{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T12:13:31Z","timestamp":1785413611291,"version":"3.56.0"},"reference-count":27,"publisher":"MDPI AG","issue":"4","license":[{"start":{"date-parts":[[2026,4,17]],"date-time":"2026-04-17T00:00:00Z","timestamp":1776384000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001691","name":"Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["21K19539"],"award-info":[{"award-number":["21K19539"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"award":["21K19539"],"award-info":[{"award-number":["21K19539"]}],"id":[{"id":"https:\/\/ror.org\/00hhkn466","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Metabolites"],"abstract":"<jats:p>Background: Acute rejection remains a major complication following liver transplantation, yet reliable noninvasive biomarkers for its early prediction and diagnosis remain unidentified. This exploratory study characterized bile and serum metabolites associated with acute rejection in living donor liver transplantation using comprehensive metabolomic profiling combined with machine learning. Methods: Non-targeted metabolomics were performed on bile samples collected on post-operative day (POD) 1 (n = 38) and serum on POD 14 (n = 45) from liver transplant recipients. Partial least squares discriminant analysis-based variable selection was followed by logistic regression and least absolute shrinkage and selection operator models, which were evaluated via cross-validation in the discovery cohort to explore potential biomarkers for acute rejection. Results: A three-variable, bile-based model for predicting acute rejection achieved a mean cross-validated AUC of 0.872 (95% confidence interval: 0.814\u20130.930). Glycohyocholic acid and sulfolithocholylglycine were the main contributors. A nine-variable serum model for the Rejection Activity Index, including the change in \u03b3-glutamyl transferase, showed a mean cross-validated R2 of 0.728 (95% confidence interval: 0.609\u20130.846), with methionine, creatine, and oxidized fatty acids contributing prominently. Conclusions: These findings suggest that metabolomic profiling combined with machine learning may provide candidate biomarkers for acute rejection after liver transplantation. However, given the exploratory nature of the study and the lack of external validation, the clinical utility of these metabolite signatures remains to be determined. Therefore, external validation in larger, independent cohorts will be required.<\/jats:p>","DOI":"10.3390\/metabo16040273","type":"journal-article","created":{"date-parts":[[2026,4,17]],"date-time":"2026-04-17T14:24:51Z","timestamp":1776435891000},"page":"273","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Bile and Serum Metabolomics in Living Donor Liver Transplantation: Exploratory Insights into Acute Rejection Biomarkers"],"prefix":"10.3390","volume":"16","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2728-2648","authenticated-orcid":false,"given":"Yuta","family":"Hirata","sequence":"first","affiliation":[{"name":"Department of Surgery, Division of Gastroenterological, General and Transplant Surgery, Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0961-9622","authenticated-orcid":false,"given":"Yasunaru","family":"Sakuma","sequence":"additional","affiliation":[{"name":"Department of Surgery, Division of Gastroenterological, General and Transplant Surgery, Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8241-3006","authenticated-orcid":false,"given":"Hideo","family":"Ogiso","sequence":"additional","affiliation":[{"name":"Department of Translational Research, Clinical Research Center, Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3681-7736","authenticated-orcid":false,"given":"Taiichi","family":"Wakiya","sequence":"additional","affiliation":[{"name":"Department of Surgery, Division of Gastroenterological, General and Transplant Surgery, Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5211-5983","authenticated-orcid":false,"given":"Takahiko","family":"Omameuda","sequence":"additional","affiliation":[{"name":"Department of Surgery, Division of Gastroenterological, General and Transplant Surgery, Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2125-7513","authenticated-orcid":false,"given":"Toshio","family":"Horiuchi","sequence":"additional","affiliation":[{"name":"Department of Surgery, Division of Gastroenterological, General and Transplant Surgery, Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Noriki","family":"Okada","sequence":"additional","affiliation":[{"name":"Department of Surgery, Division of Gastroenterological, General and Transplant Surgery, Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yukihiro","family":"Sanada","sequence":"additional","affiliation":[{"name":"Department of Surgery, Division of Gastroenterological, General and Transplant Surgery, Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yasuharu","family":"Onishi","sequence":"additional","affiliation":[{"name":"Department of Surgery, Division of Gastroenterological, General and Transplant Surgery, Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8371-9410","authenticated-orcid":false,"given":"Hironori","family":"Yamaguchi","sequence":"additional","affiliation":[{"name":"Department of Surgery, Division of Gastroenterological, General and Transplant Surgery, Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ryozo","family":"Nagai","sequence":"additional","affiliation":[{"name":"Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kenichi","family":"Aizawa","sequence":"additional","affiliation":[{"name":"Department of Translational Research, Clinical Research Center, Jichi Medical University, Tochigi 329-0498, Shimotsuke, Japan"},{"name":"Clinical Pharmacology Center, Jichi Medical University Hospital, Tochigi 329-0498, Shimotsuke, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,4,17]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"358","DOI":"10.1016\/j.jceh.2017.10.003","article-title":"Acute and Chronic Rejection After Liver Transplantation: What A Clinician Needs to Know","volume":"7","author":"Choudhary","year":"2017","journal-title":"J. Clin. Exp. Hepatol."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Ozturk, N.B., Bartosek, N., Toruner, M.D., Mumtaz, A., Simsek, C., Dao, D., Saberi, B., and Gurakar, A. (2024). Approach to Liver Transplantation: Is There a Difference between East and West?. J. Clin. Med., 13.","DOI":"10.3390\/jcm13071890"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1061","DOI":"10.3748\/wjg.v21.i4.1061","article-title":"Markers of acute rejection and graft acceptance in liver transplantation","volume":"21","author":"Germani","year":"2015","journal-title":"World J. Gastroenterol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"139","DOI":"10.1016\/j.jceh.2022.06.010","article-title":"Current Status of Biomarkers and Molecular Diagnostic Tools for Rejection in Liver Transplantation: Light at the End of the Tunnel?","volume":"13","author":"Mohamed","year":"2023","journal-title":"J. Clin. Exp. Hepatol."},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Morsiani, C., Collura, S., Sevini, F., Ciurca, E., Bertuzzo, V.R., Franceschi, C., Grazi, G.L., Cescon, M., and Capri, M. (2023). Circulating miR-122-5p, miR-92a-3p, and miR-18a-5p as Potential Biomarkers in Human Liver Transplantation Follow-Up. Int. J. Mol. Sci., 24.","DOI":"10.3390\/ijms24043457"},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Garc\u00eda-Fern\u00e1ndez, N., Macher, H.C., Su\u00e1rez-Artacho, G., G\u00f3mez-Bravo, M.\u00c1., Molinero, P., Guerrero, J.M., Porras-L\u00f3pez, M., and Rubio, A. (2023). Donor-Specific Cell-Free DNA qPCR Quantification as a Noninvasive Accurate Biomarker for Early Rejection Detection in Liver Transplantation. J. Clin. Med., 12.","DOI":"10.3390\/jcm12010036"},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Wang, W., Li, W., Cao, L., Wang, B., Liu, C., Qin, Y., Guo, B., and Huang, C. (2022). Serum extracellular vesicle MicroRNAs as candidate biomarkers for acute rejection in patients subjected to liver transplant. Front. Genet., 13.","DOI":"10.3389\/fgene.2022.1015049"},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Budkowska, M., Ostrycharz, E., Serwin, N.M., Nazarewski, \u0141., Rykowski, P., Pietrzak, R., and Zieniewicz, K. (2023). Biomarkers of the Complement SystemActivation (C3a, C5a, sC5b-9) in Serum of Patients before and after Liver Transplantation. Biomedicines, 11.","DOI":"10.3390\/biomedicines11072070"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"18663","DOI":"10.1038\/s41598-022-18957-4","article-title":"Metabolomics profiling in acute liver transplant rejection in a pediatric population","volume":"12","author":"Frediani","year":"2022","journal-title":"Sci. Rep."},{"key":"ref_10","first-page":"372","article-title":"A panel of biomarkers in the prediction for early allograft dysfunction and mortality after living donor liver transplantation","volume":"13","author":"Tsai","year":"2021","journal-title":"Am. J. Transl. Res."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"286","DOI":"10.1111\/j.1432-2277.2004.tb00445.x","article-title":"Bile acids analysis: A tool to assess graft function in human liver transplantation","volume":"17","author":"Melendez","year":"2004","journal-title":"Transpl. Int."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"2063","DOI":"10.1002\/elps.201000512","article-title":"Bile UPLC-MS fingerprinting and bile acid fluxes during human liver transplantation","volume":"32","author":"McDermott","year":"2011","journal-title":"Electrophoresis"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"313","DOI":"10.1007\/978-1-0716-0239-3_16","article-title":"Predictive Modeling for Metabolomics Data","volume":"2104","author":"Ghosh","year":"2020","journal-title":"Methods Mol. Biol."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Barberis, E., Khoso, S., Sica, A., Falasca, M., Gennari, A., Dondero, F., Afantitis, A., and Manfredi, M. (2022). Precision Medicine Approaches with Metabolomics and Artificial Intelligence. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms231911269"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"810","DOI":"10.1016\/j.transproceed.2012.01.008","article-title":"Veno-occlusive disease\/sinusoidal obstruction syndrome associated with potential antibody-mediated rejection after pediatric living donor liver transplantation: A case report","volume":"44","author":"Yamada","year":"2012","journal-title":"Transplant. Proc."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1290","DOI":"10.1097\/00007890-200211150-00016","article-title":"Real-time monitoring of acute liver-allograft rejection using the banff schema","volume":"74","author":"Demetris","year":"2002","journal-title":"Transplantation"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"658","DOI":"10.1002\/hep.510250328","article-title":"Banff schema for grading liver allograft rejection: An international consensus document","volume":"25","author":"Demetris","year":"1997","journal-title":"Hepatology"},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Sonoda, H., Ogiso, H., Aoki, Y., Morishima, K., Sasanuma, H., Sata, N., Kitayama, J., Yamashita, H., Yamaguchi, H., and Nagai, R. (2026). Diagnostic Biomarkers for Pancreatic Ductal Adenocarcinoma Using Non-Targeted Metabolomic Analysis. Cancers, 18.","DOI":"10.3390\/cancers18040684"},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Ogiso, H., Miura, K., Nagai, R., Osaka, H., and Aizawa, K. (2024). Non-Targeted Metabolomics Reveal Apomorphine \u2019 s Therapeutic Effects and Lysophospholipid Alterations in Steatohepatitis. Antioxidants, 13.","DOI":"10.3390\/antiox13111293"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"7946","DOI":"10.1021\/acs.analchem.6b00770","article-title":"Hydrogen Rearrangement Rules: Computational MS\/MS Fragmentation and Structure Elucidation Using MS-FINDER Software","volume":"88","author":"Tsugawa","year":"2016","journal-title":"Anal. Chem."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"523","DOI":"10.1038\/nmeth.3393","article-title":"MS-DIAL: Data-independent MS\/MS deconvolution for comprehensive metabolome analysis","volume":"12","author":"Tsugawa","year":"2015","journal-title":"Nat. Methods"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"500","DOI":"10.1111\/j.1600-6143.2010.03414.x","article-title":"The significance of donor-specific HLA antibodies in rejection and ductopenia development in ABO-compatible liver transplantation","volume":"11","author":"Musat","year":"2011","journal-title":"Am. J. Transplant."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"641","DOI":"10.3390\/microorganisms3040641","article-title":"The gut microbiota as a therapeutic target in ibd and metabolic disease: A role for the bile acid receptors fxr and tgr5","volume":"3","author":"Baars","year":"2015","journal-title":"Microorganisms"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"G554","DOI":"10.1152\/ajpgi.00223.2019","article-title":"Bile acid receptors FXR and TGR5 signaling in fatty liver diseases and therapy","volume":"318","author":"Chiang","year":"2020","journal-title":"Am. J. Physiol.-Gastrointest. Liver Physiol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"117","DOI":"10.1007\/BF02774550","article-title":"Impaired metabolism of methionine in severe liver diseases I. Clinical and pathophysiological significance of elevated serum methionine levels","volume":"17","author":"Higashi","year":"1982","journal-title":"Gastroenterol. Jpn."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"2805","DOI":"10.1002\/jum.15968","article-title":"Gamma-Glutamyl Transferase Combined With Conventional Ultrasound Features in Diag-nosing Biliary Atresia: A Two-Center Retrospective Analysis","volume":"41","author":"Weng","year":"2022","journal-title":"J. Ultrasound Med."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"702","DOI":"10.1136\/gut.13.9.702","article-title":"Serum-glytamyl transpeptidase activity in liver disease","volume":"13","author":"Whitfield","year":"1972","journal-title":"Gut"}],"container-title":["Metabolites"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2218-1989\/16\/4\/273\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,4,25]],"date-time":"2026-04-25T04:16:49Z","timestamp":1777090609000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2218-1989\/16\/4\/273"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,4,17]]},"references-count":27,"journal-issue":{"issue":"4","published-online":{"date-parts":[[2026,4]]}},"alternative-id":["metabo16040273"],"URL":"https:\/\/doi.org\/10.3390\/metabo16040273","relation":{},"ISSN":["2218-1989"],"issn-type":[{"value":"2218-1989","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,4,17]]}}}