprotein convertase subtilisin/kexin type 9 (PCSK9) is one of the key regulators of cholesterol metabolism. Despite this, its role as a player in atherosclerosis development is still matter of debate. Here, we investigated the relationships between this protein and several markers of subclinical atherosclerosis. (2) Methods: the IMPROVE study enrolled 3703 European subjects (54–79 years; 48% men; with ≥3 vascular risk factors), asymptomatic for cardiovascular diseases. PCSK9 levels were measured by ELISA. B-mode ultrasound was used to measure markers of carotid subclinical atherosclerosis. (3) Results: in the crude analysis, PCSK9 levels were associated with several baseline measures of carotid intima-media thickness (cIMT) (all p < 0.0001); with cIMT change over time (Fastest-IMTmax-progr) (p = 0.01); with inter-adventitia common carotid artery diameter (ICCAD) (p < 0.0001); and with the echolucency (Grey Scale Median; GSM) of both carotid plaque and plaque-free common carotid IMT (both p < 0.0001). However, after adjustment for age, sex, latitude, and pharmacological treatment, all the afore-mentioned correlations were no longer statistically significant. The lack of correlation was also observed after stratification for sex, latitude, and pharmacological treatments. (4) Conclusions: in subjects who are asymptomatic for cardiovascular diseases, PCSK9 plasma levels do not correlate with vascular damage and/or subclinical atherosclerosis of extracranial carotid arteries.

The Mediterranean diet (MD) prevents cardiovascular disease by different putative mechanisms, including modifications in the blood fatty acid (FA) profile. Polytherapy for secondary cardiovascular prevention might mask the effect of MD on the FA profile. This study was aimed to assess whether MD, in comparison with a low-fat diet (LFD), favorably modifies the blood FA profile in patients with coronary heart disease (CHD) on polytherapy. One hundred and twenty patients with a recent history of coronary stenting, randomized to MD or to LFD, completed 3 months of this open-label dietary intervention study. Diet Mediterranean-ness was evaluated using the Mediterranean Diet Adherence Screener (MeDAS) score. Both diets significantly reduced saturated FA (p < 0.01). Putative favorable changes in total n-3 FA (p = 0.03) and eicosapentaenoic acid plus docosahexaenoic acid (EPA + DHA; p = 0.04) were significantly larger with MD than with LFD. At 3 months, in the whole cohort, the MeDAS score correlated inversely with palmitic acid (R = −0.21, p = 0.02), and with palmitoleic acid (R = −0.32, p = 0.007), and positively with total n-3 FA (R = 0.19, p = 0.03), EPA (R = 0.28, p = 0.002), and EPA + DHA (R = 0.21, p = 0.02). In CHD patients on polytherapy, both MD and LFD shift FA blood composition towards a healthier profile, with a more favorable effect of MD on omega−3 levels.

The identification of new biomarkers allowing an early and more accurate characterization of patients with ST-segment elevation myocardial infarction (STEMI) is still needed, and exosomes represent an attractive diagnostic tool in this context. However, the characterization of their protein cargo in relation to cardiovascular clinical manifestation is still lacking. To this end, 35 STEMI patients (17 experiencing resuscitated out-of-hospital cardiac arrest (OHCA-STEMI) and 18 uncomplicated) and 32 patients with chronic coronary syndrome (CCS) were enrolled. Plasma exosomes were characterized by the nanoparticle tracking analysis and Western blotting. Exosomes from STEMI patients displayed a higher concentration and size and a greater expression of platelet (GPIIb) and vascular endothelial (VE-cadherin) markers, but a similar amount of cardiac troponin compared to CCS. In addition, a difference in exosome expression of acute-phase proteins (ceruloplasmin, transthyretin and fibronectin) between STEMI and CCS patients was found. GPIIb and brain-associated marker PLP1 accurately discriminated between OHCA and uncomplicated STEMI. In conclusion, the exosome profile of STEMI patients has peculiar features that differentiate it from that of CCS patients, reflecting the pathophysiological mechanisms involved in STEMI. Additionally, the exosome expression of brain- and platelet-specific markers might allow the identification of patients experiencing ischemic brain injury in STEMI.

Coronary artery disease (CAD) patients are at high ischemic risk, and new biomarkers reflecting atherosclerotic disease severity and coronary plaque vulnerability are required. The Brain-Derived Neurotrophic Factor (BDNF) affects endothelial and macrophage activation suggesting its involvement in atherosclerotic plaque behavior. To investigate whether plasma BDNF is associated with in vivo coronary plaque features, assessed by optical coherence tomography (OCT), in both acute myocardial infarction (AMI) and stable angina (SA) patients, we enrolled 55 CAD patients (31 SA and 24 AMI), and 21 healthy subjects (HS). BDNF was lower in CAD patients than in HS (p < 0.0001), and it decreased with the presence, clinical acuity and severity of CAD. The greater BDNF levels were associated with OCT features of plaque vulnerability in overall CAD as well as in SA and AMI patients (p < 0.03). Specifically, in SA patients, BDNF correlated positively with macrophages’ infiltration within atherosclerotic plaque (p = 0.01) and inversely with minimal lumen area (p = 0.02). In AMI patients a negative correlation between BDNF and cap thickness was found (p = 0.02). Despite a small study population, our data suggest a relationship between BDNF and coronary plaque vulnerability, showing that vulnerable plaque is positively associated with plasma BDNF levels, regardless of the clinical CAD manifestation.

Acute phase reaction (APR) is a systemic inflammation triggered by several conditions associated with lipid profile alterations. We evaluated whether APR also associates with changes in cholesterol synthesis and absorption, HDL structure, composition, and cholesterol efflux capacity (CEC). We analyzed 59 subjects with APR related to infections, oncologic causes, or autoimmune diseases and 39 controls. We detected no difference in markers of cholesterol synthesis and absorption. Conversely, a significant reduction of LpA-I- and LpAI:AII-containing HDL (−28% and −44.8%, respectively) and of medium-sized HDL (−10.5%) occurred in APR. Total HDL CEC was impaired in APR subjects (−18%). Evaluating specific CEC pathways, we found significant reductions in CEC by aqueous diffusion and by the transporters scavenger receptor B-I and ABCG1 (−25.5, −41.1 and −30.4%, respectively). ABCA1-mediated CEC was not affected. Analyses adjusted for age and gender provided similar results. In addition, correcting for HDL-cholesterol (HDL-C) levels, the differences in aqueous diffusion total and ABCG1-CEC remained significant. APR subjects displayed higher levels of HDL serum amyloid A (+20-folds; P = 0.003). In conclusion, APR does not associate with cholesterol synthesis and absorption changes but with alterations of HDL composition and a marked impairment of HDL CEC, partly independent of HDL-C serum level reduction.

The aim of this study was to evaluate the vasoprotective effects of HDL isolated from carriers of LCAT deficiency, which are characterized by a selective depletion of LpA-I:A-II particles and predominance of preβ migrating HDL. HDLs were isolated from LCAT-deficient carriers and tested in vitro for their capacity to promote NO production and to inhibit vascular cell adhesion molecule-1 (VCAM-1) expression in cultured endothelial cells. HDLs from carriers were more effective than control HDLs in promoting eNOS activation with a gene-dose-dependent effect (PTrend = 0.048). As a consequence, NO production induced by HDL from carriers was significantly higher than that promoted by control HDL (1.63 ± 0.24-fold vs. 1.34 ± 0.07-fold, P= 0.031). HDLs from carriers were also more effective than control HDLs in inhibiting the expression of VCAM-1 (homozygotes, 65.0 ± 8.6%; heterozygotes, 53.1 ± 7.2%; controls, 44.4 ± 4.1%; PTrend = 0.0003). The increased efficiency of carrier HDL was likely due to the depletion in LpA-I:A-II particles. The in vitro findings might explain why carriers of LCAT deficiency showed flow-mediated vasodilation and plasma-soluble cell adhesion molecule concentrations comparable to controls, despite low HDL-cholesterol levels. These results indicate that selective depletion of apoA-II-containing HDL, as observed in carriers of LCAT deficiency, leads to an increased capacity of HDL to stimulate endothelial NO production, suggesting that changes in HDL apolipoprotein composition may be the target of therapeutic interventions designed to improve HDL functionality.

Data presented in this article are related to the research article entitled “A priori-defined Mediterranean-like dietary pattern predicts cardiovascular events better in north Europe than in Mediterranean countries” [Veglia et al., 2018]. Data contain information about the incidence of cardiovascular events in a high-risk European population (IMPROVE study) [Baldassarre et al., 2010, 2012, 2013]. Combined vascular events, as well as cardio- and cerebro-vascular events were stratified according to a priori-defined simple Mediterranean Diet (MD) score, based on just seven nutritional items (minimal adherence was 0 and maximal adherence was 7).

Cigarette smoking is a major independent risk factor for cardiovascular diseases (CVD). The underlying mechanisms, however, are not clearly understood. Lungs are the primary route of exposure to smoke, with pulmonary cells and surfactant being the first structures directly exposed, resulting in the leakage of the immature proteoform of surfactant protein B (proSP-B). Herein, we evaluated whether proSP-B joined the cargo of high-density lipoprotein (HDL) proteins in healthy young subjects (n = 106) without any CVD risk factor other than smoking, and if HDL-associated proSP-B (HDL-SPB) correlated with pulmonary function parameters, systemic inflammation, and oxidative stress. At univariable analysis, HDL-SPB resulted significantly higher in smokers (2.2-fold, p < 0.001) than in non-smokers. No significant differences have been detected between smokers and non-smokers for inflammation, oxidation variables, and alveolar-capillary diffusion markers. In a multivariable model, HDL-SPB was independently associated with smoking. In conclusion, HDL-SPB is not only a precocious and sensitive index of the acute effects of smoke, but it might be also a potential causal factor in the onset of the vascular damage induced by modified HDL. These findings contribute to the emerging concept that the quality of the HDL proteome, rather than the quantity of particles, plays a central role in CVD risk protection.

The impact of cholesteryl ester transfer protein (CETP) on atherosclerosis is highly debated. This study aimed to investigate the associations between plasma CETP or CETP genotypes and carotid intima-media thickness (cIMT) and the influence of high-density lipoprotein cholesterol (HDL-C) on these associations. Plasma CETP and HDL-C concentrations were measured in 552 subjects free of any pharmacological treatment from the IMPROVE cohort, which includes 3711 European subjects at high cardiovascular risk. CETP single-nucleotide polymorphisms (SNPs) and cIMT measures (cIMTmax; cIMTmean–max of bifurcations, common and internal carotids; plaque-free common carotid [PF CC]-IMTmean) were available for the full cohort. In drug-free subjects, plasma CETP correlated with HDL-C levels (r = 0.19, p < 0.0001), but not with cIMT variables. When stratified according to HDL-C quartiles, CETP positively correlated with cIMTmax and cIMTmean–max, but not with PF CC-IMTmean, in the top HDL-C quartile only. Positive associations between the CETP concentration and cIMTmax or cIMTmean–max were found in the top HDL-C quartile, whereas HDL-C levels were negatively correlated with cIMTmax and cIMTmean–max when the CETP concentration was below the median (HDL-C × CETP interaction, p = 0.001 and p = 0.003 for cIMTmax and cIMTmean–max, respectively). In the full cohort, three CETP SNPs (rs34760410, rs12920974, rs12708968) were positively associated with cIMTmax. rs12444708 exhibited a significant interaction with HDL-C levels in the prediction of cIMTmax. In conclusion, a significant interplay was found between plasma CETP and/or CETP genotype and HDL-C in the prediction of carotid plaque thickness, as indexed by cIMTmax. This suggests that the association of HDL-C with carotid atherosclerosis is CETP-dependent.

Background: The lack of specific vaccines or drugs against coronavirus disease 2019 (COVID-19) warrants studies focusing on alternative clinical approaches to reduce the spread of this pandemic disease. In this study, we investigated whether anti-influenza vaccination plays a role in minimizing the diffusion of COVID-19 in the Italian population aged 65 and over. Methods: Four COVID-19 outcomes were used: severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) seroprevalence, hospitalizations for COVID-19 symptoms, admissions to intensive care units for reasons related to SARS-CoV-2, and deaths attributable to COVID-19. Results: At univariate analyses, the influenza vaccination coverage rates correlated negatively with all COVID-19 outcomes (Beta ranging from −134 to −0.61; all p < 0.01). At multivariable analyses, influenza vaccination coverage rates correlated independently with SARS-CoV-2 seroprevalence (Beta (95% C.I.): −130 (−198, −62); p = 0.001), hospitalizations for COVID-19 symptoms (Beta (95% C.I.): −4.16 (−6.27, −2.05); p = 0.001), admission to intensive care units for reasons related to SARS-CoV-2 (Beta (95% C.I.): −0.58 (−1.05, −0.12); p = 0.017), and number of deaths attributable to COVID-19 (Beta (95% C.I.): −3.29 (−5.66, −0.93); p = 0.010). The R2 observed in the unadjusted analysis increased from 82% to 159% for all the considered outcomes after multivariable analyses. Conclusions: In the Italian population, the coverage rate of the influenza vaccination in people aged 65 and over is associated with a reduced spread and a less severe clinical expression of COVID-19. This finding warrants ad hoc studies to investigate the role of influenza vaccination in preventing the spread of COVID-19.

The senescence of vascular smooth muscle cells (VSMCs), characterized by the acquisition of senescence-associated secretory phenotype (SASP), is relevant for VSMCs osteoblastic differentiation and vascular calcification (VC). MicroRNA-34a (miR-34a) is a driver of such phenomena and could play a role in vascular inflammaging. Herein, we analyzed the relationship between miR-34a and the prototypical SASP component IL6 in in vitro and in vivo models. miR-34a and IL6 levels increased and positively correlated in aortas of 21 months-old male C57BL/6J mice and in human aortic smooth muscle cells (HASMCs) isolated from donors of different age and undergone senescence. Lentiviral overexpression of miR-34a in HASMCs enhanced IL6 secretion. HASMCs senescence and calcification accelerated after exposure to conditioned medium of miR-34a-overexpressing cells. Analysis of miR-34a-induced secretome revealed enhancement of several pro-inflammatory cytokines and chemokines, including IL6, pro-senescent growth factors and matrix-degrading molecules. Moreover, induction of aortas medial calcification and concomitant IL6 expression, with an overdose of vitamin D, was reduced in male C57BL/6J Mir34a−/− mice. Finally, a positive correlation was observed between circulating miR-34a and IL6 in healthy subjects of 20-90 years. Hence, the vascular age-associated miR-34a promotes VSMCs SASP activation and contributes to arterial inflammation and dysfunctions such as VC.

[This corrects the article DOI: 10.1371/journal.pone.0191172.].

AIMS:Carotid intima media thickness (CIMT) predicts cardiovascular (CVD) events, but the predictive value of CIMT change is debated. We assessed the relation between CIMT change and events in individuals at high cardiovascular risk. METHODS AND RESULTS:From 31 cohorts with two CIMT scans (total n = 89070) on average 3.6 years apart and clinical follow-up, subcohorts were drawn: (A) individuals with at least 3 cardiovascular risk factors without previous CVD events, (B) individuals with carotid plaques without previous CVD events, and (C) individuals with previous CVD events. Cox regression models were fit to estimate the hazard ratio (HR) of the combined endpoint (myocardial infarction, stroke or vascular death) per standard deviation (SD) of CIMT change, adjusted for CVD risk factors. These HRs were pooled across studies. In groups A, B and C we observed 3483, 2845 and 1165 endpoint events, respectively. Average common CIMT was 0.79mm (SD 0.16mm), and annual common CIMT change was 0.01mm (SD 0.07mm), both in group A. The pooled HR per SD of annual common CIMT change (0.02 to 0.43mm) was 0.99 (95% confidence interval: 0.95-1.02) in group A, 0.98 (0.93-1.04) in group B, and 0.95 (0.89-1.04) in group C. The HR per SD of common CIMT (average of the first and the second CIMT scan, 0.09 to 0.75mm) was 1.15 (1.07-1.23) in group A, 1.13 (1.05-1.22) in group B, and 1.12 (1.05-1.20) in group C. CONCLUSIONS:We confirm that common CIMT is associated with future CVD events in individuals at high risk. CIMT change does not relate to future event risk in high-risk individuals.

Recent advances in highly multiplexed immunoassays have allowed systematic large-scale measurement of hundreds of plasma proteins in large cohort studies. In combination with genotyping, such studies offer the prospect to 1) identify mechanisms involved with regulation of protein expression in plasma, and 2) determine whether the plasma proteins are likely to be causally implicated in disease. We report here the results of genome-wide association (GWA) studies of 83 proteins considered relevant to cardiovascular disease (CVD), measured in 3,394 individuals with multiple CVD risk factors. We identified 79 genome-wide significant (p

BackgroundAcute kidney injury (AKI) has been associated with increased mortality in ST‐segment elevation myocardial infarction. We compared the mortality predictive accuracy of the 3 AKI definitions used most widely for patients with ST‐segment elevation myocardial infarction undergoing primary percutaneous coronary intervention. Methods and ResultsWe included 3771 patients with ST‐segment elevation myocardial infarction treated with primary percutaneous coronary intervention at 2 Italian hospitals. AKI incidence was evaluated according to creatinine increases of ≥25% (AKI‐25), ≥0.3 mg/dL (AKI‐0.3), and ≥0.5 mg/dL (AKI‐0.5). The primary end point was in‐hospital mortality. Overall, 557 (15%), 522 (14%), and 270 (7%) patients developed AKI‐25, AKI‐0.3, and AKI‐0.5, respectively (P

Lecithin:cholesterol acyltransferase (LCAT) is the enzyme responsible for cholesterol esterification in plasma. LCAT is a major factor in HDL remodeling and metabolism, and it has long been believed to play a critical role in macrophage reverse cholesterol transport (RCT). The effect of LCAT on human atherogenesis is still controversial. In the present study, the plasma LCAT concentration was measured in all subjects (n = 540) not on drug treatment at the time of enrollment in the multicenter, longitudinal, observational IMPROVE study. Mean and maximum intima-media thickness (IMT) of the whole carotid tree was measured by B-mode ultrasonography in all subjects. In the entire cohort, LCAT quartiles were not associated with carotid mean and maximum IMT (P for trend 0.95 and 0.18, respectively), also after adjustment for age, gender, HDL-cholesterol (HDL-C), and triglycerides. No association between carotid IMT and LCAT quartiles was observed in men (P=0.30 and P=0.99 for mean and maximum IMT, respectively), whereas carotid IMT increased with LCAT quartiles in women (P for trend 0.14 and 0.019 for mean and maximum IMT, respectively). The present findings support the concept that LCAT is not required for an efficient reverse cholesterol transport and that a low plasma LCAT concentration and activity is not associated with increased atherosclerosis.

Brain-Derived Neurotrophic Factor (BDNF) Val66Met polymorphism has been associated with increased susceptibility to develop mood disorders and recently it has been also linked with cardiovascular disease (CVD). Interestingly, stressful conditions unveil the anxious/depressive-like behavioral phenotype in heterozygous BDNFVal66Met (BDNFVal/Met) mice, suggesting an important relationship in terms of gene-environment interaction (GxE). However, the interplay between stress and BDNFVal/Met in relation to CVD is completely unknown. Here, we showed that BDNFVal/Met mice display a greater propensity to arterial thrombosis than wild type BDNFVal/Val mice after 7 days of restraint stress (RS). RS markedly increased the number of leukocytes and platelets, and induced hyper-responsive platelets as showed by increased circulating platelet/leukocyte aggregates and enhanced expression of P-selectin and GPIIbIIIa in heterozygous mutant mice. In addition, stressed BDNFVal/Met mice had a greater number of large and reticulated platelets but comparable number and maturation profile of bone marrow megakaryocytes compared to BDNFVal/Val mice. Interestingly, RS led to a significant reduction of BDNF expression accompanied by an increased activity of tissue factor in the aorta of both BDNFVal/Val and BDNFVal/Met mice. In conclusion, we provide evidence that sub-chronic stress unveils prothrombotic phenotype in heterozygous BDNF Val66Met mice affecting both the number and functionality of blood circulating cells, and the expression of key thrombotic molecules in aorta. Human studies will be crucial to understand whether this GxE interaction need to be taken into account in risk stratification of coronary artery disease (CAD) patients.

Variants at the interleukin 6 receptor (IL6R) gene regulate inflammation and are associated with risk of coronary heart disease (CHD). The aim of the present study was to investigate the effects of IL6R haplotypes on circulating levels of inflammatory biomarkers and risk of CHD. We performed a discovery analysis in SHEEP, a myocardial infarction (MI) case control study (n = 2,774) and replicated our results in two large, independent European populations, PROCARDIS, a CHD case control study (n = 7,998), and IMPROVE (n = 3,711) a prospective cardiovascular cohort study. Two major haplotype blocks (rs12083537A/G and rs4075015A/T--block 1; and rs8192282G/A, rs4553185T/C, rs8192284A/C, rs4240872T/C and rs7514452T/C--block 2) were identified in the IL6R gene. IL6R haplotype associations with C-reactive protein (CRP), fibrinogen, IL6, soluble IL6R (sIL6R), IL6, IL8 and TNF-α in SHEEP, CRP and fibrinogen in PROCARDIS and CRP in IMPROVE as well as association with risk of MI and CHD, were analyzed by THESIAS. Haplotypes in block 1 were associated neither with circulating inflammatory biomarkers nor with the MI/CHD risk. Haplotypes in block 2 were associated with circulating levels of CRP, in all three study populations, with fibrinogen in SHEEP and PROCARDIS, with IL8 and sIL6Rin SHEEP and with a modest, non significant, increase (7%) in MI/CHD risk in the three populations studied. Our results indicate that IL6R haplotypes regulate the circulating levels of inflammatory biomarkers. Lack of association with the risk of CHD may be explained by the combined effect of SNPs with opposite effect on the CHD risk, the sample size as well as by structural changes affecting sIL6R stability in the circulation.

Mutations in the CETP gene resulting in defective CETP activity have been shown to cause remarkable elevations of plasma HDL-C levels, with the accumulation in plasma of large, buoyant HDL particles enriched in apolipoprotein E. Genetic CETP deficiency thus represents a unique tool to evaluate how structural alterations of HDL impact on HDL atheroprotective functions. Aim of the present study was to assess the ability of HDL obtained from CETP-deficient subjects to protect endothelial cells from the development of endothelial dysfunction. HDL isolated from one homozygous and seven heterozygous carriers of CETP null mutations were evaluated for their ability to down-regulate cytokine-induced cell adhesion molecule expression and to promote NO production in cultured endothelial cells. When compared at the same protein concentration, HDL and HDL3 from carriers proved to be as effective as control HDL and HDL3 in down-regulating cytokine-induced VCAM-1, while carrier HDL2 were more effective than control HDL2 in inhibiting VCAM-1 expression. On the other hand, HDL and HDL fractions from carriers of CETP deficiency were significantly less effective than control HDL and HDL fractions in stimulating NO production, due to a reduced eNOS activating capacity, likely because of a reduced S1P content. In conclusion, the present findings support the notion that genetic CETP deficiency, by affecting HDL particle structure, impacts on HDL vasculoprotective functions. Understanding of these effects might be important for predicting the outcomes of pharmacological CETP inhibition.

Chronic obstructive pulmonary disease (COPD) independently associates with an increased risk of coronary artery disease (CAD), but it has not been fully investigated whether this co-morbidity involves shared pathophysiological mechanisms. To identify potential common pathways across the two diseases, we tested all recently published single nucleotide polymorphisms (SNPs) associated with human lung function (spirometry) for association with carotid intima-media thickness (cIMT) in 3,378 subjects with multiple CAD risk factors, and for association with CAD in a case-control study of 5,775 CAD cases and 7,265 controls. SNPs rs2865531, located in the CFDP1 gene, and rs9978142, located in the KCNE2 gene, were significantly associated with CAD. In addition, SNP rs9978142 and SNP rs3995090 located in the HTR4 gene, were associated with average and maximal cIMT measures. Genetic risk scores combining the most robustly spirometry-associated SNPs from the literature were modestly associated with CAD, (odds ratio (OR) (95% confidence interval (CI95) = 1.06 (1.03, 1.09); P-value = 1.5 × 10(-4), per allele). In conclusion, our study suggests that some genetic loci implicated in determining human lung function also influence cIMT and susceptibility to CAD. The present results should help elucidate the molecular underpinnings of the co-morbidity observed across COPD and CAD.

All the enzymatic factors/cofactors involved in nitric oxide (NO) metabolism have been recently found in red blood cells. Increased oxidative stress impairs NO bioavailability and has been described in plasma of coronary artery disease (CAD) patients. The aim of the study was to highlight a potential dysfunction of the metabolic profile of NO in red blood cells and in plasma from CAD patients compared with healthy controls.We determined L-arginine/NO pathway by liquid-chromatography tandem mass spectrometry and high performance liquid chromatography methods. The ratio of oxidized and reduced forms of glutathione, as index of oxidative stress, was measured by liquid-chromatography tandem mass spectrometry method. NO synthase expression and activity were evaluated by immunofluorescence staining and ex-vivo experiments of L-[(15)N2]arginine conversion to L-[(15)N]citrulline respectively.Increased amounts of asymmetric and symmetric dimethylarginines were found both in red blood cells and in plasma of CAD patients in respect to controls. Interestingly NO synthase expression and activity were reduced in CAD red blood cells. In contrast, oxidized/reduced glutathione ratio was increased in CAD and was associated to arginase activity.Our study analyzed for the first time the whole metabolic pathway of L-arginine/NO, both in red blood cells and in plasma, highlighting an impairment of NO pathway in erythrocytes from CAD patients, associated with decreased NO synthase expression/activity and increased oxidative stress.

BACKGROUND: Mechanisms linking chronic kidney disease (CKD) and adverse outcomes in acute coronary syndromes (ACS) are not fully understood. Among potential key players, reduced nitric oxide (NO) synthesis due to its endogenous inhibitors, asymmetric (ADMA) and symmetric (SDMA) dimethylarginine could be involved. We measured plasma concentration of arginine, ADMA and SDMA and investigated their relationship with CKD and long-term outcome in non-ST-elevation myocardial infarction (NSTEMI). METHODOLOGY/PRINCIPAL FINDINGS: We prospectively measured arginine, ADMA, and SDMA at hospital admission in 104 NSTEMI patients. CKD was defined as an estimated glomerular filtration rate (eGFR)