Matrix metalloproteinase 2 (MMP-2) and cardiovascular events in atrial fibrillation patients: A nested case control study from the ATHERO-AF cohort Danilo Menichelli, Emanuela Falcinelli, Pasquale Pignatelli, Roberto Carnevale, Alessandra D'Amico, Francesco Violi, Paolo Gresele, Daniele Pastori, Francesco Paciullo Thrombosis Research, 2026 Atrial fibrillation (AF) is associated with an increased risk of major adverse cardiovascular events (CVEs) including myocardial infarction, ischemic stroke and peripheral arterial ischemic events despite optimal anticoagulant therapy. Matrix metalloproteinase 2 (MMP-2) levels have been associated with endothelial dysfunction and atherosclerosis, and in AF patients, MMPs seem to play a role in atrial fibrosis but no solid data on the association with CV risk do exist. We measured MMP-2 plasma levels, and its selective naturally occurring inhibitor TIMP-2 in 211 AF patients with (n = 86) or without (n = 125) CVEs, in a nested case-control study from the prospective ATHERO-AF study. The two groups were balanced for clinical characteristics. The median value of MMP-2 was 731 ng/ml in patients with and 953 ng/ml in patients without CVEs (p < 0.05). ROC curve analysis identified a cut-off value of 825 ng/ml associated with CVEs. Patients with MMP-2 above this level had a lower incidence of CVEs (OR: 0.38, CI 0.2-0.6) and stroke (OR 0.37 CI 0.1-0.9). Additionally, multivariable Cox proportional hazards analysis confirmed an inverse association between high levels of MMP-2, CVEs and stroke (HR 0.563, 95%CI 0.358-0.886, p = 0.013 and HR 0.335, 95%CI 0.146-0.771, p = 0.010, respectively). Results were confirmed using continuous values. These data suggest a possible protective role of MMP-2 against CVEs occurrence in patients with AF and highlight the need of further investigations on the potential role of MMP-2 as a prognostic biomarker in non-valvular AF.
Thrombaxane-B2 as a predictor marker of atrial fibrillation Luigi Petramala, Roberto Carnevale, Luca Marino, Luca D'Ambrosio, Simona Bartimoccia, Elisa Alessandra D'Amico, Vittorio Picchio, Angela Leonardo, Elisa Dietrich, Maurizio Forte, Giacomo Frati, Gioacchino Galardo, Francesco Circosta, Sebastiano Sciarretta, Claudio Letizia International Journal of Cardiology Cardiovascular Risk and Prevention, 2026 Atrial fibrillation (AF) is the most frequent cardiac arrhythmia in clinical practice, and its global health burden is progressively rising due to the aging of the world population, progressively higher prevalence of obesity, and improved attention to its early detection; AF is linked to increased risk of several complications, as well as death, stroke and peripheral embolism. In recent years many efforts have been made to fully understand the pathophysiology of AF and its complications. Several epidemiological studies have been conducted to early identification of AF, in order to pay more attention to AF risk factors, including arterial hypertension, dyslipidemia, type 2 diabetes mellitus, obstructive sleep apnea syndrome (OSAS), physical inactivity, and alcohol excess. While the traditional view attributes the primary cause of AF onset to myocardial remodelling (i.e. atrial dilatation, ventricular hypertrophy), cutting-edge research has increasingly highlighted a pivotal role for systemic and myocardial inflammation in the etiology of this arrhythmia. In this study we aimed to evaluate the clinical presentation and the inflammatory immunophenotype of individuals presenting with AF episode, evaluating several biomarkers, such as IFN-γ and thromboxane B2 (TXB 2 ), a potent vasoconstrictor and promoter of platelet aggregation.
Platelet Function and Markers of Atherothrombotic Risk in Individuals With Parathyroid Disorders Anda Mihaela Naciu, Annunziata Nusca, Andrea Palermo, Francesco Piccirillo, Gaia Tabacco, Alessandra D’Amico, Alfonso Maria Di Tommaso, Giulia Sterpetti, Michele Mattia Viscusi, Federico Bernardini, Maurizio Forte, Luca D’Ambrosio, Giacomo Frati, Cristina Nocella, Nicola Napoli, Roberto Carnevale, Sebastiano Sciarretta, Francesco Grigioni Journal of Clinical Endocrinology and Metabolism, 2025 Context Both primary hyperparathyroidism (PHPT) and chronic hypoparathyroidism (HypoPT) are associated with the onset and development of cardiovascular diseases (CVDs). However, the molecular mechanisms underlying the effects of parathyroid disorders on endothelial dysfunction and platelet aggregation, two main determinants of CVDs, are not completely understood. Objective This work aimed to evaluate the effects of PHPT and HypoPT on oxidative stress and endothelial and platelet function. Methods This monocentric cross-sectional study at an outpatient clinic included 40 individuals with HypoPT, 40 with PHPT, and 40 age- and sex-matched control participants. Main outcome measures included circulating levels of markers of oxidative stress, endothelial function, and platelet activation, calcium metabolism parameters, flow-mediated vasodilation (FMD), and carotid intimal-media thickness (IMT). Results HypoPT and PHPT patients showed increased oxidative stress markers as compared to control participants (P &lt; .001). Among patients with parathyroid disorders, those with PHPT demonstrated the highest reduction of nitric oxide (P &lt; .001 vs HypoPT and controls) and FMD (P &lt; .001 and P = .001) and a marked increase of IMT (P &lt; .001 and P = .001). We also observed an increased platelet aggregation in patients with parathyroid disorders, with the highest values in PHPT patients (P &lt; .001, PHPT vs controls; P = .006, HypoPT vs controls; P &lt; .001, PHPT vs HypoPT), along with increased levels of soluble P selectin and thromboxane B2. Conclusion PHPT and HypoPT patients have increased markers of atherothrombotic risk due to endothelial and platelet function alterations. Our results suggests that parathyroid hormone may influence platelet reactivity. Further research is needed to determine if personalized antiplatelet therapy is necessary in individuals with parathyroid disorders.
Tobacco Smoke Exposure and Oxidative Stress: The Role of Circulating Lipopolysaccharides in Heated and Conventional Products Lorenzo Loffredo, Enrico Maggio, Simona Bartimoccia, Arianna Magna, Chiara Maria Totè, Chiara Bagnato, Bianca Laura Cinicola, Federica Armeli, Angela Leonardo, Alessandra D’Amico, Ernesto Greco, Giacomo Frati, Giuseppe Biondi-Zoccai, Alberto Spalice, Antonio Angeloni, Pasquale Pignatelli, Francesco Violi, Anna Maria Zicari, Roberto Carnevale, Smoking Prevention Study Group Antioxidants, 2025 Background: Exposure to tobacco smoke, from conventional tobacco cigarettes (CTC) or heated tobacco products (HTPs), increases oxidative stress, causing endothelial dysfunction and higher cardiovascular risk. It is unclear whether smoke exposure also promotes low-grade endotoxemia, potentially activating NADPH oxidase and further impairing endothelial function. This study assessed serum lipopolysaccharide (LPS) levels in children and adults actively or passively exposed to conventional cigarette smoke or HTPs, compared with non-exposed controls. Methods: We conducted a cross-sectional study comprising 26 children passively exposed to HTPs, 26 children exposed to CTC, and 26 unexposed controls, as well as 20 adult chronic HTP users, 20 chronic CTC, and 20 non-smoking adults. Circulating LPS was measured alongside oxidative stress markers (NOX2, H2O2), endothelial function, intestinal permeability (zonulin), and nicotine exposure (serum cotinine). Results: Exposed children had higher cotinine, LPS, and zonulin than controls, with no differences between HTP and CTC groups. Multiple linear regression analysis identified cotinine (β = 0.343; p = 0.005) and zonulin (β = 0.441; p < 0.001) as independent LPS predictors. In adults, LPS and zonulin were higher in both smoker groups versus controls; zonulin (β = 0.477; p < 0.001) and nitric oxide bioavailability (β = −0.307; p = 0.007) independently predicted LPS. Conclusions: Passive and active exposure to CTC or HTPs increases low-grade endotoxemia and zonulin, potentially driving NOX2-mediated oxidative stress.
PTH-driven modulation of platelet activity via the NOX2 pathway in postsurgical hypoparathyroidism Alessandra D'Amico, Gaia Tabacco, Cristina Nocella, Anda Mihaela Naciu, Annunziata Nusca, Francesco Piccirillo, Michele Mattia Viscusi, Federico Bernardini, Valentina Valenti, Angela Leonardo, Sebastiano Sciarretta, Ernesto Greco, Gianmarco Sarto, Beatrice Simeone, Luca D'Ambrosio, Giacomo Frati, Athanasios D. Anastasilakis, Francesco Grigioni, Nicola Napoli, Maurizio Forte, Andrea Palermo, Roberto Carnevale Redox Biology, 2025 Postsurgical chronic hypoparathyroidism (HypoPT) has been linked to an increased cardiovascular risk, but the underlying pathophysiological mechanisms remain incompletely understood. Emerging evidence suggests a potential direct role of parathyroid hormone (PTH) in modulating platelet function and oxidative stress, both contributors to atherothrombosis. Our study aimed to investigate the impact of PTH on platelet function and activation, with a particular focus on NOX2-mediated platelet activation in patients with HypoPT. We conducted a cross-sectional study involving 24 patients with HypoPT and 40 age- and sex-matched healthy controls. Clinical, biochemical, and platelet function parameters were assessed. In a subgroup of five HypoPT patients, changes were evaluated after 24 months of PTH (1-34) therapy. Platelet aggregation, oxidative stress biomarkers (sNOX2-dp, H 2 O 2 , 8-OHdG), and thrombus formation (T-TAS) were measured. The in vitro effect of PTH (1-34) was tested on isolated platelets. Patients with HypoPT exhibited enhanced platelet activation, increased oxidative stress, and accelerated thrombus formation compared to controls. Enhanced platelet activation and increased oxidative stress observed in HypoPT were further amplified in HypoPT subjects treated with PTH (1-34). In vitro, PTH (1-34) increased oxidative stress and platelet aggregation only in platelets from HypoPT patients, through a specific signaling pathway involving PTH1R activation, intracellular calcium release, protein kinase C (PKC) activation and NOX2-dependent ROS generation. HypoPT is associated with heightened platelet reactivity and thrombotic risk. PTH therapy may exacerbate these alterations through a defined molecular mechanism. These findings highlight the need for careful cardiovascular monitoring in HypoPT patients, particularly those receiving PTH analogues. • Hypoparathyroidism is linked to enhanced oxidative stress and platelet activation. • Recombinant PTH (1-34) exacerbates platelet activation and thrombus formation in HypoPT. • PTH signaling involves PTH1R–calcium–PKC–NOX2 axis in platelets. • In vitro, PTH increases oxidative stress and platelet aggregation in HypoPT but not in control platelets. • Findings support cardiovascular risk evaluation in HypoPT patients on PTH analogues.
Impact of Hospitalization on Sarcopenia, NADPH-Oxidase 2, Oxidative Stress, and Low-Grade Endotoxemia in Elderly Patients Chiara Bagnato, Arianna Magna, Elena Mereu, Sciaila Bernardini, Simona Bartimoccia, Roberta Marti, Pietro Enea Lazzerini, Alessandra D’Amico, Evaristo Ettorre, Giovambattista Desideri, Pasquale Pignatelli, Francesco Violi, Roberto Carnevale, Lorenzo Loffredo, and Antioxidants, 2025 Background: Hospitalization in older adults often worsens sarcopenia due to prolonged bed rest, poor nutrition, and inactivity. This study examined how hospitalization impacts muscle mass, focusing on oxidative stress and gut-derived endotoxemia. Methods: Thirty-one hospitalized older adults were compared with 31 outpatients. Ultrasound was used to measure the thickness of the rectus femoris (RF), intercostal, and diaphragmatic muscles at admission and discharge. Serum levels of LPS, zonulin, sNOX2-dp, and H2O2 were also assessed. Results: Hospitalized patients had higher serum levels of sNOX2-dp, H2O2, LPS, and zonulin than outpatients. In hospitalized patients, significant increases were observed at discharge compared to admission levels in sNOX2-dp (20.9 ± 6.5 to 23.8 ± 7.5 pg/mL; p = 0.004), H2O2 (24.4 ± 9.8 to 32.8 ± 14.5 µM; p = 0.01), LPS (30.4 ± 12.6 to 43.3 ± 16.35 pg/mL; p < 0.001), and zonulin (2.06 ± 1.23 to 2.95 ± 1.33 ng/mL; p < 0.001). Ultrasound data revealed a reduction in RF muscle thickness (−35%) (0.58 ± 0.29 to 0.38 ± 0.31 cm, p < 0.001), intercostal muscle thickness (−28%) (0.22 ± 0.08 to 0.16 ± 0.06 cm, p < 0.001), and diaphragmatic muscle thickness (−26%) (0.19 ± 0.06 to 0.14 ± 0.04 cm, p < 0.001) at discharge compared to admission. Additionally, muscle strength, measured using the hand-grip test, showed a 25% reduction. Regression analysis revealed correlations between RF muscle loss and increases in sNOX2-dp and H2O2, as well as between NOX2, H2O2, and LPS with zonulin. Conclusions: Hospitalization in older adult patients elevates NOX2 blood levels, correlating with reduced muscle mass. Increased low-grade endotoxemia may trigger NOX2 activation, generating oxidative stress that accelerates muscle degeneration and can lead to sarcopenia.
Multidisciplinary Clinical Study on Retinal, Circulatory, and Respiratory Damage in Smoking-Dependent Subjects Marcella Nebbioso, Annarita Vestri, Magda Gharbiya, Mattia D’Andrea, Matteo Calbucci, Federico Pasqualotto, Serena Esposito, Alessandra D’Amico, Valentina Castellani, Sandra Cinzia Carlesimo, Paolo Giuseppe Limoli, Alessandro Lambiase Medicina Lithuania, 2025 Background and Objectives: Cigarette smoking is a widely prevalent risk factor in the global population, despite its well-recognized systemic impact. In this pilot study, an association was hypothesized between alterations in hemorheological and respiratory characteristics and damage at the chorioretinal level, considering that traditional cigarette smoking may increase oxidative stress, platelet activation, and thrombotic phenomena at the systemic level. Fundoscopy can provide information about the characteristics of the cerebral district and the entire circulatory system. Therefore, the aim of this research was to evaluate the impact of cigarette smoking on chorioretinal vascularization and pulmonary and blood parameters through investigations with optical coherence tomography angiography (OCTA), spirometry, and the total thrombus formation analysis system (T-TAS). Materials and Methods: Thirty subjects were recruited, divided into 20 traditional cigarette smokers (SMs) and 10 non-SMs, who underwent a comprehensive ocular examination, including OCTA. Spirometric evaluation and blood sampling were also performed on both groups to study pulmonary functional capacity, as well as T-TAS. Results: An analysis of the obtained data confirmed the systemic impact of smoking, evidenced by an increase in T-TAS and a decrease in forced expiratory volume in 1 s expressed in liters (FEV1 L) in SMs compared to the non-SMs group. Additionally, OCTA revealed a statistically significant alteration in macular vascular density (FD) in the right eye (RE) of the examined SMs. The other parameters evaluated did not show statistically significant differences. Conclusions: It is believed that FD, FEV1, and T-TAS may be promising values in correlating the alterations observed in SMs, as highlighted by the changes detected with OCTA, spirometry, and hemorheological data. Further research is needed to confirm and expand the results already obtained and to evaluate the systemic vascular damage and oxidative stress caused by tobacco consumption.
Atrial natriuretic peptide (ANP) modulates stress-induced autophagy in endothelial cells Maurizio Forte, Simona Marchitti, Flavio di Nonno, Donatella Pietrangelo, Rosita Stanzione, Maria Cotugno, Luca D'Ambrosio, Alessandra D'Amico, Vittoria Cammisotto, Gianmarco Sarto, Erica Rocco, Beatrice Simeone, Sonia Schiavon, Daniele Vecchio, Roberto Carnevale, Salvatore Raffa, Giacomo Frati, Massimo Volpe, Sebastiano Sciarretta, Speranza Rubattu Biochimica Et Biophysica Acta Molecular Cell Research, 2025 Atrial natriuretic peptide (ANP), a cardiac hormone involved in the regulation of water/sodium balance and blood pressure, is also secreted by endothelial cells, where it exerts protective effects in response to stress. Autophagy is an intracellular self-renewal process involved in the degradation of dysfunctional cytoplasmic elements. ANP was recently reported to act as an extracellular regulator of cardiac autophagy. However, its role in the regulation of endothelial autophagy has never been investigated. Here, we tested the effects of ANP in the regulation of autophagy in human umbilical vein endothelial cells (HUVECs). We found that ANP rapidly increases autophagy and autophagic flux at physiological concentrations through its predominant pathway, mediated by natriuretic peptide receptor type A (NPR-A) and protein kinase G (PKG). We further observed that ANP is rapidly secreted by HUVEC under stress conditions, where it mediates stress-induced autophagy through autocrine and paracrine mechanisms. Finally, we found that the protective effects of ANP in response to high-salt loading or tumor necrosis factor (TNF)-α are blunted by concomitant inhibition of autophagy. Overall, our results suggest that ANP acts as an endogenous autophagy activator in endothelial cells. The autophagy mechanism mediates the protective endothelial effects exerted by ANP. • Atrial natriuretic peptide (ANP) stimulates autophagy in endothelial cells. • ANP-induced autophagy is mediated by the NPR-A/PKG/cGMP pathway. • ANP is secreted by endothelial cells in response to stress and regulates stress-induced autophagy. • The protective endothelial effects of ANP are mediated by autophagy activation.
Stenosing Crohn’s disease patients display gut autophagy/oxidative stress imbalance Leonardo Schirone, Maria Carla Di Paolo, Daniele Vecchio, Cristina Nocella, Alessandra D’Amico, Riccardo Urgesi, Lorella Pallotta, Gianfranco Fanello, Giuseppe Villotti, Maria Giovanna Graziani, Mariangela Peruzzi, Elena De Falco, Roberto Carnevale, Sebastiano Sciarretta, Giacomo Frati, Cristiano Pagnini Scientific Reports, 2024 In this pilot study, we assessed the role of autophagy in Crohn's Disease (CD), particularly in patients with a stenosing phenotype. Through the analysis of biopsied specimens from 36 patients, including 11 controls and 25 CD patients, categorized into inflammatory and stenosing groups, we identified a significant reduction in the autophagosomal marker Lc3b-II in patients with active inflammation and stenosis. This was paralleled by an increase in oxidative stress markers, including sNOX2-dp and H2O2, and a decrease in the antioxidant capacity measured by HBA, suggesting an imbalance in autophagy and oxidative stress mechanisms. Additionally, our findings show a correlation between autophagy markers and oxidative stress levels, indicating that autophagy dysfunction may play a pivotal role in CD and in the progression of a stenosing disease phenotype, by failing to eliminate detrimental molecules and pathogenic bacteria, thereby promoting fibrosis. This study is the first to demonstrate in vivo autophagy inhibition in stenosing CD patients and suggests that stimulating autophagic processes could offer a new avenue for the prevention and treatment of intestinal fibrosis in CD. Our results highlight the importance of exploring the interactions between autophagy, the fibrotic process, and the inflammatory cascade, opening avenues for potential therapeutic interventions in CD management.
Exposure to serum from exclusive heated tobacco product smokers induces mTOR activation and fibrotic features in human cardiac stromal cells Vittorio Picchio, Francesca Pagano, Roberto Carnevale, Alessandra D'Amico, Claudia Cozzolino, Erica Floris, Antonella Bordin, Leonardo Schirone, Daniele Vecchio, Wael Saade, Fabio Miraldi, Elena De Falco, Sebastiano Sciarretta, Mariangela Peruzzi, Giuseppe Biondi-Zoccai, Giacomo Frati, Isotta Chimenti Biochimica Et Biophysica Acta Molecular Basis of Disease, 2024 Chronic smokers have increased risk of fibrosis-related atrial fibrillation. The use of heated-tobacco products (HTPs) is increasing exponentially, and their health impact is still uncertain. We aim to investigate the effects of circulating molecules in exclusive HTP chronic smokers on the fibrotic behavior of human atrial cardiac stromal cells (CSCs). CSCs were isolated from atrial tissue of elective cardiac surgery patients, and exposed to serum lots from young healthy subjects, stratified in exclusive HTP smokers, tobacco combustion cigarette (TCC) smokers, or nonsmokers (NS). CSCs treated with TCC serum displayed impaired migration and increased expression of pro-inflammatory cytokines. Cells cultured with HTP serum showed increased levels of pro-fibrotic markers, and reduced expression of connexin-43. Both TCC and HTP sera increased collagen release and reduced secretion of angiogenic protective factors from CSCs, compared to NS serum. Paracrine support to tube-formation by endothelial cells and to viability of cardiomyocytes was significantly impaired. Treatment with sera of both smokers groups impaired H2O2/NO release balance by CSCs and reduced early phosphorylation of several pathways compared to NS serum, leading to mTOR activation. Cotreatment with rapamycin was able to reduce mTOR phosphorylation and differentiation into aSMA-positive myofibroblasts in CSCs exposed to TCC and HTP sera. In conclusion, the circulating molecules in the serum of chronic exclusive HTP smokers induce fibrotic behavior in CSCs through activation of the mTOR pathway, and reduce their beneficial paracrine effects on endothelial cells and cardiomyocytes. These results point to a potential risk for cardiac fibrosis in chronic HTP users.
Neutrophil Cathepsin G and Thrombosis in COVID-19 Francesco Violi, Simona Bartimoccia, Roberto Cangemi, Cristina Nocella, Alessandra D’Amico, Alessandra Oliva, Mario Venditti, Francesco Pugliese, Claudio Maria Mastroianni, Lorenzo Ridola, Miriam Lichtner, Vincenzo Cardinale, Domenico Alvaro, Vittoria Cammisotto, Valentina Castellani, Pasquale Pignatelli, Roberto Carnevale Circulation Research, 2024
NOX2 as a Biomarker of Academic Performance: Evidence from University Students during Examination Cristina Nocella, Alessandra D’Amico, Roberto Cangemi, Chiara Fossati, Fabio Pigozzi, Elena Mannacio, Vittoria Cammisotto, Simona Bartimoccia, Valentina Castellani, Gianmarco Sarto, Beatrice Simeone, Erica Rocco, Giacomo Frati, Sebastiano Sciarretta, Pasquale Pignatelli, Roberto Carnevale, SMiLe Group SMiLe Group Antioxidants, 2024
Impact of heat-not-burn cigarette passive smoking on children's oxidative stress, endothelial and platelet function Lorenzo Loffredo, Roberto Carnevale, Arianna Pannunzio, Bianca Laura Cinicola, Ilaria Maria Palumbo, Simona Bartimoccia, Cristina Nocella, Vittoria Cammisotto, Francesco Violi, Giuseppe Biondi-Zoccai, Giacomo Frati, Anna Maria Zicari, Arianna Magna, Raffaella Izzo, Martina Capponi, Giulia Brindisi, Francesca Salvatori, Valentina Castellani, Alessandra D’ Amico, Chiara Trivigno, Chiara Totè, Enrico Maggio, Fabio Miraldi, Marzia Duse, Pasquale Pignatelli, Alberto Spalice, Flavia Antonucci Environmental Pollution, 2024
Low Grade Endotoxemia and Oxidative Stress in Offspring of Patients with Early Myocardial Infarction Bianca Laura Cinicola, Ilaria Maria Palumbo, Arianna Pannunzio, Roberto Carnevale, Simona Bartimoccia, Vittoria Cammisotto, Martina Capponi, Giulia Brindisi, Francesca Salvatori, Francesco Barillà, Francesco Martino, Alessandra D’Amico, Roberto Poscia, Alberto Spalice, Anna Maria Zicari, Francesco Violi, Lorenzo Loffredo Antioxidants, 2023
Low-grade endotoxemia and NOX2 in patients with coronary microvascular angina Lorenzo Loffredo, Victoria Ivanov, Nicolae Ciobanu, Micaela Ivanov, Paolo Ciacci, Cristina Nocella, Vittoria Cammisotto, Federica Orlando, Aurora Paraninfi, Enrico Maggio, Alessandra D’Amico, Paolo Rosa, Mihail Popovici, Simona Bartimoccia, Francesco Barillà, Elena Deseatnicova, Evgenii Gutu, Francesco Violi, Roberto Carnevale Kardiologia Polska, 2022
