Beyond readthrough: ataluren restores mitochondrial function and reduces oxidative stress in FANCA-mutated cells via mTOR–DRP1 modulation Matilde Balbi, Elisa Guidi, Anca Manuela Hristodor, Fabio Corsolini, Vanessa Cossu, Roberta Bottega, Martina Serra, Sara Pestarino, Martina Bartolucci, Marco Cipolli, Stefano Regis, Valentino Bezzerri, Enrico Cappelli, Silvia Ravera Cell Death Discovery, 2026 Fanconi anemia (FA) is a rare inherited bone marrow failure syndrome characterized by genomic instability, mitochondrial dysfunction, and oxidative stress. While the therapeutic potential of ataluren, a translational readthrough-inducing drug, has been investigated in FA cells carrying nonsense mutations, its broader metabolic impact remains unclear. Here, we demonstrate that ataluren (tested at 2.5, 5, and 10 μM) modulates cellular energy metabolism and redox homeostasis in FA lymphoblasts harboring either nonsense or missense mutations in the FANCA gene. At low doses (2.5 μM for 72 h), ataluren improved the ATP/AMP ratio, enhanced oxidative phosphorylation efficiency, and reduced lipid peroxidation and oxidative DNA damage. These effects were independent of mutation type and were not associated with compensatory glycolysis, as lactate dehydrogenase activity remained unchanged. Strikingly, ataluren restored the P/O ratio under pyruvate/malate-driven respiration to near-normal values, indicating improved coupling between oxygen consumption and ATP synthesis. Mechanistically, ataluren reduced DRP1 protein levels and attenuated mTOR-S6 signaling, suggesting that mitochondrial dynamics and bioenergetic efficiency are modulated via the mTOR–DRP1 axis. Additionally, ataluren lowered IMPDH activity, contributing to reduced cell proliferation and DNA damage without impairing cellular energy status. Notably, these beneficial effects persisted under immune stimulation, where ataluren mitigated the metabolic and oxidative burden imposed by lymphocyte activation. Our findings unveil a pleiotropic role for ataluren that extends beyond its canonical readthrough activity, highlighting its potential as a metabolic modulator for FA and possibly other DNA repair–deficient disorders.
IgM Hyposialylation Modulates Podocyte Vulnerability in Patients With Idiopathic Nephrotic Syndrome Sonia Spinelli, Sofia Gaudiano, Andrea Garbarino, Francesca Lugani, Edoardo La Porta, Andrea Petretto, Martina Bartolucci, Chiara Lavarello, Nicole Grinovero, Ilaria Musante, Paolo Scudieri, Antonella Trivelli, Giorgio Piaggio, Alberto Magnasco, Maria Ludovica Degl’Innocenti, Simona Granata, Gianluigi Zaza, Enrico Verrina, Giovanni Candiano, Maurizio Bruschi Kidney International Reports, 2026 Introduction: Altered Ig glycosylation has been implicated in antibody-mediated podocytopathies; however, the functional relevance of IgM sialylation remains poorly defined. Previous evidence suggests that circulating cationic or hyposialylated IgM may contribute to podocyte vulnerability in idiopathic nephrotic syndrome (iNS). Methods: Serum IgM from 86 pediatric and adult patients with podocytopathies, 20 patients with membranous nephropathy (MN), 20 patients with lupus nephritis (LN), and 30 healthy controls were analyzed by lectin-based enzyme-linked immunosorbent assay (ELISA) using biotinylated lectins to assess terminal N-glycan residues, including Sambucus nigra agglutinin (SNA) and Ricinus communis agglutinin I (RCA-I). Serum levels of the sialyltransferase ST6GAL1 and the sialidases neuraminidase-1 (NEU1) and neuraminidase-3 (NEU3) were quantified. Cultured human podocytes were exposed to native, desialylated, or resialylated IgM and analyzed by confocal microscopy, quantitative proteomics, phosphoproteomics, and metabolic assays. Results: IgM from patients with iNS showed reduced SNA binding, which inversely correlated with proteinuria and circulating NEU1/NEU3 levels. In paired samples, SNA reactivity decreased during relapse and increased during remission. ST6GAL1 was undetectable across all groups, whereas phospholipase A2 receptor 1(PLA2R1)-positive MN displayed reduced RCA-I binding. Podocytes exposed to hyposialylated or desialylated IgM exhibited disorganization of the actin cytoskeleton, reduced nephrin signal, increased lipid peroxidation, and decreased ATP levels. Resialylated IgM displayed podocyte morphological and metabolic features not statistically distinguishable from those observed under control conditions. Proteomic and phosphoproteomic analyses highlighted modulation of mitogen-activated protein kinase (MAPK)-, mechanistic Target of Rapamycin (mTOR-), adenosine monophosphate-activated protein kinase (AMPK), and cytoskeleton-related pathways. Conclusion: IgM sialylation status tracks disease activity and modulates podocyte structural, metabolic, and signaling responses, supporting immune glycan remodeling as a disease-associated modifier of podocyte vulnerability in iNS.
Exploring the impact of age, sex and life experiences on plasma inflammatory profiles through comparative proteomics Martina Bartolucci, Olga Utyro, Anita Muraglia, Alessia Repetto, Vanessa Agostini, Monica Pizzonia, Silvia Ottaviani, Gino Tripodi, Gilberto Filaci, Ranieri Cancedda, Andrea Petretto, Maddalena Mastrogiacomo Frontiers in Immunology, 2026 Background In the heterochronic parabiosis model it has been shown that blood from elderly animals exhibits markedly reduced rejuvenating effects compared to that of young organisms. Furthermore, human plasma from older subjects, when used as a supplement in cell culture media, is significantly less effective than plasma derived from younger individuals. This study analyzed plasma from a cohort of 229 subjects by a proteomic approach to reveal age-related changes. Methods A mass spectrometry-based proteomic analysis was performed on plasma samples from 3 age-groups: a prepubertal, a healthy young adult group and a cohort of individuals over 75 years old with three different life-experiences. An additional parallel study was conducted by a Milliplex Luminex assay. Results The proteomic analysis revealed a chronic inflammatory state in the elderly population, along with complement activation and impaired regulation of blood coagulation. This inflammatory condition was confirmed by Luminex assay, showing elevated levels of classical pro-inflammatory cytokines in the plasma of elderly individuals. Moreover, the elderly group showed a reduced production of antibody light chains, suggesting concurrent immunosenescence. In the older group we identified 25 upregulated proteins whose elevated abundance, combined with acquired immune aging may constitute a plasma proteomic signature of aging. The degree of upregulation of these signature proteins varied among elderly subgroups with different life-experience. A good physical condition and/or cognitive function correlated with a lower expression of the aging-related proteomic profile. Furthermore, several sex-specific differences were identified in the plasma profiles of young donors. Reversely, among elderly individuals, no major differences were observed, except for an increased level of Pregnancy Zone Protein (PZP) in females. Conclusions Proteomic analysis of plasma revealed protein variations associated with aging, primarily involving inflammation-related pathways, immunosenescence features, and sex-linked differences. This study highlights the pathological characteristics underlying the aging process.
Omics data integration analysis identified new biological insights into chronic antibody-mediated rejection (CAMR) Maurizio Bruschi, Simona Granata, Francesca Leone, Laura Barberio, Giovanni Candiano, Paola Pontrelli, Andrea Petretto, Martina Bartolucci, Sonia Spinelli, Loreto Gesualdo, Gianluigi Zaza Journal of Translational Medicine, 2025 BACKGROUND: In the last two decades, many studies based on omics technologies have contributed to defining the clinical, immunological, and histological fingerprints of chronic antibody-mediated rejection (CAMR), the leading cause of long-term kidney allograft failure. However, the full biological machinery underlying CAMR has only been partially defined, likely due to the fact thatsingle-omics technologies capture only specific aspects of the biological system and fail to provide a comprehensive understanding of this clinical complication. METHODS: This study integrated mass spectrometry-based proteomic profiling of serum samples from 19 patients with clinical and histological evidence of CAMR and 26 kidney transplant recipients with normal graft function and histology (CTR) with transcriptomic analysis of peripheral blood mononuclear cells (PBMCs) from an independent cohort of 10 CAMR and 8 CTR patients. Data analysis was conducted using unsupervised hierarchical clustering (multidimensional scaling with k-means) and Spearman's correlation test. Partial least squares discriminant analysis (PLS-DA) with the importance in projection (VIP) score identified key proteins differentiating CAMR from CTR. ELISA was used to validate the omics results. RESULTS: Proteomic analysis identified 18 proteins that significantly differentiated CAMR from CTR (p < 0.01): five were more abundant (CHI3L1, LYZ, PRSS2, CPQ, IGLV3-32), while 13 were less abundant (SERPINA5, SERPING1, KNG1, CAMP, VNN1, BTD, WDR1, PON3, AHNAK2, MELTF, CA1, CD44, CUL1). Transcriptomic profiling revealed 6 downregulated and 33 upregulated genes in CAMR versus CTR (p < 0.01). Notably, only 2 biological elements were significantly deregulated in both omics analyses: chitinase-3-like protein 1 (CHI3L1) and plasma protease inhibitor C1 (SERPING1). CHI3L1, previously associated with the severity of tissue damage in kidney diseases, was up-regulated in CAMR in both transcriptomics and proteomics, while SERPING1, a serine esterase inhibitor that blocks the classical and lectin pathway of complement, was up-regulated in CAMR in transcriptomics but down-regulated in proteomics. ELISA validated the omics results, and the ROC curve showed that CHI3L1 has good discrimination power between CAMR and CTR (AUC of ROC curve of 0.81). CONCLUSIONS: Our multi-omics data, although performed in a relatively small cohort of patients, revealed new systemic biological elements involved in the pathogenesis of CAMR and identified CHI3L1 as a new potential biomarker and/or therapeutic target for this important clinical complication. Future validation of these findings in larger patient cohorts should be conducted to better evaluate their clinical utility.
NET Proteomic Profiling Reveals New Pathways Potentially Implicated in Dendritic Cell-Mediated Inflammation in DADA2 Patients Sara Signa, Martina Bartolucci, Martina Bonacini, Arinna Bertoni, Genny Del Zotto, Anna Corcione, Andrea Petretto, Silvia Della Bella, Roberta Bertelli, Dario Di Silvestre, Andrea Lomagno, Pierluigi Mauri, Roberta Caorsi, Maurizio Bruschi, Simone Balin, Paola Bocca, Stefano Volpi, Maria Grazia Catanoso, Alessia Cafaro, Gino Tripodi, Lorenzo Pellottieri, Domenico Mavilio, Antonella Insalaco, Stefania Croci, Carlo Salvarani, Marco Gattorno, Francesca Schena Journal of Clinical Immunology, 2025 PURPOSE: Adenosine deaminase 2 Deficiency (DADA2) is an autoinflammatory disease characterized by systemic vasculopathy, strokes and mild immunodeficiency. Recently NETosis has been implicated in the pathogenesis of Deficiency of Adenosine Deaminase 2. To deep investigate the possible effects of NETs on the immune system we characterized proteomic profile of NETs from DADA2 as compared to HD and Polyarteritis Nodosa (PAN) patients. To determine if NETs contain possibly immunogenic antigens we study functional aspects on Dendritic Cells after in vitro stimulation with NETs. METHODS: Twenty-three DADA2 patients were enrolled. We analyzed NETosis by Imaging Flow Citometry. We evaluated NETs remnants and DNAse in the plasma samples by ELISA assay whereas DNAse activity by DNA digestion. We used quantitative proteomics approach and network analysis to identify NET proteins and pathways in 6 DADA2, 7 PAN and 7 HD. We analyzed circulating and monocyte-derived dendritic cells by flow cytometry. RESULTS: Neutrophils from DADA2 patients show a significant increased suicidal NETosis. DNAse enzymes were not normal in the level or activity. By proteomic analysis we identified 1356 proteins among which a hundred of proteins were significantly up or down-modulated in DADA2 NETs as compared to normal and disease control NETs in resting condition and after stimulation with PMA, Adenosine and TNFα. DADA2 NETs are significantly more efficient than normal NETs in stimulating patients' monocyte-derived dendritic cells. CONCLUSION: We identified different pathways significantly modulated in DADA2 NETs versus PAN/HD NETs. This peculiar protein profile could contribute in activating inflammatory pathways in Dendritic cells in DADA2.
Dysregulated cortical excitability and tau phosphorylation in a β3 integrin mouse model of autism Carmela Vitale, Fanny Jaudon, Rafael Luján, Martina Bartolucci, Lucia Celora, Elisa Reisoli, Riccardo Ruggeri, Andrea Petretto, Agnes Thalhammer, Lorenzo A Cingolani Brain, 2025 Autism spectrum disorder is a complex neurodevelopmental disease characterized by altered cortical network excitability. Recent genetic studies have identified deep layer V cortical pyramidal neurons in the frontal cortex as central to autism pathophysiology, yet the cortical circuits, plasticity mechanisms and molecular signalling pathways involved remain poorly understood. Layer V pyramidal neurons consist of two main types with distinct functional roles: intratelencephalic neurons, which respond to low-frequency stimulation and project within the cortex and striatum, and pyramidal tract neurons, which are tuned to theta-frequency inputs and convey information to subcortical structures. Determining which of these two neuron types is more critical to autism pathophysiology and whether disruptions in their synaptic connectivity or intrinsic excitability contribute to autism-related dysfunctions would significantly advance our understanding of the disorder. Integrins, a family of cell adhesion molecules, are vital for neuronal function. The gene encoding β3 integrin (ITGB3) is genetically linked to autism spectrum disorder, with rare mutations identified in affected individuals, while Itgb3 knockout mice exhibit autism-like behaviours, including impaired social memory and increased grooming. However, it remains unclear why loss of β3 integrin is associated with autism spectrum disorder, how it disrupts cortical circuits, and which plasticity mechanisms and molecular pathways are involved. Here, we demonstrate that β3 integrin selectively regulates the excitability of pyramidal tract neurons in the medial prefrontal cortex. Using electrophysiology, proteomics and molecular approaches, we show that β3 integrin regulates the gain, adaptation and precision of action potential discharge by controlling the surface expression of Ca2+-activated SK2 channels. Genetic ablation of Itgb3 impaired intrinsic excitability and SK2 channel function in pyramidal tract neurons, with no effects in intratelencephalic neurons. Furthermore, we identified Tau, a protein traditionally linked to neurodegenerative diseases, as part of the SK2 channel interactome. Proteomic analyses revealed altered protein kinase A-dependent phosphorylation of Tau in Itgb3 knockout mice, while protein kinase A inhibition restored SK2 channel currents, thereby connecting phosphorylation changes to excitability deficits. Our findings expand the current mechanistic framework linking signalling pathway dysfunctions to cortical excitability deficits, highlighting the dysregulation of pyramidal tract neuron excitability as a core feature of autism pathophysiology and demonstrating the involvement of β3 integrin, SK2 channels, Tau and PKA in this process. Because pyramidal tract neurons serve as final integrators of cortical computations before relaying information outside the cortex, their impaired excitability may disrupt communication with subcortical targets, contributing to the complex pathophysiology of autism spectrum disorder.
Polydopamine Nanoparticles as a Potential Non-Pharmaceutical Antioxidant Tool against Mitochondrial Disorders Matteo Battaglini, Francesco Schiavone, Alessio Carmignani, Attilio Marino, Valentina Naef, Andrea Petretto, Martina Bartolucci, Fabiana Longo, Francesca Maltecca, Filippo Maria Santorelli, Gianni Ciofani ACS Applied Nano Materials, 2025 Mitochondrial disorders are hereditary diseases caused by mutations in nuclear or mitochondrial DNA that impair organelle function.Key features include excessive production of reactive oxygen species (ROS), mitochondrial abnormalities, and metabolic dysfunctions.Systemically, these defects can lead to severe conditions affecting the central nervous system, muscles, heart, and gastrointestinal tract.Organic antioxidants such as idebenone and resveratrol have been explored as potential treatments; in the framework of nanotechnological antioxidants, polydopamine nanoparticles (PDNPs), derived from the oxidative self-polymerization of dopamine, are highly biocompatible, biodegradable, easy to functionalize, and possess potent ROSscavenging and photothermal properties.In this study, we investigated PDNPs as a nonpharmaceutical therapy for mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes (MELAS) and progressive external ophthalmoplegia (PEO).PDNPs were evaluated in fibroblasts from healthy donors and patients with MELAS and PEO.Molecular characterization was performed via proteomic analysis, followed by assessment of PDNP biocompatibility, internalization, intracellular localization, and antioxidant effects.Their protective activity was also confirmed in vivo, exploiting zebrafish embryos.Our findings demonstrate that PDNPs effectively protect cells from ROS-induced damage, oxidative stress, apoptosis, and mitochondrial dysfunction.Additionally, PDNPs were able to preserve zebrafish embryos against pro-oxidative stimuli.Overall, this work highlights the potential of polydopamine nanostructures as promising therapeutic tools for mitigating the molecular hallmarks of mitochondrial disorders and supporting future clinical applications.
Evaluation of the Effects of Sumac (Rhus coriaria) Extract-Loaded Ethosomes on an In Vitro Wound Healing Model Melis Emanet, Matteo Battaglini, Alessio Carmignani, Federico Catalano, Martina Bartolucci, Andrea Petretto, Gianni Ciofani ACS Omega, 2025 Wound healing involves a series of complex bioprocesses, including repairing skin damage, maintaining its barrier features, and preserving all other skin functions. Since the skin is the primary organ exposed to external factors, these bioprocesses can be interrupted by potential exogenous toxicants. Efforts to mitigate the effects of these toxicants can help accelerate the healing process, facilitating complete wound recovery. In this context, sumac () extract, rich in polyphenolics with antioxidant and anti-inflammatory properties, can be exploited to overcome oxidant and inflammation-dependent burdens. Ethosomes, lipid-based intradermal delivery vehicles, have been selected for the delivery of sumac extract, as they enhance penetration through the skin layers. Considering their remarkable flexibility and deformability, ethosomes can minimize drug leakage even under harmful penetration conditions. Given the diverse bioactive content of sumac extract, ethosomes have been considered ideal for delivering both hydrophilic and lipophilic active compounds. Sumac extract (SuExt)-loaded ethosomes (SuExt-ethosomes) were therefore produced and characterized. These nanocarriers demonstrated significant cellular internalization and cytocompatibility in human dermal fibroblasts (HDFs), along with excellent antioxidant and anti-inflammatory activity. A comprehensive investigation, supported by proteomic analysis, revealed that SuExt-ethosomes present promising wound healing potential, supporting future investigations in preclinical models.
Proteomics characterization of mule milk throughout lactation Diana Fanelli, Martina Bartolucci, Cristina Lamberti, Duccio Panzani, Carlotta Bocci, Rebecca Moroni, Nicole Grinovero, Simona Cirrincione, Giovanna Monti, Maria Gabriella Giuffrida, Francesco Camillo, Andrea Petretto, Andrea Degl’Innocenti Heliyon, 2025 Cow milk is a widespread food, but several children are allergic to it – chiefly because of adverse reactions to its proteins. Nutritionally valid, hypoallergenic surrogates are highly sought after, and among them donkey milk is prized for its tolerability and human-like composition. Still, donkeys produce very little milk, with disastrous consequences for marketability and supply chain. Mules are crosses between a donkey stallion and a horse mare, and female ones can be set pregnant via embryo transfer for later lactation, conceivably within economically sustainable production strategies. Owing to their bigger average body size, mules should produce more milk than donkeys, yet the protein profile and allergenicity of mule milk are vastly unknown. Here we present a proteomics characterization of mule milk, obtained following a mule-in-mule surrogate gestation. In addition, preliminary experiments on blood sera from children allergic to cow milk may provide a first hint that mule milk is less allergenic than its cattle analog.
Two-photon polymerization of miniaturized 3D scaffolds optimized for studies on glioblastoma multiforme in spaceflight-like microgravity conditions Giada Graziana Genchi, Claudio Conci, Özlem Şen, Alessandra Nardini, Martina Bartolucci, Attilio Marino, Rebeca Martinez Vazquez, Giulio Cerullo, Roberto Osellame, Andrea Petretto, Manuela Teresa Raimondi, Gianni Ciofani Biofabrication, 2025 The obtainment of innovative models recalling complex tumour architectures and activities in vitro is a challenging drive in the understanding of pathology molecular bases, yet it is a crucial path to the identification of targets for advanced oncotherapy. Cell environment recapitulation by 3D scaffolding and gravitational unloading of cell cultures represent powerful means in tumour biomimicry processes, but their simultaneous adoption has consistently been explored only in the latest decade. Here, an unprecedented bioengineering approach capitalizing on spaceflight biology practice is proposed for modelling of glioblastoma multiforme, a highly aggressive neoplasm that affects the central nervous system and has poorly effective pharmacological and radiological countermeasures. Tumour modelling was pursued by the original implementation of two-photon polymerization in fast prototyping of 3D scaffolds on flexible substrates for U87-MG glioma cell culture, and by the exposure of cell-laden scaffolds to simulated microgravity (s-μg). Realistic spaceflight conditions were applied to collect preliminary information suitable for testing of U87-MG cell-laden scaffold in low Earth orbit. Responses of glioma cells anchored to 3D scaffolds were investigated by microscopy, quantitative reverse transcription-polymerase chain reaction and proteomic analyses, revealing synergic regulatory effects of cell scaffolding and s-μg on markers of tumour cell growth, metabolism and invasiveness.
Proteomics profiling and machine learning in nusinersen-treated patients with spinal muscular atrophy Chiara Panicucci, Eray Sahin, Martina Bartolucci, Sara Casalini, Noemi Brolatti, Marina Pedemonte, Serena Baratto, Sara Pintus, Elisa Principi, Adele D’Amico, Marika Pane, Marina Sframeli, Sonia Messina, Emilio Albamonte, Valeria A. Sansone, Eugenio Mercuri, Enrico Bertini, Ugur Sezerman, Andrea Petretto, Claudio Bruno Cellular and Molecular Life Sciences, 2024
Extracellular vesicles from II trimester human amniotic fluid as paracrine conveyors counteracting oxidative stress Giorgia Senesi, Laura Guerricchio, Maddalena Ghelardoni, Nadia Bertola, Stefano Rebellato, Nicole Grinovero, Martina Bartolucci, Ambra Costa, Andrea Raimondi, Cristina Grange, Sara Bolis, Valentina Massa, Dario Paladini, Domenico Coviello, Assunta Pandolfi, Benedetta Bussolati, Andrea Petretto, Grazia Fazio, Silvia Ravera, Lucio Barile, Carolina Balbi, Sveva Bollini Redox Biology, 2024
Identification of biochemical and molecular markers of early aging in childhood cancer survivors Silvia Ravera, Tiziana Vigliarolo, Silvia Bruno, Fabio Morandi, Danilo Marimpietri, Federica Sabatini, Monica Dagnino, Andrea Petretto, Martina Bartolucci, Monica Muraca, Eleonora Biasin, Riccardo Haupt, Marco Zecca, Franca Fagioli, Daniela Cilloni, Marina Podestà, Francesco Frassoni Cancers, 2021
Extramitochondrial energy production in platelets Silvia Ravera, Maria Grazia Signorello, Martina Bartolucci, Sara Ferrando, Lucia Manni, Federico Caicci, Daniela Calzia, Isabella Panfoli, Alessandro Morelli, Giuliana Leoncini Biology of the Cell, 2018
Exosomes from human mesenchymal stem cells conduct aerobic metabolism in term and preterm newborn infants Isabella Panfoli, Silvia Ravera, Marina Podestà, Claudia Cossu, Laura Santucci, Martina Bartolucci, Maurizio Bruschi, Daniela Calzia, Federica Sabatini, Matteo Bruschettini, Luca Antonio Ramenghi, Olga Romantsik, Danilo Marimpietri, Vito Pistoia, Gianmarco Ghiggeri, Francesco Frassoni, Giovanni Candiano FASEB Journal, 2016
Human urinary exosome proteome unveils its aerobic respiratory ability Maurizio Bruschi, Laura Santucci, Silvia Ravera, Giovanni Candiano, Martina Bartolucci, Daniela Calzia, Chiara Lavarello, Elvira Inglese, Luca A. Ramenghi, Andrea Petretto, Gian Marco Ghiggeri, Isabella Panfoli Journal of Proteomics, 2016
The human urinary exosome as a potential metabolic effector cargo Maurizio Bruschi, Silvia Ravera, Laura Santucci, Giovanni Candiano, Martina Bartolucci, Daniela Calzia, Chiara Lavarello, Elvira Inglese, Andrea Petretto, Gianmarco Ghiggeri, Isabella Panfoli Expert Review of Proteomics, 2015
