{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,30]],"date-time":"2026-06-30T11:45:35Z","timestamp":1782819935020,"version":"3.54.5"},"reference-count":44,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2026,6,25]],"date-time":"2026-06-25T00:00:00Z","timestamp":1782345600000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2026,6,30]],"date-time":"2026-06-30T00:00:00Z","timestamp":1782777600000},"content-version":"vor","delay-in-days":5,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"funder":[{"DOI":"10.13039\/501100012320","name":"Otto-von-Guericke-Universit\u00e4t Magdeburg","doi-asserted-by":"crossref","id":[{"id":"10.13039\/501100012320","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Sci Rep"],"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>\n                    MicroRNAs (miRNA) are considered promising biomarkers for inflammatory and neoplastic diseases in fecal samples, but despite reproducible miRNA detection, there is no consensus on appropriate normalization in fecal samples. In this study, we aimed to explore an endogenous normalizer for miRNA analysis in fecal samples. For this purpose, we performed a multistep study with 3 independent cohorts. In the first step, we investigated miRNA profiling using serial dilutions of blood and fecal samples from healthy subjects. In the second part, we tested the analysis using a cohort of IBD patients (\n                    <jats:italic>n<\/jats:italic>\n                    \u2009=\u200930) with active disease and in remission and 59 patients with different liver pathologies or healthy subjects. Finally, the data were validated using an independent cohort of patients with liver disease (\n                    <jats:italic>n<\/jats:italic>\n                    \u2009=\u2009360). After extraction, miRNAs were analyzed using Affymetrix GeneChip microarray technology in the screening and confirmation cohorts, and TaqMan qPCR was performed for validation. Analysis of the blood stool mixture cohort revealed 5 miRNAs that were stable in all three subgroups. Among them, miR-638 was found to have a Log\n                    <jats:sub>2<\/jats:sub>\n                    FC of 0.0004 and was selected for further analysis as one of the best studied miRNAs from previous reports. The stability of miR-638 in microarray analysis was demonstrated in two independent cohorts of patients with IBD and liver disease. We observed no effect of age or other factors on miR-638 levels in fecal samples. Furthermore, its relatively high concentration in feces and its stability against potential blood contamination may offer great advantages over the previously used miR-16, suggesting miR-638 as a potential endogenous fecal normalizer.\n                  <\/jats:p>","DOI":"10.1038\/s41598-026-57583-2","type":"journal-article","created":{"date-parts":[[2026,6,25]],"date-time":"2026-06-25T04:55:57Z","timestamp":1782363357000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Systematic assessment for potential endogenous normalizers for microRNA analysis in fecal samples"],"prefix":"10.1038","volume":"16","author":[{"given":"Theresa","family":"Lederer","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Konrad","family":"Lehr","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Cosima","family":"Thon","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Katharina","family":"Magin","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andreas","family":"Jeron","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Denny","family":"Schanze","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Martin","family":"Zenker","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ali","family":"Canbay","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alexander","family":"Link","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2026,6,25]]},"reference":[{"key":"57583_CR1","doi-asserted-by":"publisher","first-page":"1766","DOI":"10.1158\/1055-9965.EPI-10-0027","volume":"19","author":"A Link","year":"2010","unstructured":"Link, A. et al. Fecal microRNAs as novel biomarkers for colon cancer screening. Cancer Epidemiol. Biomarkers Prev. 19, 1766\u20131774 (2010).","journal-title":"Cancer Epidemiol. Biomarkers Prev."},{"key":"57583_CR2","doi-asserted-by":"publisher","first-page":"3313","DOI":"10.3748\/wjg.v24.i30.3313","volume":"24","author":"A Link","year":"2018","unstructured":"Link, A. & Kupcinskas, J. MicroRNAs as non-invasive diagnostic biomarkers for gastric cancer: Current insights and future perspectives. World J. Gastroenterol. 24, 3313\u20133329 (2018).","journal-title":"World J. Gastroenterol."},{"key":"57583_CR3","doi-asserted-by":"crossref","unstructured":"Link, A., Kupcinskas, J., Wex, T. & Malfertheiner, P. Macro-role of MicroRNA in gastric cancer. 30, 255\u2013267 (2012).","DOI":"10.1159\/000336919"},{"key":"57583_CR4","doi-asserted-by":"crossref","unstructured":"Link, A., Becker, V., Goel, A., Wex, T. & Malfertheiner, P. Feasibility of fecal microRNAs as novel biomarkers for pancreatic cancer. PLoS One 7, (2012).","DOI":"10.1371\/journal.pone.0042933"},{"key":"57583_CR5","doi-asserted-by":"crossref","unstructured":"Giza, D. E. & Calin, G. A. MicroRNA and chronic lymphocytic leukemia. in Advances in Experimental Medicine and Biology vol. 889 23\u201340Springer New York LLC, (2015).","DOI":"10.1007\/978-3-319-23730-5_2"},{"key":"57583_CR6","doi-asserted-by":"publisher","first-page":"522","DOI":"10.1038\/nrg1379","volume":"5","author":"L He","year":"2004","unstructured":"He, L., Hannon, G. J. & MicroRNAs Small RNAs with a big role in gene regulation. Nat. Rev. Genet. 5, 522\u2013531 (2004).","journal-title":"Nat. Rev. Genet."},{"key":"57583_CR7","doi-asserted-by":"publisher","first-page":"S52","DOI":"10.1038\/onc.2009.353","volume":"27","author":"P Trang","year":"2008","unstructured":"Trang, P., Weidhaas, J. B. & Slack, F. J. MicroRNAs as potential cancer therapeutics. Oncogene 27, S52\u2013S57 (2008).","journal-title":"Oncogene"},{"key":"57583_CR8","doi-asserted-by":"publisher","first-page":"136","DOI":"10.12688\/f1000research.2-136.v1","volume":"2","author":"P Hydbring","year":"2013","unstructured":"Hydbring, P. & Badalian-Very, G. Clinical applications of microRNAs. F1000Res 2, 136 (2013).","journal-title":"F1000Res"},{"key":"57583_CR9","doi-asserted-by":"publisher","first-page":"358","DOI":"10.1038\/nrg3198","volume":"13","author":"CC Pritchard","year":"2012","unstructured":"Pritchard, C. C., Cheng, H. H. & Tewari, M. MicroRNA profiling: Approaches and considerations. Nat. Rev. Genet. 13, 358\u2013369 (2012).","journal-title":"Nat. Rev. Genet."},{"key":"57583_CR10","doi-asserted-by":"publisher","first-page":"113","DOI":"10.1016\/bs.acc.2018.07.003","volume":"87","author":"M Faraldi","year":"2018","unstructured":"Faraldi, M., Gomarasca, M., Banfi, G. & Lombardi, G. Free Circulating miRNAs Measurement in Clinical Settings: The Still Unsolved Issue of the Normalization. Adv. Clin. Chem. 87, 113\u2013139 (2018).","journal-title":"Adv. Clin. Chem."},{"key":"57583_CR11","doi-asserted-by":"publisher","first-page":"843","DOI":"10.1016\/0092-8674(93)90529-Y","volume":"75","author":"RC Lee","year":"1993","unstructured":"Lee, R. C., Feinbaum, R. L. & Ambrost, V. The C. elegans Heterochronic Gene lin-4 Encodes Small RNAs with Antisense Complementarity to &II-14. Cell 75, 843\u2013854 (1993).","journal-title":"Cell"},{"key":"57583_CR12","doi-asserted-by":"publisher","first-page":"1030","DOI":"10.1016\/S0140-6736(10)60353-4","volume":"375","author":"D Cunningham","year":"2010","unstructured":"Cunningham, D. et al. Colorectal cancer. Lancet 375, 1030\u20131047 (2010).","journal-title":"Lancet"},{"key":"57583_CR13","doi-asserted-by":"publisher","first-page":"1590","DOI":"10.1016\/S0140-6736(12)60026-9","volume":"380","author":"DC Baumgart","year":"2012","unstructured":"Baumgart, D. C. & Sandborn, W. J. Crohn\u2019s disease. Lancet 380, 1590\u20131605 (2012).","journal-title":"Lancet"},{"key":"57583_CR14","doi-asserted-by":"publisher","first-page":"1756","DOI":"10.1016\/S0140-6736(16)32126-2","volume":"389","author":"R Ungaro","year":"2017","unstructured":"Ungaro, R., Mehandru, S., Allen, P. B., Peyrin-Biroulet, L. & Colombel, J. F. Ulcerative colitis. Lancet 389, 1756\u20131770 (2017).","journal-title":"Lancet"},{"key":"57583_CR15","first-page":"1","volume":"21","author":"H Wang","year":"2020","unstructured":"Wang, H. MicroRNAs and apoptosis in colorectal cancer. Int. J. Mol. Sci. 21, 1\u201315 (2020).","journal-title":"Int. J. Mol. Sci."},{"key":"57583_CR16","doi-asserted-by":"publisher","first-page":"2112","DOI":"10.7150\/ijbs.59904","volume":"17","author":"H Jung","year":"2021","unstructured":"Jung, H. et al. Roles of micrornas in inflammatory bowel disease. Int. J. Biol. Sci. 17, 2112\u20132123 (2021).","journal-title":"Int. J. Biol. Sci."},{"key":"57583_CR17","doi-asserted-by":"publisher","first-page":"159","DOI":"10.1093\/ibd\/izx009","volume":"24","author":"U Netz","year":"2018","unstructured":"Netz, U. et al. Plasma microRNA Profile Differentiates Crohn\u2019s Colitis from Ulcerative Colitis. Inflamm. Bowel Dis. 24, 159\u2013165 (2018).","journal-title":"Inflamm. Bowel Dis."},{"key":"57583_CR18","doi-asserted-by":"publisher","first-page":"1856","DOI":"10.1002\/med.21893","volume":"42","author":"D Liu","year":"2022","unstructured":"Liu, D., Saikam, V., Skrada, K. A., Merlin, D. & Iyer, S. S. Inflammatory bowel disease biomarkers. Med. Res. Rev. 42, 1856\u20131887 (2022).","journal-title":"Med. Res. Rev."},{"key":"57583_CR19","doi-asserted-by":"crossref","unstructured":"Francavilla, A. et al. Faecal miRNA profiles associated with age, sex, BMI, and lifestyle habits in healthy individuals. Sci Rep 11, (2021).","DOI":"10.1038\/s41598-021-00014-1"},{"key":"57583_CR20","doi-asserted-by":"publisher","first-page":"4104","DOI":"10.3748\/wjg.v24.i36.4104","volume":"24","author":"JL Torres","year":"2018","unstructured":"Torres, J. L. et al. Role of microRNAs in alcohol-induced liver disorders and non-alcoholic fatty liver disease. World J. Gastroenterol. 24, 4104\u20134118 (2018).","journal-title":"World J. Gastroenterol."},{"key":"57583_CR21","doi-asserted-by":"crossref","unstructured":"Newman, L. A. et al. Selective Isolation of Liver-Derived Extracellular Vesicles Redefines Performance of miRNA Biomarkers for Non-Alcoholic Fatty Liver Disease. Biomedicines 10, (2022).","DOI":"10.3390\/biomedicines10010195"},{"key":"57583_CR22","doi-asserted-by":"crossref","unstructured":"Klieser, E. et al. The crosstalk of miRNA and oxidative stress in the liver: From physiology to pathology and clinical implications. Int J. Mol. Sci 20, (2019).","DOI":"10.3390\/ijms20215266"},{"key":"57583_CR23","doi-asserted-by":"publisher","first-page":"2078","DOI":"10.1002\/jcp.30699","volume":"237","author":"A Mahmoudi","year":"2022","unstructured":"Mahmoudi, A., Butler, A. E., Jamialahmadi, T. & Sahebkar, A. The role of exosomal miRNA in nonalcoholic fatty liver disease. J. Cell. Physiol. 237, 2078\u20132094 (2022).","journal-title":"J. Cell. Physiol."},{"key":"57583_CR24","doi-asserted-by":"publisher","first-page":"1","DOI":"10.3390\/ijms21197368","volume":"21","author":"M Gholizadeh","year":"2020","unstructured":"Gholizadeh, M. et al. Identifying differentially expressed micrornas, target genes, and key pathways deregulated in patients with liver diseases. Int. J. Mol. Sci. 21, 1\u201316 (2020).","journal-title":"Int. J. Mol. Sci."},{"key":"57583_CR25","doi-asserted-by":"publisher","first-page":"217","DOI":"10.1055\/s-2004-832936","volume":"24","author":"S Mandayam","year":"2004","unstructured":"Mandayam, S., Mazen Jamal, M. & Morgan, T. R. Epidemiology of Alcoholic Liver Disease. Semin Liver Dis. 24, 217\u2013232 (2004).","journal-title":"Semin Liver Dis."},{"key":"57583_CR26","doi-asserted-by":"publisher","first-page":"846","DOI":"10.4254\/wjh.v7.i6.846","volume":"7","author":"SM Abd El-Kader","year":"2015","unstructured":"Abd El-Kader, S. M. & El-Den Ashmawy, E. M. Non-alcoholic fatty liver disease: The diagnosis and management. World J. Hepatol. 7, 846\u2013858 (2015).","journal-title":"World J. Hepatol."},{"key":"57583_CR27","doi-asserted-by":"publisher","first-page":"1021","DOI":"10.4065\/76.10.1021","volume":"76","author":"KVN Menon","year":"2001","unstructured":"Menon, K. V. N., Gores, G. J. & Shah, V. H. Pathogenesis, diagnosis, and treatment of alcoholic liver disease. Mayo Clin. Proc. 76, 1021\u20131029 (2001).","journal-title":"Mayo Clin. Proc."},{"key":"57583_CR28","doi-asserted-by":"publisher","first-page":"2065","DOI":"10.1136\/gutjnl-2018-318146","volume":"68","author":"M Gjorgjieva","year":"2019","unstructured":"Gjorgjieva, M., Sobolewski, C., Dolicka, D., Correia De Sousa, M. & Foti, M. MiRNAs and NAFLD: From pathophysiology to therapy. Gut 68, 2065\u20132079 (2019).","journal-title":"Gut"},{"key":"57583_CR29","doi-asserted-by":"publisher","first-page":"e25441","DOI":"10.1097\/MD.0000000000025441","volume":"100","author":"L Hu","year":"2021","unstructured":"Hu, L., Huang, M., Yuan, Q. & Kong, F. Prognostic and clinicopathological significance of miR-638 in cancer patients: A meta-analysis. Medicine 100, e25441 (2021).","journal-title":"Medicine"},{"key":"57583_CR30","unstructured":"Xu, Y. et al. Long Noncoding RNA NR2F1-AS1 Enhances the Migration and Invasion of Hepatocellular Carcinoma via Modulating miR-642a\/DEK Pathway. J. Oncol. (2021). (2021)."},{"key":"57583_CR31","doi-asserted-by":"crossref","unstructured":"Yu, Z. et al. miR-642 serves as a tumor suppressor in hepatocellular carcinoma by regulating sema4c and p38 mapk signaling pathway. Oncol Lett 20, (2020).","DOI":"10.3892\/ol.2020.11935"},{"key":"57583_CR32","doi-asserted-by":"publisher","first-page":"6475","DOI":"10.1007\/s11033-022-07476-0","volume":"49","author":"M Fathi","year":"2022","unstructured":"Fathi, M., Aghdaie, H. A., Ghafouri-Fard, S. & Shams, R. Evaluation of potential of miR-8073 and miR-642 as diagnostic markers in pancreatic cancer. Mol. Biol. Rep. 49, 6475\u20136481 (2022).","journal-title":"Mol. Biol. Rep."},{"key":"57583_CR33","doi-asserted-by":"crossref","unstructured":"Liu, B. X., Yang, J., Zeng, C. & Chen, Y. MACC1 Correlates with Tumor Progression and Immune Cell Infiltration of Colon Adenocarcinoma and is Regulated by the lncRNA ZFAS1\/miR-642a-5p Axis. J. Oncol. (2022). (2022).","DOI":"10.1155\/2022\/8179208"},{"key":"57583_CR34","doi-asserted-by":"publisher","first-page":"4802","DOI":"10.1002\/jcb.28174","volume":"120","author":"S Madadi","year":"2019","unstructured":"Madadi, S. & Soleimani, M. Comparison of miR-16 and cel-miR-39 as reference controls for serum miRNA normalization in colorectal cancer. J. Cell. Biochem. 120, 4802\u20134803 (2019).","journal-title":"J. Cell. Biochem."},{"key":"57583_CR35","doi-asserted-by":"publisher","first-page":"e25441","DOI":"10.1097\/MD.0000000000025441","volume":"100","author":"L Hu","year":"2021","unstructured":"Hu, L., Huang, M., Yuan, Q. & Kong, F. Prognostic and clinicopathological significance of miR-638 in cancer patients: A meta-analysis. Medicine 100, e25441 (2021).","journal-title":"Medicine"},{"key":"57583_CR36","doi-asserted-by":"publisher","first-page":"825","DOI":"10.1007\/s12640-022-00504-8","volume":"40","author":"L Deng","year":"2022","unstructured":"Deng, L., Gong, K. & Wang, G. Hsa_circ_0008344 Promotes Glioma Tumor Progression and Angiogenesis Presumably by Regulating miR-638\/SZRD1 Pathway. Neurotox. Res. 40, 825\u2013836 (2022).","journal-title":"Neurotox. Res."},{"key":"57583_CR37","doi-asserted-by":"crossref","unstructured":"Nascimento, N. P. G., Gally, T. B., Borges, G. F., Campos, L. C. G. & Kaneto, C. M. Systematic review of circulating MICRORNAS as biomarkers of cervical carcinogenesis. BMC Cancer 22, (2022).","DOI":"10.1186\/s12885-022-09936-z"},{"key":"57583_CR38","doi-asserted-by":"publisher","first-page":"2130","DOI":"10.1016\/j.asjsur.2022.11.054","volume":"46","author":"D Yan","year":"2022","unstructured":"Yan, D., Zhu, C. & Liu, W. Low expression of microRNA-638 is associated with poor overall survival from hepatocellular carcinoma: A meta-analysis. Asian J. Surg. 46, 2130\u20132131 (2022).","journal-title":"Asian J. Surg."},{"key":"57583_CR39","doi-asserted-by":"crossref","unstructured":"Alkandari, A. et al. Improved physiology and metabolic flux after Roux-en-Y gastric bypass is associated with temporal changes in the circulating microRNAome: A longitudinal study in humans. BMC Obes 5, (2018).","DOI":"10.1186\/s40608-018-0199-z"},{"key":"57583_CR40","unstructured":"Dobrowolny, G. et al. A longitudinal study defined circulating microRNAs as reliable biomarkers for disease prognosis and progression in ALS human patients. Cell Death Discov 7, (2021)."},{"key":"57583_CR41","doi-asserted-by":"publisher","first-page":"900","DOI":"10.1038\/s41370-021-00306-8","volume":"31","author":"AM Jung","year":"2021","unstructured":"Jung, A. M. et al. Longitudinal evaluation of whole blood miRNA expression in firefighters. J. Expo Sci. Environ. Epidemiol. 31, 900\u2013912 (2021).","journal-title":"J. Expo Sci. Environ. Epidemiol."},{"key":"57583_CR42","doi-asserted-by":"crossref","unstructured":"Link, J. et al. The Translational Impact of Plant-Derived Xeno-miRNA miR-168 in Gastrointestinal Cancers and Preneoplastic Conditions. Diagnostics 13, (2023).","DOI":"10.3390\/diagnostics13162701"},{"key":"57583_CR43","doi-asserted-by":"publisher","first-page":"498","DOI":"10.3390\/genes15040498","volume":"15","author":"T Lederer","year":"2024","unstructured":"Lederer, T., Hipler, N. M., Thon, C., Kupcinskas, J. & Link, A. Comparison of Fecal MicroRNA Isolation Using Various Total RNA Isolation Kits. Genes (Basel). 15, 498 (2024).","journal-title":"Genes (Basel)"},{"key":"57583_CR44","doi-asserted-by":"publisher","first-page":"7742","DOI":"10.1038\/s41598-026-41688-9","volume":"16","author":"T Lederer","year":"2026","unstructured":"Lederer, T. et al. Combining tissue-derived microRNAs with clinical risk models for prediction of HCC recurrence after liver transplantation: A proof-of-concept study. Sci. Rep. 16, 7742 (2026).","journal-title":"Sci. Rep."}],"container-title":["Scientific Reports"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.nature.com\/articles\/s41598-026-57583-2","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/www.nature.com\/articles\/s41598-026-57583-2.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/www.nature.com\/articles\/s41598-026-57583-2.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,6,30]],"date-time":"2026-06-30T10:54:31Z","timestamp":1782816871000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.nature.com\/articles\/s41598-026-57583-2"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,6,25]]},"references-count":44,"journal-issue":{"issue":"1","published-online":{"date-parts":[[2026,12]]}},"alternative-id":["57583"],"URL":"https:\/\/doi.org\/10.1038\/s41598-026-57583-2","relation":{},"ISSN":["2045-2322"],"issn-type":[{"value":"2045-2322","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,6,25]]},"assertion":[{"value":"9 April 2026","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"8 June 2026","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"25 June 2026","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Declarations"}},{"value":"Competing Interests Statement: Alexander Link is a Senior Editorial Board Member of Scientific Reports.","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Competing interests"}},{"value":"This prospective study was conducted in accordance with the World Medical association Declaration of Helsinki \u2013 \u201cEthical Principles for Medical Research Involving Human Subjects.\u201d The study protocol was approved by the Institutional Review Board of Otto von Guericke University Magdeburg (number 99\/10; 78\/19 and 65\/19). Each participant in the prospective part of the study provided written informed consent before enrollment.","order":3,"name":"Ethics","group":{"name":"EthicsHeading","label":"Ethical standards"}}],"article-number":"19925"}}