{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T18:24:19Z","timestamp":1785435859603,"version":"3.56.0"},"reference-count":50,"publisher":"MDPI AG","issue":"15","license":[{"start":{"date-parts":[[2026,7,24]],"date-time":"2026-07-24T00:00:00Z","timestamp":1784851200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100007300","name":"China Medical University","doi-asserted-by":"publisher","award":["CMU-112-S-44"],"award-info":[{"award-number":["CMU-112-S-44"]}],"id":[{"id":"10.13039\/501100007300","id-type":"DOI","asserted-by":"publisher"}]},{"award":["CMU-112-S-44"],"award-info":[{"award-number":["CMU-112-S-44"]}],"id":[{"id":"https:\/\/ror.org\/00v408z34","id-type":"ROR","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100004391","name":"China Medical University Hospital","doi-asserted-by":"publisher","award":["DMR-115-127"],"award-info":[{"award-number":["DMR-115-127"]}],"id":[{"id":"10.13039\/501100004391","id-type":"DOI","asserted-by":"publisher"}]},{"award":["DMR-115-127"],"award-info":[{"award-number":["DMR-115-127"]}],"id":[{"id":"https:\/\/ror.org\/0368s4g32","id-type":"ROR","asserted-by":"publisher"}]},{"name":"Hualien Tzu Chi Medical Center","award":["TCMF-CM3-112-01"],"award-info":[{"award-number":["TCMF-CM3-112-01"]}]},{"award":["TCMF-CM3-112-01"],"award-info":[{"award-number":["TCMF-CM3-112-01"]}],"id":[{"id":"https:\/\/ror.org\/030k6vb67","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Int. J. Mol. Sci.","IJMS"],"abstract":"<jats:p>Metabolic dysfunction-associated steatotic liver disease (MASLD) is the most prevalent chronic liver disease worldwide and is characterized by excessive hepatic lipid accumulation and metabolic dysfunction. Although microRNA-34a (miR-34a) has been implicated in hepatic lipid metabolism, the molecular mechanisms underlying its contribution to MASLD remain incompletely understood. Here, we investigated the role of miR-34a in FFA-induced hepatic steatosis using HepG2 cells and explored RXR-associated signaling as a potential therapeutic strategy. RT-qPCR quantified miR-34a expression; direct target interactions were validated using dual-luciferase reporter assays; proteomic alterations were characterized by iTRAQ-based proteomics followed by Ingenuity Pathway Analysis (IPA); and transcriptomic responses to 9-cis-retinoic acid (9-cis-RA) were analyzed by RNA sequencing. FFA treatment significantly increased miR-34a expression, and dual-luciferase assays confirmed that miR-34a directly targets the 3\u2032-UTRs of RXR\u03b1, PPAR\u03b1, and SIRT1. Integrated proteomic and transcriptomic analyses consistently identified LXR\/RXR signaling as one of the principal pathways associated with miR-34a dysregulation and 9-cis-RA treatment. Pharmacological activation of RXR-associated signaling with the pan-RXR agonist 9-cis-RA attenuated intracellular lipid accumulation and reduced the expression of key regulators of lipogenesis and fatty acid uptake, including FASN, SCD1, FABP4, and CD36. Collectively, these findings support the miR-34a\u2013RXR\u03b1 axis as one regulatory component within a broader nuclear receptor network associated with hepatic lipid homeostasis and support further investigation of RXR-associated signaling as a potential therapeutic strategy for MASLD. Nevertheless, confirmation of receptor-specific mechanisms and validation in more physiologically relevant experimental models will be required.<\/jats:p>","DOI":"10.3390\/ijms27156609","type":"journal-article","created":{"date-parts":[[2026,7,27]],"date-time":"2026-07-27T09:47:41Z","timestamp":1785145661000},"page":"6609","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["MicroRNA-34a Promotes Hepatic Lipid Accumulation Through RXR\u03b1 Suppression and Is Reversed by 9-cis-Retinoic Acid in Steatotic Hepatocytes"],"prefix":"10.3390","volume":"27","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-7302-8248","authenticated-orcid":false,"given":"Jai-Sing","family":"Yang","sequence":"first","affiliation":[{"name":"Department of Medical Research, China Medical University Hospital, China Medical University, Taichung 404327, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hong-Yi","family":"Chiu","sequence":"additional","affiliation":[{"name":"Department of Pharmacy, Hualien Tzu Chi Hospital, Buddhist Tzu Chi Medical Foundation, Hualien 970473, Taiwan"},{"name":"Graduate Institute of Clinical Pharmacy, College of Medicine, Tzu Chi University, Hualien 970473, Taiwan"},{"name":"Department of Pharmacy, College of Medicine, Tzu Chi University, Hualien 970473, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Syun-Rong","family":"Jhan","sequence":"additional","affiliation":[{"name":"Department of Biological Science and Technology, China Medical University, Taichung 406040, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yao-An","family":"Liu","sequence":"additional","affiliation":[{"name":"Department of Biological Science and Technology, China Medical University, Taichung 406040, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chao-Jung","family":"Chen","sequence":"additional","affiliation":[{"name":"Graduate Institute of Integrated Medicine, China Medical University, Taichung 404327, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9018-607X","authenticated-orcid":false,"given":"Shih-Chang","family":"Tsai","sequence":"additional","affiliation":[{"name":"Department of Biological Science and Technology, China Medical University, Taichung 406040, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,7,24]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"202","DOI":"10.1016\/j.jhep.2020.03.039","article-title":"A new definition for metabolic dysfunction-associated fatty liver disease: An international expert consensus statement","volume":"73","author":"Eslam","year":"2020","journal-title":"J. Hepatol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1542","DOI":"10.1016\/j.jhep.2023.06.003","article-title":"A multisociety Delphi consensus statement on new fatty liver disease nomenclature","volume":"79","author":"Rinella","year":"2023","journal-title":"J. Hepatol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"908","DOI":"10.1038\/s41591-018-0104-9","article-title":"Mechanisms of NAFLD development and therapeutic strategies","volume":"24","author":"Friedman","year":"2018","journal-title":"Nat. Med."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1037","DOI":"10.21873\/invivo.13178","article-title":"Liraglutide with Metformin Therapy Ameliorates Hepatic Steatosis and Liver Injury in a Mouse Model of Non-alcoholic Steatohepatitis","volume":"37","author":"Chiu","year":"2023","journal-title":"In Vivo"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"20","DOI":"10.1016\/j.cell.2018.03.006","article-title":"Metazoan MicroRNAs","volume":"173","author":"Bartel","year":"2018","journal-title":"Cell"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"239","DOI":"10.1038\/nrm3313","article-title":"MicroRNAs in metabolism and metabolic disorders","volume":"13","author":"Rottiers","year":"2012","journal-title":"Nat. Rev. Mol. Cell Biol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"2065","DOI":"10.1136\/gutjnl-2018-318146","article-title":"miRNAs and NAFLD: From pathophysiology to therapy","volume":"68","author":"Gjorgjieva","year":"2019","journal-title":"Gut"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1810","DOI":"10.1002\/hep.22569","article-title":"Nonalcoholic steatohepatitis is associated with altered hepatic MicroRNA expression","volume":"48","author":"Cheung","year":"2008","journal-title":"Hepatology"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"119","DOI":"10.1016\/j.jhep.2012.08.008","article-title":"miR-34a\/SIRT1\/p53 is suppressed by ursodeoxycholic acid in the rat liver and activated by disease severity in human non-alcoholic fatty liver disease","volume":"58","author":"Castro","year":"2013","journal-title":"J. Hepatol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/S0083-6729(06)75001-4","article-title":"RXR: From partnership to leadership in metabolic regulations","volume":"75","author":"Desvergne","year":"2007","journal-title":"Vitam. Horm."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"13421","DOI":"10.1073\/pnas.0801613105","article-title":"miR-34a repression of SIRT1 regulates apoptosis","volume":"105","author":"Yamakuchi","year":"2008","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1062","DOI":"10.1111\/acel.12135","article-title":"Elevated microRNA-34a in obesity reduces NAD+ levels and SIRT1 activity by directly targeting NAMPT","volume":"12","author":"Choi","year":"2013","journal-title":"Aging Cell"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"101648","DOI":"10.1016\/j.gendis.2025.101648","article-title":"Elevated miR-34a induced by lipotoxicity and inflammation mediates pathophysiological communication between hepatocytes and hepatic stellate cells in liver fibrosis","volume":"12","author":"Duan","year":"2025","journal-title":"Genes Dis."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Erceg, S., Munjas, J., Sopic, M., Tomasevic, R., Mitrovic, M., Kotur-Stevuljevic, J., Mamic, M., Vujcic, S., Klisic, A., and Ninic, A. (2025). Expression Analysis of Circulating miR-21, miR-34a and miR-122 and Redox Status Markers in Metabolic Dysfunction-Associated Steatotic Liver Disease Patients with and Without Type 2 Diabetes. Int. J. Mol. Sci., 26.","DOI":"10.3390\/ijms26062392"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"841","DOI":"10.1016\/0092-8674(95)90200-7","article-title":"The RXR heterodimers and orphan receptors","volume":"83","author":"Mangelsdorf","year":"1995","journal-title":"Cell"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"255","DOI":"10.1016\/j.cell.2014.03.012","article-title":"Nuclear Receptors, RXR, and the Big Bang","volume":"157","author":"Evans","year":"2014","journal-title":"Cell"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"676","DOI":"10.1016\/j.tem.2010.06.009","article-title":"Retinoid X receptors: Common heterodimerization partners with distinct functions","volume":"21","author":"Lefebvre","year":"2010","journal-title":"Trends Endocrinol. Metab."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"4436","DOI":"10.1128\/MCB.20.12.4436-4444.2000","article-title":"Hepatocyte-specific mutation establishes retinoid X receptor alpha as a heterodimeric integrator of multiple physiological processes in the liver","volume":"20","author":"Wan","year":"2000","journal-title":"Mol. Cell. Biol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1083","DOI":"10.1016\/j.bbagrm.2016.03.002","article-title":"Nuclear receptors and nonalcoholic fatty liver disease","volume":"1859","author":"Cave","year":"2016","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Xu, R., Zhang, L., Pan, H., and Zhang, Y. (2024). Retinoid X receptor heterodimers in hepatic function: Structural insights and therapeutic potential. Front. Pharmacol., 15.","DOI":"10.3389\/fphar.2024.1464655"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"179","DOI":"10.1016\/j.bcp.2014.05.002","article-title":"Retinoid X receptor alpha in human liver is regulated by miR-34a","volume":"90","author":"Oda","year":"2014","journal-title":"Biochem. Pharmacol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"397","DOI":"10.1016\/0092-8674(92)90479-V","article-title":"9-cis retinoic acid is a high affinity ligand for the retinoid X receptor","volume":"68","author":"Heyman","year":"1992","journal-title":"Cell"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"359","DOI":"10.1038\/355359a0","article-title":"9-cis retinoic acid stereoisomer binds and activates the nuclear receptor RXR alpha","volume":"355","author":"Levin","year":"1992","journal-title":"Nature"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1027","DOI":"10.1002\/cmdc.201600105","article-title":"An Endogenous Mammalian Retinoid X Receptor Ligand, At Last!","volume":"11","author":"Krezel","year":"2016","journal-title":"ChemMedChem"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"101331","DOI":"10.1016\/j.jbc.2021.101331","article-title":"A retinoic acid receptor beta2 agonist attenuates transcriptome and metabolome changes underlying nonalcohol-associated fatty liver disease","volume":"297","author":"Tang","year":"2021","journal-title":"J. Biol. Chem."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"13729","DOI":"10.1038\/srep13729","article-title":"Effect of miR-34a in regulating steatosis by targeting PPARalpha expression in nonalcoholic fatty liver disease","volume":"5","author":"Ding","year":"2015","journal-title":"Sci. Rep."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"12604","DOI":"10.1074\/jbc.M109.094524","article-title":"A pathway involving farnesoid X receptor and small heterodimer partner positively regulates hepatic sirtuin 1 levels via microRNA-34a inhibition","volume":"285","author":"Lee","year":"2010","journal-title":"J. Biol. Chem."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"985","DOI":"10.1210\/me.2003-0061","article-title":"Liver X receptor signaling pathways in cardiovascular disease","volume":"17","author":"Tontonoz","year":"2003","journal-title":"Mol. Endocrinol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"213","DOI":"10.1038\/nrm3312","article-title":"Transcriptional integration of metabolism by the nuclear sterol-activated receptors LXR and FXR","volume":"13","author":"Calkin","year":"2012","journal-title":"Nat. Rev. Mol. Cell Biol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2819","DOI":"10.1101\/gad.844900","article-title":"Regulation of mouse sterol regulatory element-binding protein-1c gene (SREBP-1c) by oxysterol receptors, LXRalpha and LXRbeta","volume":"14","author":"Repa","year":"2000","journal-title":"Genes Dev."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"1125","DOI":"10.1172\/JCI0215593","article-title":"SREBPs: Activators of the complete program of cholesterol and fatty acid synthesis in the liver","volume":"109","author":"Horton","year":"2002","journal-title":"J. Clin. Investig."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"S126","DOI":"10.1038\/sj.cdd.4401533","article-title":"Retinoid X receptors: X-ploring their (patho)physiological functions","volume":"11","author":"Szanto","year":"2004","journal-title":"Cell Death Differ."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"720","DOI":"10.1016\/j.jhep.2014.10.039","article-title":"Molecular mechanism of PPARalpha action and its impact on lipid metabolism, inflammation and fibrosis in non-alcoholic fatty liver disease","volume":"62","author":"Pawlak","year":"2015","journal-title":"J. Hepatol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"866","DOI":"10.1194\/jlr.M071357","article-title":"FoxO3 increases miR-34a to cause palmitate-induced cholangiocyte lipoapoptosis","volume":"58","author":"Natarajan","year":"2017","journal-title":"J. Lipid Res."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"13","DOI":"10.1038\/s44324-024-00013-6","article-title":"Unlocking therapeutic potential: Exploring cross-talk among emerging nuclear receptors to combat metabolic dysfunction in steatotic liver disease","volume":"2","author":"Antwi","year":"2024","journal-title":"NPJ Metab. Health Dis."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"32523","DOI":"10.1074\/jbc.M003826200","article-title":"Defective uptake and utilization of long chain fatty acids in muscle and adipose tissues of CD36 knockout mice","volume":"275","author":"Coburn","year":"2000","journal-title":"J. Biol. Chem."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"570","DOI":"10.1210\/en.2015-1866","article-title":"Hepatocyte-Specific Disruption of CD36 Attenuates Fatty Liver and Improves Insulin Sensitivity in HFD-Fed Mice","volume":"157","author":"Wilson","year":"2016","journal-title":"Endocrinology"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"91","DOI":"10.1016\/S0163-7827(03)00039-0","article-title":"Regulation of stearoyl-CoA desaturases and role in metabolism","volume":"43","author":"Ntambi","year":"2004","journal-title":"Prog. Lipid Res."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"289","DOI":"10.1016\/S0163-7827(02)00067-X","article-title":"Structural and functional organization of the animal fatty acid synthase","volume":"42","author":"Smith","year":"2003","journal-title":"Prog. Lipid Res."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"6692","DOI":"10.4238\/2013.December.13.3","article-title":"RXR agonists inhibit high glucose-induced upregulation of inflammation by suppressing activation of the NADPH oxidase-nuclear factor-kappaB pathway in human endothelial cells","volume":"12","author":"Ning","year":"2013","journal-title":"Genet. Mol. Res."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"89","DOI":"10.1016\/S0928-0987(01)00211-1","article-title":"The role of hepatocyte RXR alpha in xenobiotic-sensing nuclear receptor-mediated pathways","volume":"15","author":"Cai","year":"2002","journal-title":"Eur. J. Pharm. Sci."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"1202","DOI":"10.1136\/gutjnl-2015-310798","article-title":"Liver PPARalpha is crucial for whole-body fatty acid homeostasis and is protective against NAFLD","volume":"65","author":"Montagner","year":"2016","journal-title":"Gut"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"142","DOI":"10.1016\/j.pharmthera.2017.05.011","article-title":"Targeting nuclear receptors for the treatment of fatty liver disease","volume":"179","author":"Tanaka","year":"2017","journal-title":"Pharmacol. Ther."},{"key":"ref_44","doi-asserted-by":"crossref","unstructured":"Wagner, N., and Wagner, K.D. (2020). The Role of PPARs in Disease. Cells, 9.","DOI":"10.3390\/cells9112367"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"92","DOI":"10.1016\/j.crfs.2020.03.008","article-title":"Piceatannol attenuates fat accumulation and oxidative stress in steatosis-induced HepG2 cells","volume":"3","author":"Yang","year":"2020","journal-title":"Curr. Res. Food Sci."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"207","DOI":"10.1002\/jbt.10085","article-title":"Influence of culture time on the expression of drug-metabolizing enzymes in primary human hepatocytes and hepatoma cell line HepG2","volume":"17","author":"Wilkening","year":"2003","journal-title":"J. Biochem. Mol. Toxicol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"1961","DOI":"10.1084\/jem.20122508","article-title":"Retinoic acid controls the homeostasis of pre-cDC-derived splenic and intestinal dendritic cells","volume":"210","author":"Klebanoff","year":"2013","journal-title":"J. Exp. Med."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"481","DOI":"10.1097\/SHK.0000000000002752","article-title":"Retinoic Acid Attenuates Sepsis-Induced Liver Injury Via RIG-I Inhibition-Mediated Suppression of TNF-alpha\/RIPK1\/RIPK3\/MLKL Pathway","volume":"65","author":"Liu","year":"2026","journal-title":"Shock"},{"key":"ref_49","doi-asserted-by":"crossref","unstructured":"Yen, Y.T., Hsu, C.I., Chen, Y.C., and Tsai, S.C. (2025). Integrative Analysis of miR-21, PTEN, and Immune Signatures in Colorectal Cancer. Int. J. Mol. Sci., 26.","DOI":"10.3390\/ijms262412118"},{"key":"ref_50","doi-asserted-by":"crossref","unstructured":"Forbes-Hernandez, T.Y., Giampieri, F., Gasparrini, M., Afrin, S., Mazzoni, L., Cordero, M.D., Mezzetti, B., Quiles, J.L., and Battino, M. (2017). Lipid Accumulation in HepG2 Cells Is Attenuated by Strawberry Extract through AMPK Activation. Nutrients, 9.","DOI":"10.3390\/nu9060621"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/27\/15\/6609\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,7,28]],"date-time":"2026-07-28T07:57:06Z","timestamp":1785225426000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/27\/15\/6609"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,7,24]]},"references-count":50,"journal-issue":{"issue":"15","published-online":{"date-parts":[[2026,8]]}},"alternative-id":["ijms27156609"],"URL":"https:\/\/doi.org\/10.3390\/ijms27156609","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,7,24]]}}}