{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,22]],"date-time":"2026-10-22T23:43:56Z","timestamp":1792712636018,"version":"4.5.0"},"reference-count":40,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2026,1,26]],"date-time":"2026-01-26T00:00:00Z","timestamp":1769385600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100002428","name":"Austrian Science Fund","doi-asserted-by":"publisher","award":["10.55776\/KLI1036"],"award-info":[{"award-number":["10.55776\/KLI1036"]}],"id":[{"id":"10.13039\/501100002428","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>The total cavopulmonary anastomosis (Fontan procedure), a palliative procedure for single-ventricle congenital heart disease, improves survival but is associated with progressive multiorgan complications and high long-term morbidity. Prior blood-based proteomic studies in adults have been limited to targeted antibody-based panels or focused on methodological comparisons. Systemic molecular alterations in younger, clinically heterogeneous patients, particularly in untargeted pathways, remain incompletely characterized. Serum samples from 48 Fontan patients and 48 age- and sex-matched healthy controls were analyzed using mass spectrometry with TMT labeling. 2228 proteins were quantified, of which 124 were significantly differentially abundant (fold change &gt; 1.5 or &lt;0.67, FDR-adjusted p &lt; 0.05). Network analysis identified three major functional clusters: extracellular matrix (ECM) organization (predominantly increased), actin cytoskeleton organization, and platelet-related pathways (both predominantly decreased). Stratified analyses showed reduced ECM protein abundance in high-risk patients, suggesting a shift from active remodeling toward a more established fibrotic state, and uniquely elevated cytochrome b5 reductase 3 (CYB5R3), implicating altered redox homeostasis, nitric oxide metabolism, and cellular aging. Overall, our findings extend prior targeted analyses, reveal potential biomarkers such as CYB5R3 and underscore the complexity of the Fontan circulation, with implications for risk stratification and therapeutic targeting.<\/jats:p>","DOI":"10.3390\/ijms27031220","type":"journal-article","created":{"date-parts":[[2026,1,26]],"date-time":"2026-01-26T10:24:36Z","timestamp":1769423076000},"page":"1220","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Untargeted Serum Proteomics in the Fontan Circulation Reveals Three Distinct Molecular Signatures of Fontan Physiology with CYB5R3 Among Key Proteins"],"prefix":"10.3390","volume":"27","author":[{"ORCID":"https:\/\/orcid.org\/0009-0000-6086-3308","authenticated-orcid":false,"given":"Alexander","family":"Blaha","sequence":"first","affiliation":[{"name":"Department of Child and Adolescent Health, Division of Pediatrics III\u2014Cardiology, Pulmonology, Allergology and Cystic Fibrosis, Medical University of Innsbruck, 6020 Innsbruck, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0003-8746-5758","authenticated-orcid":false,"given":"David","family":"Renaud","sequence":"additional","affiliation":[{"name":"Department of Child and Adolescent Health, Division of Pediatrics III\u2014Cardiology, Pulmonology, Allergology and Cystic Fibrosis, Medical University of Innsbruck, 6020 Innsbruck, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0008-8066-9658","authenticated-orcid":false,"given":"Fatima","family":"Ageed","sequence":"additional","affiliation":[{"name":"Department of Child and Adolescent Health, Division of Pediatrics III\u2014Cardiology, Pulmonology, Allergology and Cystic Fibrosis, Medical University of Innsbruck, 6020 Innsbruck, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bettina","family":"Sarg","sequence":"additional","affiliation":[{"name":"Biocenter, Protein Core Facility, Medical University of Innsbruck, 6020 Innsbruck, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1646-3256","authenticated-orcid":false,"given":"Klaus","family":"Faserl","sequence":"additional","affiliation":[{"name":"Biocenter, Protein Core Facility, Medical University of Innsbruck, 6020 Innsbruck, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2284-7424","authenticated-orcid":false,"given":"Alexander","family":"Kirchmair","sequence":"additional","affiliation":[{"name":"Biocenter, Bioinformatics Core Facility, Medical University of Innsbruck, 6020 Innsbruck, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1754-690X","authenticated-orcid":false,"given":"Dietmar","family":"Rieder","sequence":"additional","affiliation":[{"name":"Biocenter, Institute of Bioinformatics, Medical University of Innsbruck, 6020 Innsbruck, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Isabel","family":"Mihajlovic","sequence":"additional","affiliation":[{"name":"Department of Child and Adolescent Health, Division of Pediatrics III\u2014Cardiology, Pulmonology, Allergology and Cystic Fibrosis, Medical University of Innsbruck, 6020 Innsbruck, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nele","family":"Str\u00f6bel","sequence":"additional","affiliation":[{"name":"Department of Child and Adolescent Health, Division of Pediatrics III\u2014Cardiology, Pulmonology, Allergology and Cystic Fibrosis, Medical University of Innsbruck, 6020 Innsbruck, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kai Thorsten","family":"Laser","sequence":"additional","affiliation":[{"name":"Department of Pediatrics and Adolescent Medicine, University Hospital of Giessen and Marburg, 35392 Giessen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3626-3270","authenticated-orcid":false,"given":"Miriam","family":"Michel","sequence":"additional","affiliation":[{"name":"Department of Child and Adolescent Health, Division of Pediatrics III\u2014Cardiology, Pulmonology, Allergology and Cystic Fibrosis, Medical University of Innsbruck, 6020 Innsbruck, Austria"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,1,26]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1111","DOI":"10.1007\/s00392-011-0355-7","article-title":"Frequency and spectrum of congenital heart defects among live births in Germany: A study of the Competence Network for Congenital Heart Defects","volume":"100","author":"Schwedler","year":"2011","journal-title":"Clin. Res. Cardiol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"800","DOI":"10.1161\/CIRCULATIONAHA.105.592378","article-title":"Univentricular heart","volume":"115","author":"Khairy","year":"2007","journal-title":"Circulation"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"240","DOI":"10.1136\/thx.26.3.240","article-title":"Surgical repair of tricuspid atresia","volume":"26","author":"Fontan","year":"1971","journal-title":"Thorax"},{"key":"ref_4","first-page":"S32","article-title":"Redefining expectations of long-term survival after the Fontan procedure: Twenty-five years of follow-up from the entire population of Australia and New Zealand","volume":"130","author":"Iyengar","year":"2014","journal-title":"Circulation"},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Renaud, D., Scholl-B\u00fcrgi, S., Karall, D., and Michel, M. (2023). Comparative Metabolomics in Single Ventricle Patients after Fontan Palliation: A Strong Case for a Targeted Metabolic Therapy. Metabolites, 13.","DOI":"10.3390\/metabo13080932"},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Michel, M., Renaud, D., Schmidt, R., Einkemmer, M., Laser, L.V., Michel, E., Dubowy, K.O., Karall, D., Laser, K.T., and Scholl-B\u00fcrgi, S. (2024). Altered Serum Proteins Suggest Inflammation, Fibrogenesis and Angiogenesis in Adult Patients with a Fontan Circulation. Int. J. Mol. Sci., 25.","DOI":"10.3390\/ijms25105416"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"e038648","DOI":"10.1161\/JAHA.124.038648","article-title":"Systemic Changes in Adults with a Fontan Circulation: Insights from the Plasma Proteome","volume":"14","author":"Kelly","year":"2025","journal-title":"J. Am. Heart Assoc."},{"key":"ref_8","first-page":"100584","article-title":"Correlation between Olink and SomaScan proteomics platforms in adults with a Fontan circulation","volume":"20","author":"Assi","year":"2025","journal-title":"Int. J. Cardiol. Congenit. Heart Dis."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"e012136","DOI":"10.1161\/CIRCHEARTFAILURE.124.012136","article-title":"Plasma Proteomics of the Fontan Circulation Reveal Signatures of Oxidative Stress and Cell Death","volume":"18","author":"Lecointe","year":"2025","journal-title":"Circ. Heart Fail."},{"key":"ref_10","unstructured":"UniPro (2026, January 18). Uniprot. Available online: https:\/\/www.uniprot.org."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Thomson, L.M., Mancuso, C.A., Wolfe, K.R., Khailova, L., Niemiec, S., Ali, E., DiMaria, M., Mitchell, M., Twite, M., and Morgan, G. (2023). The proteomic fingerprint in infants with single ventricle heart disease in the interstage period: Evidence of chronic inflammation and widespread activation of biological networks. Front. Pediatr., 11.","DOI":"10.3389\/fped.2023.1308700"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1920","DOI":"10.1164\/ajrccm.158.6.9804076","article-title":"Alveolar hypoxia increases gene expression of extracellular matrix proteins and platelet-derived growth factor-B in lung parenchyma","volume":"158","author":"Berg","year":"1998","journal-title":"Am. J. Respir. Crit. Care Med."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1087","DOI":"10.1038\/s41598-024-84151-3","article-title":"Regulation of fibronectin and collagens type I, III and VI by TNF-\u03b1, TGF-\u03b2, IL-13, and tofacitinib","volume":"15","author":"Gillesberg","year":"2025","journal-title":"Sci. Rep."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"56","DOI":"10.1016\/j.ijcard.2020.01.012","article-title":"Biomarker profile in stable Fontan patients","volume":"305","author":"Saraf","year":"2020","journal-title":"Int. J. Cardiol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"7809","DOI":"10.1074\/jbc.M510722200","article-title":"Heparan sulfate plays a central role in a dynamic in vitro model of protein-losing enteropathy","volume":"281","author":"Bode","year":"2006","journal-title":"J. Biol. Chem."},{"key":"ref_16","first-page":"988","article-title":"Plastic Bronchitis and Protein-Losing Enteropathy in the Fontan Patient: Evolving Understanding and Emerging Therapies","volume":"38","author":"Mackie","year":"2022","journal-title":"Can. J. Cardiol."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"777","DOI":"10.1161\/01.RES.0000015592.95986.03","article-title":"Hypoxia causes downregulation of protein and RNA synthesis in noncontracting Mammalian cardiomyocytes","volume":"90","author":"Casey","year":"2002","journal-title":"Circ. Res."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1335","DOI":"10.1038\/s41390-023-02969-5","article-title":"Decreased erythrocyte aggregation in Glenn and Fontan: Univentricular circulation as a rheologic disease model","volume":"95","author":"Suriany","year":"2024","journal-title":"Pediatr. Res."},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Meinel, K., Korak, F., Dusleag, M., Strini, T., Baumgartner, D., Burmas, A., Sallmon, H., Zieger, B., Schlagenhauf, A., and Koestenberger, M. (2023). Mild Acquired von Willebrand Syndrome and Cholestasis in Pediatric and Adult Patients with Fontan Circulation. J. Clin. Med., 12.","DOI":"10.3390\/jcm12031240"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"893","DOI":"10.1016\/j.cell.2006.02.031","article-title":"TGF-\u03b2 regulation by Emilin1: New links in the etiology of hypertension","volume":"124","author":"Raman","year":"2006","journal-title":"Cell"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"289","DOI":"10.1016\/S0945-053X(00)00074-3","article-title":"The EMILIN protein family","volume":"19","author":"Colombatti","year":"2000","journal-title":"Matrix Biol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"102654","DOI":"10.1016\/j.jbc.2022.102654","article-title":"Cytochrome b5 reductases: Redox regulators of cell homeostasis","volume":"298","author":"Hall","year":"2022","journal-title":"J. Biol. Chem."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"63","DOI":"10.18632\/aging.100112","article-title":"CYB5R3: A key player in aerobic metabolism and aging?","volume":"2","author":"Siendones","year":"2009","journal-title":"Aging"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"72","DOI":"10.1016\/j.yjmcc.2021.09.005","article-title":"Smooth muscle cell CYB5R3 preserves cardiac and vascular function under chronic hypoxic stress","volume":"162","author":"Durgin","year":"2022","journal-title":"J. Mol. Cell. Cardiol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"298","DOI":"10.1111\/j.1365-2141.2008.07017.x","article-title":"Recessive congenital methaemoglobinaemia: Cytochrome b5 reductase deficiency","volume":"141","author":"Percy","year":"2008","journal-title":"Br. J. Haematol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"384","DOI":"10.1016\/j.jjcc.2014.02.021","article-title":"Iron deficiency and hematological changes in adult patients after Fontan operation","volume":"64","author":"Plazak","year":"2014","journal-title":"J. Cardiol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"e002693","DOI":"10.1136\/openhrt-2024-002693","article-title":"Iron deficiency in patients with a Fontan circulation and its impact on exercise capacity","volume":"11","author":"Rivrud","year":"2024","journal-title":"Open Heart"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"8930","DOI":"10.1038\/s41598-020-65852-x","article-title":"Targeted metabolomic analysis of serum amino acids in the adult Fontan patient with a dominant left ventricle","volume":"10","author":"Michel","year":"2020","journal-title":"Sci. Rep."},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Surendran, A., Ravandi, A., and Shah, A.H. (2025). Plasma Lipidomic Alterations in Fontan Circulation Reflect Cardiovascular Functional Reserve. Metabolites, 15.","DOI":"10.3390\/metabo15090592"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"100865","DOI":"10.1016\/j.jacadv.2024.100865","article-title":"Accelerated Epigenetic Aging in Children and Adults with a Fontan Circulation","volume":"3","author":"Drury","year":"2024","journal-title":"JACC Adv."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"15941","DOI":"10.1021\/acs.analchem.9b04474","article-title":"TMTpro: Design, Synthesis, and Initial Evaluation of a Proline-Based Isobaric 16-Plex Tandem Mass Tag Reagent Set","volume":"91","author":"Thompson","year":"2019","journal-title":"Anal. Chem."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"14","DOI":"10.1016\/j.ab.2015.03.003","article-title":"Evaluating the effects of preanalytical variables on the stability of the human plasma proteome","volume":"478","author":"Hassis","year":"2015","journal-title":"Anal. Biochem."},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Kramer, P., Schleiger, A., Schafstedde, M., Danne, F., Nordmeyer, J., Berger, F., and Ovroutski, S. (2022). A Multimodal Score Accurately Classifies Fontan Failure and Late Mortality in Adult Fontan Patients. Front. Cardiovasc. Med., 9.","DOI":"10.3389\/fcvm.2022.767503"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"198","DOI":"10.1093\/ehjci\/jeu179","article-title":"Impact of respiration on stroke volumes in paediatric controls and in patients after Fontan procedure assessed by MR real-time phase-velocity mapping","volume":"16","author":"Barth","year":"2015","journal-title":"Eur. Heart J. Cardiovasc. Imaging"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"158","DOI":"10.1016\/j.ijcard.2015.05.174","article-title":"Peak oxygen uptake, ventilatory efficiency and QRS-duration predict event free survival in patients late after surgical repair of tetralogy of Fallot","volume":"196","author":"Hager","year":"2015","journal-title":"Int. J. Cardiol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"4610","DOI":"10.1021\/acs.jproteome.1c00378","article-title":"Evaluation of Spin Columns for Human Plasma Depletion to Facilitate MS-Based Proteomics Analysis of Plasma","volume":"20","author":"Cao","year":"2021","journal-title":"J. Proteome Res."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"464777","DOI":"10.1016\/j.chroma.2024.464777","article-title":"High throughput multidimensional liquid chromatography approach for online protein removal and characterization of polysorbates and poloxamer in monoclonal antibody formulations","volume":"1720","author":"Zegota","year":"2024","journal-title":"J. Chromatogr. A"},{"key":"ref_38","doi-asserted-by":"crossref","unstructured":"Indelicato, E., Faserl, K., Amprosi, M., Nachbauer, W., Schneider, R., Wanschitz, J., Sarg, B., and Boesch, S. (2023). Skeletal muscle proteome analysis underpins multifaceted mitochondrial dysfunction in Friedreich\u2019s ataxia. Front. Neurosci., 17.","DOI":"10.3389\/fnins.2023.1289027"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"289","DOI":"10.1111\/j.2517-6161.1995.tb02031.x","article-title":"Controlling the False Discovery Rate: A Practical and Powerful Approach to Multiple Testing","volume":"57","author":"Benjamini","year":"1995","journal-title":"J. R. Stat. Soc. Ser. B Methodol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"15545","DOI":"10.1073\/pnas.0506580102","article-title":"Gene set enrichment analysis: A knowledge-based approach for interpreting genome-wide expression profiles","volume":"102","author":"Subramanian","year":"2005","journal-title":"Proc. Natl. Acad. Sci. USA"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/27\/3\/1220\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,1,26]],"date-time":"2026-01-26T10:25:09Z","timestamp":1769423109000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/27\/3\/1220"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,1,26]]},"references-count":40,"journal-issue":{"issue":"3","published-online":{"date-parts":[[2026,2]]}},"alternative-id":["ijms27031220"],"URL":"https:\/\/doi.org\/10.3390\/ijms27031220","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,1,26]]}}}