{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,20]],"date-time":"2026-08-20T15:08:20Z","timestamp":1787238500070,"version":"build-2736575974"},"reference-count":55,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2023,5,19]],"date-time":"2023-05-19T00:00:00Z","timestamp":1684454400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001700","name":"Japanese Ministry of Education, Culture, Sports, Science and Technology (MEXT)","doi-asserted-by":"publisher","award":["18H03379"],"award-info":[{"award-number":["18H03379"]}],"id":[{"id":"10.13039\/501100001700","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001700","name":"Japanese Ministry of Education, Culture, Sports, Science and Technology (MEXT)","doi-asserted-by":"publisher","award":["18J23449"],"award-info":[{"award-number":["18J23449"]}],"id":[{"id":"10.13039\/501100001700","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>The nuclear estrogen receptor (ER) and G-protein-coupled ER (GPER1) play a crucial role during brain development and are involved in dendrite and spine growth as well as synapse formation. Soybean isoflavones, such as genistein, daidzein, and S-equol, a daidzein metabolite, exert their action through ER and GPER1. However, the mechanisms of action of isoflavones on brain development, particularly during dendritogenesis and neuritogenesis, have not yet been extensively studied. We evaluated the effects of isoflavones using mouse primary cerebellar culture, astrocyte-enriched culture, Neuro-2A clonal cells, and co-culture with neurons and astrocytes. Soybean isoflavone-augmented estradiol mediated dendrite arborization in Purkinje cells. Such augmentation was suppressed by co-exposure with ICI 182,780, an antagonist for ERs, or G15, a selective GPER1 antagonist. The knockdown of nuclear ERs or GPER1 also significantly reduced the arborization of dendrites. Particularly, the knockdown of ER\u03b1 showed the greatest effect. To further examine the specific molecular mechanism, we used Neuro-2A clonal cells. Isoflavones also induced neurite outgrowth of Neuro-2A cells. The knockdown of ER\u03b1 most strongly reduced isoflavone-induced neurite outgrowth compared with ER\u03b2 or GPER1 knockdown. The knockdown of ER\u03b1 also reduced the mRNA levels of ER-responsive genes (i.e., Bdnf, Camk2b, Rbfox3, Tubb3, Syn1, Dlg4, and Syp). Furthermore, isoflavones increased ER\u03b1 levels, but not ER\u03b2 or GPER1 levels, in Neuro-2A cells. The co-culture study of Neuro-2A cells and astrocytes also showed an increase in isoflavone-induced neurite growth, and co-exposure with ICI 182,780 or G15 significantly reduced the effects. In addition, isoflavones increased astrocyte proliferation via ER and GPER1. These results indicate that ER\u03b1 plays an essential role in isoflavone-induced neuritogenesis. However, GPER1 signaling is also necessary for astrocyte proliferation and astrocyte\u2013neuron communication, which may lead to isoflavone-induced neuritogenesis.<\/jats:p>","DOI":"10.3390\/ijms24109011","type":"journal-article","created":{"date-parts":[[2023,5,19]],"date-time":"2023-05-19T10:35:11Z","timestamp":1684492511000},"page":"9011","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":14,"title":["Isoflavones Mediate Dendritogenesis Mainly through Estrogen Receptor \u03b1"],"prefix":"10.3390","volume":"24","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-4813-6604","authenticated-orcid":false,"given":"Winda","family":"Ariyani","sequence":"first","affiliation":[{"name":"Metabolic Signal Research Center, Institute for Molecular and Cellular Regulation, Gunma University, 3-39-15 Showa-machi, Maebashi 371-8512, Japan"},{"name":"Department of Integrative Physiology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi 371-8511, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5076-2041","authenticated-orcid":false,"given":"Izuki","family":"Amano","sequence":"additional","affiliation":[{"name":"Department of Integrative Physiology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi 371-8511, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2247-9740","authenticated-orcid":false,"given":"Noriyuki","family":"Koibuchi","sequence":"additional","affiliation":[{"name":"Department of Integrative Physiology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi 371-8511, Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,5,19]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"211","DOI":"10.2478\/v10102-009-0021-3","article-title":"Isoflavonoids\u2014An overview of their biological activities and potential health benefits","volume":"2","year":"2009","journal-title":"Interdiscip. Toxicol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1280","DOI":"10.1080\/10408398.2014.989958","article-title":"An updated review of dietary isoflavones: Nutrition, processing, bioavailability and impacts on human health","volume":"57","author":"Zaheer","year":"2017","journal-title":"Crit. Rev. Food Sci. Nutr."},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"Gorzkiewicz, J., Bartosz, G., and Sadowska-Bartosz, I. (2021). The potential effects of phytoestrogens: The role in neuroprotection. Molecules, 26.","DOI":"10.3390\/molecules26102954"},{"key":"ref_4","doi-asserted-by":"crossref","unstructured":"K\u0159\u00ed\u017eov\u00e1, L., Dad\u00e1kov\u00e1, K., Ka\u0161parovsk\u00e1, J., and Ka\u0161parovsk\u00fd, T. (2019). Isoflavones. Molecules, 24.","DOI":"10.3390\/molecules24061076"},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Ariyani, W., Miyazaki, W., and Koibuchi, N. (2019). A Novel Mechanism of S-equol Action in Neurons and Astrocytes: The Possible Involvement of GPR30\/GPER1. Int. J. Mol. Sci., 20.","DOI":"10.3390\/ijms20205178"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"554941","DOI":"10.3389\/fendo.2020.554941","article-title":"Soy Isoflavones Accelerate Glial Cell Migration via GPER-Mediated Signal Transduction Pathway","volume":"11","author":"Ariyani","year":"2020","journal-title":"Front. Endocrinol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"32","DOI":"10.1016\/j.mce.2009.03.026","article-title":"Mechanisms of estrogen signaling and gene expression via GPR30","volume":"308","author":"Prossnitz","year":"2009","journal-title":"Mol. Cell. Endocrinol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"505","DOI":"10.1124\/pr.114.009712","article-title":"International Union of Basic and Clinical Pharmacology. XCVII. G Protein\u2013Coupled Estrogen Receptor and Its Pharmacologic Modulators","volume":"67","author":"Prossnitz","year":"2015","journal-title":"Pharmacol. Rev."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1336","DOI":"10.3389\/fphar.2019.01336","article-title":"Molecular mechanisms of action of genistein in cancer: Recent advances","volume":"10","author":"Tuli","year":"2019","journal-title":"Front. Pharmacol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"667643","DOI":"10.3389\/fnut.2021.667643","article-title":"Antioxidant Effect of Soymilk Fermented by Lactobacillus plantarum HFY01 on D-Galactose-Induced Premature Aging Mouse Model","volume":"8","author":"Li","year":"2021","journal-title":"Front. Nutr."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"135","DOI":"10.1007\/PL00007388","article-title":"Isoflavones, substances with multi-biological and clinical properties","volume":"40","author":"Ren","year":"2001","journal-title":"Eur. J. Nutr."},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Lecomte, S., Demay, F., Ferri, F., and Pakdel, F. (2017). Phytochemicals Targeting Estrogen Receptors: Beneficial Rather Than Adverse Effects?. Int. J. Mol. Sci., 18.","DOI":"10.3390\/ijms18071381"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"2","DOI":"10.1016\/j.neulet.2018.05.013","article-title":"Insights into cerebellar development and connectivity","volume":"688","author":"Beckinghausen","year":"2019","journal-title":"Neurosci. Lett."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"417","DOI":"10.1093\/toxsci\/kfy097","article-title":"A possible novel mechanism of action of genistein and daidzein for activating thyroid hormone receptor-mediated transcription","volume":"164","author":"Ariyani","year":"2018","journal-title":"Toxicol. Sci."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"361","DOI":"10.1016\/j.brainres.2017.01.002","article-title":"Low dietary soy isoflavonoids increase hippocampal spine synapse density in ovariectomized rats","volume":"1657","author":"MacLusky","year":"2017","journal-title":"Brain Res."},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Rivera, P., P\u00e9rez-Mart\u00edn, M., Pav\u00f3n, F.J., Serrano, A., Crespillo, A., Cifuentes, M., L\u00f3pez-\u00c1valos, M.D., Grondona, J.M., Vida, M., and Fern\u00e1ndez-Llebrez, P. (2013). Pharmacological Administration of the Isoflavone Daidzein Enhances Cell Proliferation and Reduces High Fat Diet-Induced Apoptosis and Gliosis in the Rat Hippocampus. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0064750"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1772","DOI":"10.1152\/ajpcell.2000.279.6.C1772","article-title":"\u03b2-Amyloid-induced migration of monocytes across human brain endothelial cells involves RAGE and PECAM-1","volume":"279","author":"Giri","year":"2000","journal-title":"Am. J. Physiol. Cell Physiol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"758","DOI":"10.1016\/j.bcp.2009.10.005","article-title":"Genistein and daidzein prevent low potassium-dependent apoptosis of cerebellar granule cells","volume":"79","author":"Atlante","year":"2010","journal-title":"Biochem. Pharmacol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"57","DOI":"10.1111\/j.1365-2826.2005.01268.x","article-title":"Phytoestrogen action in the adult and developing brain","volume":"17","author":"Patisaul","year":"2005","journal-title":"J. Neuroendocrinol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"5","DOI":"10.1016\/S0892-0362(01)00197-0","article-title":"Neurobehavioral effects of dietary soy phytoestrogens","volume":"24","author":"Lephart","year":"2002","journal-title":"Neurotoxicol. Teratol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"40","DOI":"10.1016\/j.physbeh.2014.03.006","article-title":"Perinatal exposure to genistein, a soy phytoestrogen, improves spatial learning and memory but impairs passive avoidance learning and memory in offspring","volume":"130","author":"Kohara","year":"2014","journal-title":"Physiol. Behav."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"1118","DOI":"10.1080\/09637486.2021.1904844","article-title":"Maternal consumption of soy and isoflavones during pregnancy and risk of childhood behavioural problems: The Kyushu Okinawa Maternal and Child Health Study","volume":"72","author":"Miyake","year":"2021","journal-title":"Int. J. Food Sci. Nutr."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"441","DOI":"10.1007\/s00394-016-1327-5","article-title":"Soy isoflavone intake and prevalence of depressive symptoms during pregnancy in Japan: Baseline data from the Kyushu Okinawa Maternal and Child Health Study","volume":"57","author":"Miyake","year":"2018","journal-title":"Eur. J. Nutr."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"87","DOI":"10.1016\/j.neulet.2011.09.067","article-title":"Morphological effects of isoflavones (daidzein and genistein) on hypothalamic oxytocin neurons in the neonatal mouse brain slice cultures","volume":"505","author":"Yoshimura","year":"2011","journal-title":"Neurosci. Lett."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"938596","DOI":"10.3389\/fendo.2022.938596","article-title":"Involvement of integrin \u03b1v\u03b23 in thyroid hormone-induced dendritogenesis","volume":"13","author":"Ariyani","year":"2022","journal-title":"Front. Endocrinol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"187","DOI":"10.1146\/annurev-neuro-070918-050443","article-title":"The Emerging Nature of Astrocyte Diversity","volume":"42","author":"Khakh","year":"2019","journal-title":"Annu. Rev. Neurosci."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"942","DOI":"10.1038\/nn.4043","article-title":"Diversity of astrocyte functions and phenotypes in neural circuits","volume":"18","author":"Khakh","year":"2015","journal-title":"Nat. Neurosci."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"272","DOI":"10.1007\/s11064-016-2089-4","article-title":"Astrocytes in Migration","volume":"42","author":"Zhan","year":"2017","journal-title":"Neurochem. Res."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"263","DOI":"10.1146\/annurev-genet-112618-043515","article-title":"Microglia in Brain Development, Homeostasis, and Neurodegeneration","volume":"53","author":"Bohlen","year":"2019","journal-title":"Annu. Rev. Genet."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1218","DOI":"10.1111\/dmcn.13546","article-title":"Principles of plasticity in the developing brain","volume":"59","author":"Kolb","year":"2017","journal-title":"Dev. Med. Child Neurol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"1014","DOI":"10.3389\/fimmu.2018.01014","article-title":"Microglial dynamics during human brain development","volume":"9","author":"Menassa","year":"2018","journal-title":"Front. Immunol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"44","DOI":"10.1016\/j.yfrne.2018.09.001","article-title":"Molecular mechanisms involved in the protective actions of Selective Estrogen Receptor Modulators in brain cells","volume":"52","author":"Ashraf","year":"2019","journal-title":"Front. Neuroendocrinol."},{"key":"ref_33","unstructured":"D\u2019Angelo, E. (2018). Physiology of the Cerebellum, Elsevier. [1st ed.]."},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"van Essen, M.J., Nayler, S., Becker, E.B.E., and Jacob, J. (2020). Deconstructing cerebellar development cell by cell. PLoS Genet., 16.","DOI":"10.1371\/journal.pgen.1008630"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"51","DOI":"10.3389\/fncir.2019.00051","article-title":"A New Projection From the Deep Cerebellar Nuclei to the Hippocampus via the Ventrolateral and Laterodorsal Thalamus in Mice","volume":"13","author":"Bohne","year":"2019","journal-title":"Front. Neural Circuits"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"249","DOI":"10.2170\/jjphysiol.54.249","article-title":"A genistein-sensitive Na+\/Ca2+ exchange is responsible for the resting [Ca2+]i and most of the Ca2+ plasma membrane fluxes in stimulated rat cerebellar type 1 astrocytes","volume":"54","author":"Rojas","year":"2004","journal-title":"Jpn. J. Physiol."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"11414","DOI":"10.1523\/JNEUROSCI.6446-11.2012","article-title":"Alternative splicing of the TRPC3 ion channel calmodulin\/ IP 3 receptor-binding domain in the hindbrain Enhances cation flux","volume":"32","author":"Kim","year":"2012","journal-title":"J. Neurosci."},{"key":"ref_38","first-page":"9","article-title":"Brain Development and the Role of Experience in the Early Years","volume":"30","author":"Tierney","year":"2009","journal-title":"Zero Three"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"146427","DOI":"10.1016\/j.brainres.2019.146427","article-title":"Mouse vs man: Organoid models of brain development & disease","volume":"1724","author":"Marshall","year":"2019","journal-title":"Brain Res."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"91","DOI":"10.1152\/physrev.00010.2007","article-title":"Estradiol and the developing brain","volume":"88","author":"Mccarthy","year":"2009","journal-title":"Physiol. Rev."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"10039","DOI":"10.1523\/JNEUROSCI.1262-16.2016","article-title":"A Critical Period in Purkinje Cell Development Is Mediated by Local Estradiol Synthesis, Disrupted by Inflammation, and Has Enduring Consequences Only for Males","volume":"36","author":"Hoffman","year":"2016","journal-title":"J. Neurosci."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"676","DOI":"10.1016\/j.phymed.2008.04.007","article-title":"The phytoestrogenic isoflavones from Trifolium pratense L. (Red clover) protects human cortical neurons from glutamate toxicity","volume":"15","author":"Occhiuto","year":"2008","journal-title":"Phytomedicine"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"80","DOI":"10.1186\/1423-0127-19-80","article-title":"Daidzein induces neuritogenesis in DRG neuronal cultures","volume":"19","author":"Yang","year":"2012","journal-title":"J. Biomed. Sci."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"59","DOI":"10.3389\/fnmol.2018.00059","article-title":"Genistein, a phytoestrogen in soybean, induces the expression of acetylcholinesterase via G protein-coupled receptor 30 in PC12 cells","volume":"11","author":"Liu","year":"2018","journal-title":"Front. Mol. Neurosci."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"351","DOI":"10.2220\/biomedres.32.351","article-title":"Genistein enhances the NGF-induced neurite outgrowth","volume":"32","author":"Nakajima","year":"2011","journal-title":"Biomed. Res."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"267","DOI":"10.1097\/00001756-200502280-00013","article-title":"Genistein protects dopaminergic neurons by inhibiting microglial activation","volume":"16","author":"Wang","year":"2005","journal-title":"Neuroreport"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"123","DOI":"10.1016\/j.neures.2008.07.001","article-title":"Protective effect of isoflavones from Trifolium pratense on dopaminergic neurons","volume":"62","author":"Chen","year":"2008","journal-title":"Neurosci. Res."},{"key":"ref_48","doi-asserted-by":"crossref","unstructured":"Subedi, L., Ji, E., Shin, D., Jin, J., Yeo, J.H., and Kim, S.Y. (2017). Equol, a Dietary Daidzein Gut Metabolite Attenuates Microglial Activation and Potentiates Neuroprotection In Vitro. Nutrients, 9.","DOI":"10.3390\/nu9030207"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"1710","DOI":"10.1093\/jn\/128.10.1710","article-title":"Pharmacokinetics of soybean isoflavones in plasma, urine and feces of men after ingestion of 60 g baked soybean powder (kinako)","volume":"128","author":"Watanabe","year":"1998","journal-title":"J. Nutr."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"1363S","DOI":"10.3945\/jn.109.119784","article-title":"Equol: Pharmacokinetics and Biological Actions","volume":"140","author":"Setchell","year":"2010","journal-title":"J. Nutr."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"2703","DOI":"10.1007\/s00204-018-2279-8","article-title":"Effects of isoflavones on breast tissue and the thyroid hormone system in humans: A comprehensive safety evaluation","volume":"92","author":"Guth","year":"2018","journal-title":"Arch. Toxicol."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"1045","DOI":"10.1158\/1940-6207.CAPR-14-0465","article-title":"Isoflavone pharmacokinetics and metabolism after consumption of a standardized soy and soy-almond bread in men with asymptomatic prostate cancer","volume":"8","author":"Clinton","year":"2015","journal-title":"Cancer Prev. Res."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"1485","DOI":"10.1124\/dmd.108.021006","article-title":"Plasma profiling of intact isoflavone metabolites by high-performance liquid chromatography and mass spectrometric identification of flavone glycosides daidzin and genistin in human plasma after administration of kinako","volume":"36","author":"Hosoda","year":"2008","journal-title":"Drug Metab. Dispos."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"115","DOI":"10.3389\/fendo.2016.00115","article-title":"Effects of gadolinium-based contrast agents on thyroid hormone receptor action and thyroid hormone-induced cerebellar purkinje cell morphogenesis","volume":"7","author":"Ariyani","year":"2016","journal-title":"Front. Endocrinol."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"5850","DOI":"10.1038\/s41598-022-09882-7","article-title":"Gadolinium-based contrast agent accelerates the migration of astrocyte via integrin \u03b1v\u03b23 signaling pathway","volume":"12","author":"Ariyani","year":"2022","journal-title":"Sci. Rep."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/24\/10\/9011\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T19:38:38Z","timestamp":1760125118000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/24\/10\/9011"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,5,19]]},"references-count":55,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2023,5]]}},"alternative-id":["ijms24109011"],"URL":"https:\/\/doi.org\/10.3390\/ijms24109011","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,5,19]]}}}