{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,26]],"date-time":"2026-09-26T13:50:31Z","timestamp":1790430631623,"version":"4.1.0"},"reference-count":39,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2022,2,4]],"date-time":"2022-02-04T00:00:00Z","timestamp":1643932800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001665","name":"Agence Nationale de la Recherche","doi-asserted-by":"publisher","award":["ANR-16-RHUS-0005"],"award-info":[{"award-number":["ANR-16-RHUS-0005"]}],"id":[{"id":"10.13039\/501100001665","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Cells"],"abstract":"<jats:p>Background: Human-induced pluripotent stem cell-derived hepatocytes (iHeps) have been shown to have considerable potential in liver diseases, toxicity, and pharmacological studies. However, there is a growing need to obtain iHeps that are truly similar to primary adult hepatocytes in terms of morphological features and functions. We generated such human iHeps, self-assembled as organoids (iHep-Orgs). Methods: iPSC-derived hepatoblasts were self-assembled into spheroids and differentiated into mature hepatocytes modulating final step of differentiation. Results: In about four weeks of culture, the albumin secretion levels and the complete disappearance of \u03b1-fetoprotein from iHep-Orgs suggested the acquisition of a greater degree of maturation than those previously reported. The expression of apical transporters and bile acid secretion evidenced the acquisition of complex hepatocyte polarity as well as the development of a functional and well-defined bile canalicular network confirmed by computational analysis. Activities recorded for CYP450, UGT1A1, and alcohol dehydrogenase, response to hormonal stimulation, and glucose metabolism were also remarkable. Finally, iHep-Orgs displayed a considerable ability to detoxify pathological concentrations of lactate and ammonia. Conclusions: With features similar to those of primary adult hepatocytes, the iHep-Orgs thus produced could be considered as a valuable tool for the development and optimization of preclinical and clinical applications.<\/jats:p>","DOI":"10.3390\/cells11030537","type":"journal-article","created":{"date-parts":[[2022,2,4]],"date-time":"2022-02-04T11:35:17Z","timestamp":1643974517000},"page":"537","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":26,"title":["Evidence of Adult Features and Functions of Hepatocytes Differentiated from Human Induced Pluripotent Stem Cells and Self-Organized as Organoids"],"prefix":"10.3390","volume":"11","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-5610-973X","authenticated-orcid":false,"given":"Antonietta","family":"Messina","sequence":"first","affiliation":[{"name":"UMR_S 1193, INSERM\/Universit\u00e9 Paris-Saclay, F-94800 Villejuif, France"},{"name":"Centre H\u00e9patobiliaire, F\u00e9d\u00e9ration Hospitalo-Universitaire (FHU) H\u00e9patinov, AP-HP, H\u00f4pital Paul Brousse, F-94800 Villejuif, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8938-8405","authenticated-orcid":false,"given":"El\u00e9anor","family":"Luce","sequence":"additional","affiliation":[{"name":"UMR_S 1193, INSERM\/Universit\u00e9 Paris-Saclay, F-94800 Villejuif, France"},{"name":"Centre H\u00e9patobiliaire, F\u00e9d\u00e9ration Hospitalo-Universitaire (FHU) H\u00e9patinov, AP-HP, H\u00f4pital Paul Brousse, F-94800 Villejuif, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7502-7441","authenticated-orcid":false,"given":"Nassima","family":"Benzoubir","sequence":"additional","affiliation":[{"name":"UMR_S 1193, INSERM\/Universit\u00e9 Paris-Saclay, F-94800 Villejuif, France"},{"name":"Centre H\u00e9patobiliaire, F\u00e9d\u00e9ration Hospitalo-Universitaire (FHU) H\u00e9patinov, AP-HP, H\u00f4pital Paul Brousse, F-94800 Villejuif, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8312-6041","authenticated-orcid":false,"given":"Mattia","family":"Pasqua","sequence":"additional","affiliation":[{"name":"Centre H\u00e9patobiliaire, F\u00e9d\u00e9ration Hospitalo-Universitaire (FHU) H\u00e9patinov, AP-HP, H\u00f4pital Paul Brousse, F-94800 Villejuif, France"},{"name":"UMR CNRS 7338 Biomechanics & Bioengineering, Universit\u00e9 de Technologie de Compi\u00e8gne, Sorbonne Universit\u00e9s, 60203 Compiegne, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5677-6906","authenticated-orcid":false,"given":"Ulysse","family":"Pereira","sequence":"additional","affiliation":[{"name":"Centre H\u00e9patobiliaire, F\u00e9d\u00e9ration Hospitalo-Universitaire (FHU) H\u00e9patinov, AP-HP, H\u00f4pital Paul Brousse, F-94800 Villejuif, France"},{"name":"UMR CNRS 7338 Biomechanics & Bioengineering, Universit\u00e9 de Technologie de Compi\u00e8gne, Sorbonne Universit\u00e9s, 60203 Compiegne, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lydie","family":"Humbert","sequence":"additional","affiliation":[{"name":"Centre de Recherche Saint-Antoine, Sorbonne Universit\u00e9, INSERM, CRSA, AP-HP, H\u00f4pital Saint Antoine, Metomics, 75012 Paris, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1891-9253","authenticated-orcid":false,"given":"Thibaut","family":"Eguether","sequence":"additional","affiliation":[{"name":"Centre de Recherche Saint-Antoine, Sorbonne Universit\u00e9, INSERM, CRSA, AP-HP, H\u00f4pital Saint Antoine, Metomics, 75012 Paris, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dominique","family":"Rainteau","sequence":"additional","affiliation":[{"name":"Centre de Recherche Saint-Antoine, Sorbonne Universit\u00e9, INSERM, CRSA, AP-HP, H\u00f4pital Saint Antoine, Metomics, 75012 Paris, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jean-Charles","family":"Duclos-Vall\u00e9e","sequence":"additional","affiliation":[{"name":"UMR_S 1193, INSERM\/Universit\u00e9 Paris-Saclay, F-94800 Villejuif, France"},{"name":"Centre H\u00e9patobiliaire, F\u00e9d\u00e9ration Hospitalo-Universitaire (FHU) H\u00e9patinov, AP-HP, H\u00f4pital Paul Brousse, F-94800 Villejuif, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8061-906X","authenticated-orcid":false,"given":"C\u00e9cile","family":"Legallais","sequence":"additional","affiliation":[{"name":"Centre H\u00e9patobiliaire, F\u00e9d\u00e9ration Hospitalo-Universitaire (FHU) H\u00e9patinov, AP-HP, H\u00f4pital Paul Brousse, F-94800 Villejuif, France"},{"name":"UMR CNRS 7338 Biomechanics & Bioengineering, Universit\u00e9 de Technologie de Compi\u00e8gne, Sorbonne Universit\u00e9s, 60203 Compiegne, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9326-4413","authenticated-orcid":false,"given":"Anne","family":"Dubart-Kupperschmitt","sequence":"additional","affiliation":[{"name":"UMR_S 1193, INSERM\/Universit\u00e9 Paris-Saclay, F-94800 Villejuif, France"},{"name":"Centre H\u00e9patobiliaire, F\u00e9d\u00e9ration Hospitalo-Universitaire (FHU) H\u00e9patinov, AP-HP, H\u00f4pital Paul Brousse, F-94800 Villejuif, France"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2022,2,4]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1113","DOI":"10.1016\/j.apsb.2019.10.001","article-title":"Current trends in drug metabolism and pharmaco-kinetics","volume":"9","author":"Li","year":"2019","journal-title":"Acta Pharm. Sin. B"},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Holmgren, G., Ulfenborg, B., Asplund, A., Toet, K., Andersson, C.X., Hammarstedt, A., Hanemaaijer, R., K\u00fcppers-Munther, B., and Synnergren, J. (2020). Characterization of Human Induced Pluripotent Stem Cell-Derived Hepatocytes with Mature Features and Potential for Modeling Metabolic Diseases. Int. J. Mol. Sci., 21.","DOI":"10.3390\/ijms21020469"},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"Messina, A., Luce, E., Hussein, M., and Dubart-Kupperschmitt, A. (2020). Pluripotent-Stem-Cell-Derived Hepatic Cells: Hepatocytes and Organoids for Liver Therapy and Regeneration. Cells, 9.","DOI":"10.3390\/cells9020420"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1101","DOI":"10.1002\/hep.31705","article-title":"Advanced techniques and awaited clinical applications for human pluripotent stem cell differentiation into hepatocytes","volume":"74","author":"Luce","year":"2021","journal-title":"Hepatology"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1016\/j.diff.2019.02.004","article-title":"Stem-cell derived hepatocyte-like cells for the assessment of drug-induced liver injury","volume":"106","author":"Donato","year":"2019","journal-title":"Differentiation"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"95","DOI":"10.1038\/s41422-019-0261-5","article-title":"Hepatic spheroids derived from human induced pluripotent stem cells in bio-artificial liver rescue porcine acute liver failure","volume":"30","author":"Chen","year":"2020","journal-title":"Cell Res."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"581","DOI":"10.1016\/j.jhep.2014.10.016","article-title":"Phenotypic and function-al analyses show stem cell-derived hepatocyte-like cells better mimic fetal rather than adult hepatocytes","volume":"62","author":"Baxter","year":"2015","journal-title":"J. Hepatol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"504","DOI":"10.1016\/j.biotechadv.2014.01.003","article-title":"Pluripotent stem cell-derived hepatocyte-like cells","volume":"32","author":"Schwartz","year":"2014","journal-title":"Biotechnol. Adv."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1258","DOI":"10.1053\/j.gastro.2018.01.066","article-title":"Biotechnology Challenges to In Vitro Maturation of Hepatic Stem Cells","volume":"154","author":"Chen","year":"2018","journal-title":"Gastroenterology"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"132","DOI":"10.1016\/j.biotechadv.2017.10.005","article-title":"Next generation organoids for biomedical research and applications","volume":"36","author":"Lou","year":"2018","journal-title":"Biotechnol. Adv."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"3","DOI":"10.1002\/cld.855","article-title":"Organoid Medicine in Hepatology","volume":"15","author":"Sakabe","year":"2020","journal-title":"Clin. Liver Dis."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"132","DOI":"10.1016\/j.addr.2014.02.011","article-title":"Bioreactor technologies to support liver function in vitro","volume":"69\u201370","author":"Ebrahimkhani","year":"2014","journal-title":"Adv. Drug Deliv. Rev."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1586","DOI":"10.1016\/j.cell.2016.05.082","article-title":"Modeling Development and Disease with Organoids","volume":"165","author":"Clevers","year":"2016","journal-title":"Cell"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"e1800430","DOI":"10.1002\/adhm.201800430","article-title":"Bioengineering Organs for Blood Detoxification","volume":"7","author":"Legallais","year":"2018","journal-title":"Adv. Healthc. Mater."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"608","DOI":"10.1248\/bpb.b19-00740","article-title":"Generation of Human Induced Pluripotent Stem Cell-Derived Hepatocyte-Like Cells for Cellular Medicine","volume":"43","author":"Deguchi","year":"2020","journal-title":"Biol. Pharm. Bull."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"221","DOI":"10.1186\/s13287-019-1342-6","article-title":"Low-density lipoprotein receptor-deficient hepatocytes differentiated from induced pluripotent stem cells allow familial hypercholes-terolemia modeling, CRISPR\/Cas-mediated genetic correction, and productive hepatitis C virus infection","volume":"10","author":"Caron","year":"2019","journal-title":"Stem Cell Res. Ther."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"686","DOI":"10.5966\/sctm.2013-0158","article-title":"Messenger RNA- versus Retrovirus-Based Induced Pluripotent Stem Cell Reprogramming Strategies: Analysis of Genomic Integrity","volume":"3","author":"Steichen","year":"2014","journal-title":"Stem Cells Transl. Med."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"2090","DOI":"10.1002\/term.2105","article-title":"Self-assembly of tissue spheroids on polymeric membranes: Self-assembly of spheroids on polymeric membranes","volume":"11","author":"Messina","year":"2017","journal-title":"J. Tissue Eng. Regen. Med."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"135","DOI":"10.1016\/j.jchromb.2012.05.015","article-title":"Bile acid profiling in human biological samples: Comparison of extraction procedures and application to normal and cholestatic patients","volume":"899","author":"Humbert","year":"2012","journal-title":"J. Chromatogr. B"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"409","DOI":"10.5966\/sctm.2012-0160","article-title":"Highly Efficient Differentiation of Functional Hepatocytes From Human Induced Pluripotent Stem Cells","volume":"2","author":"Ma","year":"2013","journal-title":"Stem Cells Transl. Med."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"419","DOI":"10.1124\/dmd.116.074369","article-title":"Transcriptional, Functional, and Mechanistic Comparisons of Stem Cell-Derived Hepatocytes, HepaRG Cells, and Three-Dimensional Human Hepatocyte Spheroids as Predictive In Vitro Systems for Drug-Induced Liver Injury","volume":"45","author":"Bell","year":"2017","journal-title":"Drug Metab. Dispos. Biol. Fate Chem."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"1259","DOI":"10.1089\/scd.2010.0361","article-title":"Comparative Analysis of Human Embryonic Stem Cell and Induced Pluripotent Stem Cell-Derived Hepatocyte-Like Cells Reveals Current Drawbacks and Possible Strategies for Improved Differentiation","volume":"20","author":"Jozefczuk","year":"2011","journal-title":"Stem Cells Dev."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"934","DOI":"10.1016\/j.jhep.2015.05.013","article-title":"Gene net-works and transcription factor motifs defining the differentiation of stem cells into hepatocyte-like cells","volume":"63","author":"Godoy","year":"2015","journal-title":"J. Hepatol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1813","DOI":"10.1016\/j.stemcr.2017.10.019","article-title":"Distinct Gene Expression and Epigenetic Signatures in Hepatocyte-like Cells Produced by Different Strategies from the Same Donor","volume":"9","author":"Gao","year":"2017","journal-title":"Stem Cell Rep."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"558","DOI":"10.1016\/j.stem.2017.03.017","article-title":"Large, Diverse Population Cohorts of hiPSCs and Derived Hepatocyte-like Cells Reveal Functional Genetic Variation at Blood Lipid-Associated Loci","volume":"20","author":"Pashos","year":"2017","journal-title":"Cell Stem Cell"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"90","DOI":"10.1016\/S0014-5793(01)02140-8","article-title":"Oncostatin M and hepatocyte growth factor induce hepatic maturation via distinct signaling pathways","volume":"492","author":"Kamiya","year":"2001","journal-title":"FEBS Lett."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"281","DOI":"10.1203\/00006450-199109000-00017","article-title":"Dexamethasone Stimulates Vitamin K-Dependent Carboxylase Activity in Neonatal Rats and Cultured Fetal Hepatocytes","volume":"30","author":"Wallin","year":"1991","journal-title":"Pediatr. Res."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1807","DOI":"10.1093\/carcin\/15.9.1807","article-title":"Enhancement of liver cell gap junction protein expression by glucocorticoids","volume":"15","author":"Ren","year":"1994","journal-title":"Carcinogenesis"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"37388","DOI":"10.1038\/srep37388","article-title":"Connexin 32-mediated cell-cell communication is essential for hepatic differentiation from human embryonic stem cells","volume":"6","author":"Qin","year":"2016","journal-title":"Sci. Rep."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2085","DOI":"10.1016\/S0024-3205(01)01525-9","article-title":"Vitamin K uptake in hepatocytes and hepatoma cells","volume":"70","author":"Li","year":"2002","journal-title":"Life Sci."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"265","DOI":"10.1002\/hep.27803","article-title":"Microbial-derived lithocholic acid and vitamin K 2 drive the metabolic maturation of pluripotent stem cells-derived and fetal hepatocytes","volume":"62","author":"Avior","year":"2015","journal-title":"Hepatology"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"103","DOI":"10.1016\/j.jnutbio.2017.09.022","article-title":"Vitamin K1 inversely correlates with glycemia and insulin resistance in patients with type 2 diabetes (T2D) and positively regulates SIRT1\/AMPK pathway of glucose metabolism in liver of T2D mice and hepatocytes cultured in high glucose","volume":"52","author":"Dihingia","year":"2018","journal-title":"J. Nutr. Biochem."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"174","DOI":"10.1016\/j.jnutbio.2019.05.006","article-title":"Gamma-glutamyl carboxylated Gas6 mediates the beneficial effect of vitamin K on lowering hyperlipidemia via regulating the AMPK\/SREBP1\/PPAR\u03b1 signaling cascade of lipid metabolism","volume":"70","author":"Bordoloi","year":"2019","journal-title":"J. Nutr. Biochem."},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Luce, E., Steichen, C., Allouche, M., Messina, A., Heslan, J., Lambert, T., Weber, A., Nguyen, T.H., Christophe, O., and Dubart-Kupperschmitt, A. (2021). In vitro recovery of FIX clotting activity as a marker of highly functional hepatocytes in a hemophilia B iPSC model. Hepatology, Online ahead of print.","DOI":"10.1002\/hep.32211"},{"key":"ref_35","doi-asserted-by":"crossref","unstructured":"Gieseck, R.L., Hannan, N.R.F., Bort, R., Hanley, N.A., Drake, R.A.L., Cameron, G.W.W., Wynn, T.A., and Vallier, L. (2014). Maturation of Induced Pluripotent Stem Cell Derived Hepatocytes by 3D-Culture. PLoS ONE, 9.","DOI":"10.1371\/journal.pone.0086372"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"627","DOI":"10.1016\/j.stemcr.2019.08.007","article-title":"Robust, Long-Term Culture of Endoderm-Derived Hepatic Organoids for Disease Modeling","volume":"13","author":"Akbari","year":"2019","journal-title":"Stem Cell Rep."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"970","DOI":"10.1016\/j.jhep.2019.06.030","article-title":"Generation of expandable human pluripotent stem cell-derived hepatocyte-like liver organoids","volume":"71","author":"Mun","year":"2019","journal-title":"J. Hepatol."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"e9968","DOI":"10.7717\/peerj.9968","article-title":"Generation of human liver organoids from pluripotent stem cell-derived hepatic endoderms","volume":"8","author":"Kulkeaw","year":"2020","journal-title":"PeerJ"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"831","DOI":"10.1053\/j.gastro.2020.10.002","article-title":"High-Fidelity Drug-Induced Liver Injury Screen Using Human Pluripotent Stem Cell\u2013Derived Organoids","volume":"160","author":"Shinozawa","year":"2021","journal-title":"Gastroenterology"}],"container-title":["Cells"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4409\/11\/3\/537\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T22:13:52Z","timestamp":1760134432000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4409\/11\/3\/537"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,2,4]]},"references-count":39,"journal-issue":{"issue":"3","published-online":{"date-parts":[[2022,2]]}},"alternative-id":["cells11030537"],"URL":"https:\/\/doi.org\/10.3390\/cells11030537","relation":{},"ISSN":["2073-4409"],"issn-type":[{"value":"2073-4409","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,2,4]]}}}