Radical Oxygen Species, Oxidized Low-Density Lipoproteins, and Lectin-like Oxidized Low-Density Lipoprotein Receptor 1: A Vicious Circle in Atherosclerotic Process.

Atherosclerosis is a complex condition that involves the accumulation of lipids and subsequent plaque formation in the arterial intima. There are various stimuli, cellular receptors, and pathways involved in this process, but oxidative modifications of low-density lipoprotein (ox-LDL) are particularly important in the onset and progression of atherosclerosis. Ox-LDLs promote foam-cell formation, activate proinflammatory pathways, and induce smooth-muscle-cell migration, apoptosis, and cell death. One of the major receptors for ox-LDL is LOX-1, which is upregulated in several cardiovascular diseases, including atherosclerosis. LOX-1 activation in endothelial cells promotes endothelial dysfunction and induces pro-atherogenic signaling, leading to plaque formation. The binding of ox-LDLs to LOX-1 increases the generation of reactive oxygen species (ROS), which can induce LOX-1 expression and oxidize LDLs, contributing to ox-LDL generation and further upregulating LOX-1 expression. This creates a vicious circle that is amplified in pathological conditions characterized by high plasma levels of LDLs. Although LOX-1 has harmful effects, the clinical significance of inhibiting this protein remains unclear. Further studies both in vitro and in vivo are needed to determine whether LOX-1 inhibition could be a potential therapeutic target to counteract the atherosclerotic process.

N-Acetylcysteine, N-Acetylcysteine Amide, and Thioredoxin Mimetic Peptides Regenerate Mercaptoalbumin and Exhibit Antioxidant Activity.

Albumin (HSA) is the most abundant circulating protein and plays a pivotal role in maintaining the redox state of the plasma. Three HSA proteoforms have been identified based on the redox state of cysteine 34. These proteoforms comprise of the reduced state (HSA-SH) referred to as mercaptoalbumin, non-mercaptoalbumin-1, containing a disulfide with small thiols such as cysteine (HSA-Cys), and non-mercaptoalbumin-2, representing the higher oxidized proteoform. Several clinical studies have shown a relationship between an individual's serum HSA redox status and the severity of diseases such as heart failure, diabetes mellitus, and liver disease. Furthermore, when HSA undergoes oxidation, it can worsen certain health conditions and contribute to their advancement. This study aimed to evaluate the ability of the redox compounds AD4/NACA and the thioredoxin mimetic (TXM) peptides TXM-CB3, TXM-CB13, and TXM-CB30 to regenerate HSA-SH and to enhance its redox activity. The HSA proteoforms were quantified by LC-MS, and the antioxidant activity was determined using dichlorofluorescin. Each of the compounds exhibited a significant increase in HSA-SH and a reduction in HSA-Cys levels. The increase in HSA-SH was associated with a recovery of its antioxidant activity. In this work, we unveil a novel mechanistic facet of the antioxidant activity of AD4/NACA and TXM peptides. These results suggest an additional therapeutic approach for addressing oxidative stress-related conditions.

Symptomatic post COVID patients have impaired alveolar capillary membrane function and high VE/VCO2.

BACKGROUND: Post COVID-19 syndrome is characterized by several cardiorespiratory symptoms but the origin of patients' reported symptomatology is still unclear. METHODS: Consecutive post COVID-19 patients were included. Patients underwent full clinical evaluation, symptoms dedicated questionnaires, blood tests, echocardiography, thoracic computer tomography (CT), spirometry including alveolar capillary membrane diffusion (DM) and capillary volume (Vcap) assessment by combined carbon dioxide and nitric oxide lung diffusion (DLCO/DLNO) and cardiopulmonary exercise test. We measured surfactant derive protein B (immature form) as blood marker of alveolar cell function. RESULTS: We evaluated 204 consecutive post COVID-19 patients (56.5 ± 14.5 years, 89 females) 171 ± 85 days after the end of acute COVID-19 infection. We measured: forced expiratory volume (FEV1) 99 ± 17%pred, FVC 99 ± 17%pred, DLCO 82 ± 19%, DM 47.6 ± 14.8 mL/min/mmHg, Vcap 59 ± 17 mL, residual parenchymal damage at CT 7.2 ± 3.2% of lung tissue, peakVO2 84 ± 18%pred, VE/VCO2 slope 112 [102-123]%pred. Major reported symptoms were: dyspnea 45% of cases, tiredness 60% and fatigability 77%. Low FEV1, Vcap and high VE/VCO2 slope were associated with persistence of dyspnea. Tiredness was associated with high VE/VCO2 slope and low PeakVO2 and FEV1 while fatigability with high VE/VCO2 slope. SPB was fivefold higher in post COVID-19 than in normal subjects, but not associated to any of the referred symptoms. SPB was negatively associated to Vcap. CONCLUSIONS: In patients with post COVID-19, cardiorespiratory symptoms are linked to VE/VCO2 slope. In these patients the alveolar cells are dysregulated as shown by the very high SPB. The Vcap is low likely due to post COVID-19 pulmonary endothelial/vasculature damage but DLCO is only minimally impaired being DM preserved.

Multiplexed MRM-based proteomics for identification of circulating proteins as biomarkers of cardiovascular damage progression associated with diabetes mellitus.

BACKGROUND: Type 2 diabetes mellitus (T2DM) increases the risk of coronary heart disease (CHD) by 2-4 fold, and is associated with endothelial dysfunction, dyslipidaemia, insulin resistance, and chronic hyperglycaemia. The aim of this investigation was to assess, by a multimarker mass spectrometry approach, the predictive role of circulating proteins as biomarkers of cardiovascular damage progression associated with diabetes mellitus. METHODS: The study considered 34 patients with both T2DM and CHD, 31 patients with T2DM and without CHD, and 30 patients without diabetes with a diagnosis of CHD. Plasma samples of subjects were analysed through a multiplexed targeted liquid chromatography mass spectrometry (LC-MS)-based assay, namely Multiple Reaction Monitoring (MRM), allowing the simultaneous detection of peptides derived from a protein of interest. Gene Ontology (GO) Analysis was employed to identify enriched GO terms in the biological process, molecular function, or cellular component categories. Non-parametric multivariate methods were used to classify samples from patients and evaluate the relevance of the analysed proteins' panel. RESULTS: A total of 81 proteins were successfully quantified in the human plasma samples. Gene Ontology analysis assessed terms related to blood microparticles, extracellular exosomes and collagen-containing extracellular matrix. Preliminary evaluation using analysis of variance (ANOVA) of the differences in the proteomic profile among patient groups identified 13 out of the 81 proteins as significantly different. Multivariate analysis, including cluster analysis and principal component analysis, identified relevant grouping of the 13 proteins. The first main cluster comprises apolipoprotein C-III, apolipoprotein C-II, apolipoprotein A-IV, retinol-binding protein 4, lysozyme C and cystatin-C; the second one includes, albeit with sub-grouping, alpha 2 macroglobulin, afamin, kininogen 1, vitronectin, vitamin K-dependent protein S, complement factor B and mannan-binding lectin serine protease 2. Receiver operating characteristic (ROC) curves obtained with the 13 selected proteins using a nominal logistic regression indicated a significant overall distinction (p < 0.001) among the three groups of subjects, with area under the ROC curve (AUC) ranging 0.91-0.97, and sensitivity and specificity ranging from 85 to 100%. CONCLUSIONS: Targeted mass spectrometry approach indicated 13 multiple circulating proteins as possible biomarkers of cardiovascular damage progression associated with T2DM, with excellent classification results in terms of sensitivity and specificity.

Albumin Thiolation and Oxidative Stress Status in Patients with Aortic Valve Stenosis.

Recent evidence indicates that reactive oxygen species play an important causative role in the onset and progression of valvular diseases. Here, we analyzed the oxidative modifications of albumin (HSA) occurring on Cysteine 34 and the antioxidant capacity of the serum in 44 patients with severe aortic stenosis (36 patients underwent aortic valve replacement and 8 underwent a second aortic valve substitution due to a degenerated bioprosthetic valve), and in 10 healthy donors (controls). Before surgical intervention, patients showed an increase in the oxidized form of albumin (HSA-Cys), a decrease in the native reduced form (HSA-SH), and a significant reduction in serum free sulfhydryl groups and in the total serum antioxidant activity. Patients undergoing a second valve replacement showed levels of HSA-Cys, free sulfhydryl groups, and total antioxidant activity similar to those of controls. In vitro incubation of whole blood with aspirin (ASA) significantly increased the free sulfhydryl groups, suggesting that the in vivo treatment with ASA may contribute to reducing oxidative stress. We also found that N-acetylcysteine and its amide derivative were able to regenerate HSA-SH. In conclusion, the systemic oxidative stress reflected by high levels of HSA-Cys is increased in patients with aortic valve stenosis. Thiol-disulfide breaking agents regenerate HSA-SH, thus paving the way to the use these compounds to mitigate the oxidative stress occurring in the disease.

Proteome profiling identifies circulating biomarkers associated with hepatic steatosis in subjects with Prader-Willi syndrome.

Introduction: Prader-Willi syndrome (PWS) is a rare genetic disorder characterized by loss of expression of paternal chromosome 15q11.2-q13 genes. Individuals with PWS exhibit unique physical, endocrine, and metabolic traits associated with severe obesity. Identifying liver steatosis in PWS is challenging, despite its lower prevalence compared to non-syndromic obesity. Reliable biomarkers are crucial for the early detection and management of this condition associated with the complex metabolic profile and cardiovascular risks in PWS. Methods: Circulating proteome profiling was conducted in 29 individuals with PWS (15 with steatosis, 14 without) using the Olink Target 96 metabolism and cardiometabolic panels. Correlation analysis was performed to identify the association between protein biomarkes and clinical variables, while the gene enrichment analysis was conducted to identify pathways linked to deregulated proteins. Receiver operating characteristic (ROC) curves assessed the discriminatory power of circulating protein while a logistic regression model evaluated the potential of a combination of protein biomarkers. Results: CDH2, CTSO, QDPR, CANT1, ALDH1A1, TYMP, ADGRE, KYAT1, MCFD, SEMA3F, THOP1, TXND5, SSC4D, FBP1, and CES1 exhibited a significant differential expression in liver steatosis, with a progressive increase from grade 1 to grade 3. FBP1, CES1, and QDPR showed predominant liver expression. The logistic regression model, -34.19 + 0.85 * QDPR*QDPR + 0.75 * CANT1*TYMP - 0.46 * THOP1*ALDH1A, achieved an AUC of 0.93 (95% CI: 0.63-0.99), with a sensitivity of 93% and specificity of 80% for detecting steatosis in individuals with PWS. These biomarkers showed strong correlations among themselves and were involved in an interconnected network of 62 nodes, related to seven metabolic pathways. They were also significantly associated with cholesterol, LDL, triglycerides, transaminases, HbA1c, FLI, APRI, and HOMA, and showed a negative correlation with HDL levels. Conclusion: The biomarkers identified in this study offer the potential for improved patient stratification and personalized therapeutic protocols.

Circulating Small Extracellular Vesicles Reflect the Severity of Myocardial Damage in STEMI Patients.

Circulating small extracellular vesicles (sEVs) contribute to inflammation, coagulation and vascular injury, and have great potential as diagnostic markers of disease. The ability of sEVs to reflect myocardial damage assessed by Cardiac Magnetic Resonance (CMR) in ST-segment elevation myocardial infarction (STEMI) is unknown. To fill this gap, plasma sEVs were isolated from 42 STEMI patients treated by primary percutaneous coronary intervention (pPCI) and evaluated by CMR between days 3 and 6. Nanoparticle tracking analysis showed that sEVs were greater in patients with anterior STEMI (p = 0.0001), with the culprit lesion located in LAD (p = 0.045), and in those who underwent late revascularization (p = 0.038). A smaller sEV size was observed in patients with a low myocardial salvage index (MSI, p = 0.014). Patients with microvascular obstruction (MVO) had smaller sEVs (p < 0.002) and lower expression of the platelet marker CD41-CD61 (p = 0.039). sEV size and CD41-CD61 expression were independent predictors of MVO/MSI (OR [95% CI]: 0.93 [0.87-0.98] and 0.04 [0-0.61], respectively). In conclusion, we provide evidence that the CD41-CD61 expression in sEVs reflects the CMR-assessed ischemic damage after STEMI. This finding paves the way for the development of a new strategy for the timely identification of high-risk patients and their treatment optimization.

The alveolar-capillary unit in the physiopathological conditions of heart failure: identification of a potential marker.

In this review, we describe the structure and function of the alveolar-capillary membrane and the identification of a novel potential marker of its integrity in the context of heart failure (HF). The alveolar-capillary membrane is indeed a crucial structure for the maintenance of the lung parenchyma gas exchange capacity, and the occurrence of pathological conditions determining lung fluids accumulation, such as HF, might significantly impair lung diffusion capacity altering the alveolar-capillary membrane protective functions. In the years, we found that the presence of immature forms of the surfactant protein-type B (proSP-B) in the circulation reflects alterations in the alveolar-capillary membrane integrity. We discussed our main achievements showing that proSP-B, due to its chemical properties, specifically binds to high-density lipoprotein, impairing their antioxidant activity, and likely contributing to the progression of the disease. Further, we found that immature proSP-B, not the mature protein, is related to lung abnormalities, more precisely than the lung function parameters. Thus, to the list of the potential proposed markers of HF, we add proSP-B, which represents a precise marker of alveolar-capillary membrane dysfunction in HF, correlates with prognosis, and represents a precocious marker of drug therapy.

Looking into the Kinetics of NT-proBNP and sST2 Changes in Patients with Heart Failure Treated with Sacubitril/Valsartan: A Hint to Different Therapeutic Pathways.

BACKGROUND AND OBJECTIVE: N-terminal pro-B-type natriuretic peptide (NT-proBNP) and soluble interleukin 1 receptor-like 1 ST2 (sST2) are biomarkers used to grade heart failure with reduced ejection fraction (HFrEF) severity. Both are potential targets of HFrEF treatment, but the first is associated with the patient's hemodynamic status, while the second is more indicative of the inflammatory status and of myocardial fibrosis. The aim of this study was to assess the kinetics of these biomarkers after treatment with sacubitril/valsartan in HFrEF. METHODS: We analyzed blood samples of patients with HFrEF at baseline (before sacubitril/valsartan treatment), after 1, 2, and 3 months (respectively, after a month taking the 24/26 - 49/51 - 97/103 mg twice daily, or b.i.d., doses), and 6 months after the maximum-tolerated dose was reached (end study). RESULTS: We obtained samples from 72 patients with HFrEF (age 64.0 ± 10.5 years, 83% males). NT-proBNP and sST2 values progressively and significantly reduced to 37% and 16%, respectively, with a greater reduction for NT-proBNP (p < 0.001). Specifically, NT-proBNP reduced from 1144 [593-2586] pg/mL to 743 [358-1524] pg/mL and sST2 from 27.3 [20.5-35.0] ng/mL to 23.1 [15.9-30.7] ng/mL, p for trend < 0.001 in both cases. The reduction of the two biomarkers over time occurred with statistically significant different kinetics: deferred for sST2 and faster for NT-proBNP. No significant changes in renal function and potassium levels were recorded. CONCLUSION: These findings suggest that, in patients with HF, sacubitril/valsartan effects on the cardiovascular system share a double pathway: a first, hemodynamic, faster pathway and a second, non-hemodynamic anti-fibrotic, delayed one. Both likely contribute to the sacubitril/valsartan benefits in HFrEF.

N-acetylcysteine Amide AD4/NACA and Thioredoxin Mimetic Peptides Inhibit Platelet Aggregation and Protect against Oxidative Stress.

In the present study, we tested the effect of small-molecular-weight redox molecules on collagen-induced platelet aggregation. We used N-acetylcysteine amide (AD4/NACA), the amide form of N-acetylcysteine (NAC), a thiol antioxidant with improved lipophilicity and bioavailability compared to NAC, and the thioredoxin-mimetic (TXM) peptides, TXM-CB3, TXM-CB13, and TXM-CB30. All compounds significantly inhibited platelet aggregation induced by collagen, with TXM-peptides and AD4 being more effective than NAC. The levels of TxB2 and 12-HETE, the main metabolites derived from the cyclooxygenase and lipoxygenase pathways following platelet activation, were significantly reduced in the presence of AD4, TXM peptides, or NAC, when tested at the highest concentration (0.6 mM). The effects of AD4, TXM-peptides, and NAC were also tested on the clotting time (CT) of whole blood. TXM-CB3 and TXM-CB30 showed the greatest increase in CT. Furthermore, two representative compounds, TXM-CB3 and NAC, showed an increase in the anti-oxidant free sulfhydryl groups of plasma detected via Ellman's method, suggesting a contribution of plasma factors to the antiaggregating effects. Our results suggest that these small-molecular-weight redox peptides might become useful for the prevention and/or treatment of oxidative stress conditions associated with platelet activation.

Proteome Profiling of the Dura Mater in Patients with Moyamoya Angiopathy.

Moyamoya angiopathy (MMA) is an uncommon cerebrovascular disease characterized by a progressive steno-occlusive lesion of the internal carotid artery and the compensatory development of an unstable network of collateral vessels. These vascular hallmarks are responsible for recurrent ischemic/hemorrhagic strokes. Surgical treatment represents the preferred procedure for MMA patients, and indirect revascularization may induce a spontaneous angiogenesis between the brain surface and dura mater (DM), whose function remains rather unknown. A better understanding of MMA pathogenesis is expected from the molecular characterization of DM. We performed a comprehensive, label-free, quantitative mass spectrometry-based proteomic characterization of DM. The 30 most abundant identified proteins were located in the extracellular region or exosomes and were involved in extracellular matrix organization. Gene ontology analysis revealed that most proteins were involved in binding functions and hydrolase activity. Among the 30 most abundant proteins, Filamin A is particularly relevant because considering its well-known biochemical functions and molecular features, it could be a possible second hit gene with a potential role in MMA pathogenesis. The current explorative study could pave the way for further analyses aimed at better understanding such uncommon and disabling intracranial vasculopathy.

Proteomics and Lipidomics to unveil the contribution of PCSK9 beyond cholesterol lowering: a narrative review.

Proprotein convertase subtilisin/kexin type 9 (PCSK9), one of the key regulators of the low-density lipoprotein receptor (LDLR), can play a direct role in atheroma development. Although advances in the understandings of genetic PCSK9 polymorphisms have enabled to reveal the role of PCSK9 in the complex pathophysiology of cardiovascular diseases (CVDs), increasing lines of evidence support non-cholesterol-related processes mediated by PCSK9. Owing to major improvements in mass spectrometry-based technologies, multimarker proteomic and lipidomic panels hold the promise to identify novel lipids and proteins potentially related to PCSK9. Within this context, this narrative review aims to provide an overview of the most significant proteomics and lipidomics studies related to PCSK9 effects beyond cholesterol lowering. These approaches have enabled to unveil non-common targets of PCSK9, potentially leading to the development of novel statistical models for CVD risk prediction. Finally, in the era of precision medicine, we have reported the impact of PCSK9 on extracellular vesicles (EVs) composition, an effect that could contribute to an increased prothrombotic status in CVD patients. The possibility to modulate EVs release and cargo could help counteract the development and progression of the atherosclerotic process.

Impact of Sacubitril/Valsartan on Circulating microRNA in Patients with Heart Failure.

Sacubitril/Valsartan, used for the treatment of heart failure (HF), is a combination of two drugs, an angiotensin receptor inhibitor, and a neprilysin inhibitor, which activates vasoactive peptides. Even though its beneficial effects on cardiac functions have been demonstrated, the mechanisms underpinning these effects remain poorly understood. To achieve more mechanistic insights, we analyzed the profiles of circulating miRNAs in plasma from patients with stable HF with reduced ejection function (HFrEF) and treated with Sacubitril/Valsartan for six months. miRNAs are short (22-24 nt) non-coding RNAs, which are not only emerging as sensitive and stable biomarkers for various diseases but also participate in the regulation of several biological processes. We found that in patients with high levels of miRNAs, specifically miR-29b-3p, miR-221-3p, and miR-503-5p, Sacubitril/Valsartan significantly reduced their levels at follow-up. We also found a significant negative correlation of miR-29b-3p, miR-221-3p, and miR-503-5p with VO2 at peak exercise, whose levels decrease with HF severity. Furthermore, from a functional point of view, miR-29b-3p, miR-221-3p, and miR-503-5p all target Phosphoinositide-3-Kinase Regulatory Subunit 1, which encodes regulatory subunit 1 of phosphoinositide-3-kinase. Our findings support that an additional mechanism through which Sacubitril/Valsartan exerts its functions is the modulation of miRNAs with potentially relevant roles in HFrEF pathophysiology.

Prenylcysteine Oxidase 1 Is a Key Regulator of Adipogenesis.

The process of adipogenesis involves the differentiation of preadipocytes into mature adipocytes. Excessive adipogenesis promotes obesity, a condition that increasingly threatens global health and contributes to the rapid rise of obesity-related diseases. We have recently shown that prenylcysteine oxidase 1 (PCYOX1) is a regulator of atherosclerosis-disease mechanisms, which acts through mechanisms not exclusively related to its pro-oxidant activity. To address the role of PCYOX1 in the adipogenic process, we extended our previous observations confirming that Pcyox1-/-/Apoe-/- mice fed a high-fat diet for 8 or 12 weeks showed significantly lower body weight, when compared to Pcyox1+/+/Apoe-/- mice, due to an evident reduction in visceral adipose content. We herein assessed the role of PCYOX1 in adipogenesis. Here, we found that PCYOX1 is expressed in adipose tissue, and, independently from its pro-oxidant enzymatic activity, is critical for adipogenesis. Pcyox1 gene silencing completely prevented the differentiation of 3T3-L1 preadipocytes, by acting as an upstream regulator of several key players, such as FABP4, PPARγ, C/EBPα. Proteomic analysis, performed by quantitative label-free mass spectrometry, further strengthened the role of PCYOX1 in adipogenesis by expanding the list of its downstream targets. Finally, the absence of Pcyox1 reduces the inflammatory markers in adipose tissue. These findings render PCYOX1 a novel adipogenic factor with possible pathophysiological or therapeutic potential.

Pharmacometabolomics for the Study of Lipid-Lowering Therapies: Opportunities and Challenges.

Lipid-lowering therapies are widely used to prevent the development of atherosclerotic cardiovascular disease (ASCVD) and related mortality worldwide. "Omics" technologies have been successfully applied in recent decades to investigate the mechanisms of action of these drugs, their pleiotropic effects, and their side effects, aiming to identify novel targets for future personalized medicine with an improvement of the efficacy and safety associated with the treatment. Pharmacometabolomics is a branch of metabolomics that is focused on the study of drug effects on metabolic pathways that are implicated in the variation of response to the treatment considering also the influences from a specific disease, environment, and concomitant pharmacological therapies. In this review, we summarized the most significant metabolomic studies on the effects of lipid-lowering therapies, including the most commonly used statins and fibrates to novel drugs or nutraceutical approaches. The integration of pharmacometabolomics data with the information obtained from the other "omics" approaches could help in the comprehension of the biological mechanisms underlying the use of lipid-lowering drugs in view of defining a precision medicine to improve the efficacy and reduce the side effects associated with the treatment.

Macrophage Phenotyping in Atherosclerosis by Proteomics.

Macrophages are heterogeneous and plastic cells, able to adapt their phenotype and functions to changes in the microenvironment. They are involved in several homeostatic processes and also in many human diseases, including atherosclerosis, where they participate in all the stages of the disease. For these reasons, macrophages have been studied extensively using different approaches, including proteomics. Proteomics, indeed, may be a powerful tool to better understand the behavior of these cells, and a careful analysis of the proteome of different macrophage phenotypes can help to better characterize the role of these phenotypes in atherosclerosis and provide a broad view of proteins that might potentially affect the course of the disease. In this review, we discuss the different proteomic techniques that have been used to delineate the proteomic profile of macrophage phenotypes and summarize some results that can help to elucidate the roles of macrophages and develop new strategies to counteract the progression of atherosclerosis and/or promote regression.

Novel insights about albumin in cardiovascular diseases: Focus on heart failure.

The Human Plasma Proteome has always been the most investigated compartment in proteomics-based biomarker discovery, and is considered the largest and deepest version of the human proteome, reflecting the state of the body in health and disease. Even if efforts have been always dedicated to the refinement of proteomic approaches to investigate more deeply the plasma proteome, it should not be forgotten that also highly abundant plasma proteins, like human serum albumin (HSA), often neglected in these studies, might provide fundamental physiological functions in plasma, and should be better considered. This review summarizes the important roles of HSA in the context of cardiovascular diseases (CVD), and in particular in heart failure. Notwithstanding much attention has been historically directed toward the association of HSA levels and CVD risk, the advances in the field of mass spectrometry research allow also a better characterization of the effects of oxidative modifications that could alter not only the structure but also the function of HSA.

Proteomic studies on apoB-containing lipoprotein in cardiovascular research: A comprehensive review.

The complexity of cardiovascular diseases (CVDs), which remains the leading cause of death worldwide, makes the current clinical pathway for cardiovascular risk assessment unsatisfactory, as there remains a substantial unexplained residual risk. Simultaneous assessment of a large number of plasma proteins may be a promising tool to further refine risk assessment, and lipoprotein-associated proteins have the potential to fill this gap. Technical advances now allow for high-throughput proteomic analysis in a reproducible and cost-effective manner. Proteomics has great potential to identify and quantify hundreds of candidate marker proteins in a sample and allows the translation from isolated lipoproteins to whole plasma, thus providing an individual multiplexed proteomic fingerprint. This narrative review describes the pathophysiological roles of atherogenic apoB-containing lipoproteins and the recent advances in their mass spectrometry-based proteomic characterization and quantitation for better refinement of CVD risk assessment.

PCSK9 Confers Inflammatory Properties to Extracellular Vesicles Released by Vascular Smooth Muscle Cells.

Vascular smooth muscle cells (VSMCs) are key participants in both early- and late-stage atherosclerosis and influence neighbouring cells possibly by means of bioactive molecules, some of which are packed into extracellular vesicles (EVs). Proprotein convertase subtilisin/kexin type 9 (PCSK9) is expressed and secreted by VSMCs. This study aimed to unravel the role of PCSK9 on VSMCs-derived EVs in terms of content and functionality. EVs were isolated from human VSMCs overexpressing human PCSK9 (VSMCPCSK9-EVs) and tested on endothelial cells, monocytes, macrophages and in a model of zebrafish embryos. Compared to EVs released from wild-type VSMCs, VSMCPCSK9-EVs caused a rise in the expression of adhesion molecules in endothelial cells and of pro-inflammatory cytokines in monocytes. These acquired an increased migratory capacity, a reduced oxidative phosphorylation and secreted proteins involved in immune response and immune effector processes. Concerning macrophages, VSMCPCSK9-EVs enhanced inflammatory milieu and uptake of oxidized low-density lipoproteins, whereas the migratory capacity was reduced. When injected into zebrafish embryos, VSMCPCSK9-EVs favoured the recruitment of macrophages toward the site of injection. The results of the present study provide evidence that PCSK9 plays an inflammatory role by means of EVs, at least by those derived from smooth muscle cells of vascular origin.

The Effects of Silencing PTX3 on the Proteome of Human Endothelial Cells.

The human long pentraxin PTX3 has complex regulatory roles at the crossroad of innate immunity, inflammation, and tissue repair. PTX3 can be produced by various cell types, including vascular endothelial cells (ECs), in response to pro-inflammatory cytokines or bacterial molecules. PTX3 has also been involved in the regulation of cardiovascular biology, even if ambiguous results have been so far provided in both preclinical and clinical research. In this study, we compared the proteomic profiles of human ECs (human umbilical vein ECs, HUVECs), focusing on differentially expressed proteins between the control and PTX3-silenced ECs. We identified 19 proteins that were more abundant in the proteome of control ECs and 23 proteins that were more expressed in PTX3-silenced cells. Among the latter, proteins with multifunctional roles in angiogenesis, oxidative stress, and inflammation were found, and were further validated by assessing their mRNAs with RT-qPCR. Nevertheless, the knock down of PTX3 did not affect in vitro angiogenesis. On the contrary, the lack of the protein induced an increase in pro-inflammatory markers and a shift to the more oxidative profile of PTX3-deficient ECs. Altogether, our results support the idea of a protective function for PTX3 in the control of endothelial homeostasis, and more generally, in cardiovascular biology.