{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,27]],"date-time":"2026-09-27T06:51:52Z","timestamp":1790491912801,"version":"4.1.0"},"reference-count":51,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2021,1,8]],"date-time":"2021-01-08T00:00:00Z","timestamp":1610064000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Cell Dev. Biol."],"abstract":"<jats:p>The rapid aging of worldwide populations had led to epidemic increases in the incidence of osteoporosis (OP), but while treatments are available, high cost, adverse effects, and poor compliance continue to be significant problems. Naturally occurring plant-based compounds including phytoestrogens can be good and safe candidates to treat OP, but screening for osteogenic capacity has been difficult to achieve, largely due to the requirement of using primary osteoblasts or mesenchymal stem cells (MSCs), the progenitors of osteoblasts, to conduct time-consuming <jats:italic>in vitro<\/jats:italic> and <jats:italic>in vivo<\/jats:italic> osteogenic assay. Taking advantage of MSC osteogenic capacity and utilizing a promoter reporter assay for Runx2, the master osteogenesis transcription factor, we developed a rapid <jats:italic>in vitro<\/jats:italic> screening platform to screen osteogenic small molecules including natural plant-based compounds. We screened eight plant-derived compounds from different families including flavonoids, polyphenolic compounds, alkaloids, and isothiocyanates for osteogenic capacity using the human RUNX2-promoter luciferase reporter (hRUNX2-luc) transduced into the mouse MSC line, C3H10T1\/2, with daidzein\u2014a well-studied osteogenic phytoestrogen\u2014as a positive control. Classical <jats:italic>in vitro<\/jats:italic> and <jats:italic>in vivo<\/jats:italic> osteogenesis assays were performed using primary murine and human bone marrow MSCs (BMMSCs) to validate the accuracy of this rapid screening platform. Using the MSC\/hRUNX2-luc screening platform, we were able not only to shorten the selection process for osteogenic compounds from 3\u223c4 weeks to just a few days but also simultaneously perform comparisons between multiple compounds to assess relative osteogenic potency. Predictive analyses revealed nearly absolute correlation of the MSC\/hRUNX2-luc reporter platform to the <jats:italic>in vitro<\/jats:italic> classical functional assay of mineralization using murine BMMSCs. Validation using human BMMSCs with <jats:italic>in vitro<\/jats:italic> mineralization and <jats:italic>in vivo<\/jats:italic> osteogenesis assays also demonstrated nearly absolute correlation to the MSC\/hRUNX2-luc reporter results. Our findings therefore demonstrate that the MSC\/hRUNX2 reporter platform can accurately, rapidly, and robustly screen for candidate osteogenic compounds and thus be relevant for therapeutic application in OP.<\/jats:p>","DOI":"10.3389\/fcell.2020.607383","type":"journal-article","created":{"date-parts":[[2021,1,8]],"date-time":"2021-01-08T07:48:12Z","timestamp":1610092092000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":7,"title":["A Rapid and Highly Predictive in vitro Screening Platform for Osteogenic Natural Compounds Using Human Runx2 Transcriptional Activity in Mesenchymal Stem Cells"],"prefix":"10.3389","volume":"8","author":[{"given":"Li-Tzu","family":"Wang","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yu-Wei","family":"Lee","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chyi-Huey","family":"Bai","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hui-Chun","family":"Chiang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hsiu-Huan","family":"Wang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"B. Linju","family":"Yen","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Men-Luh","family":"Yen","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2021,1,8]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","DOI":"10.3390\/ijms20215386","article-title":"Substantial overview on mesenchymal stem cell biological and physical properties as an opportunity in translational medicine.","volume":"20","author":"Abdelrazik","year":"2019","journal-title":"Int. J. Mol. Sci."},{"key":"B2","doi-asserted-by":"publisher","first-page":"364","DOI":"10.1016\/s1470-2045(02)00777-5","article-title":"Phyto-oestrogens and cancer.","volume":"3","author":"Adlercreutz","year":"2002","journal-title":"Lancet Oncol."},{"key":"B3","doi-asserted-by":"publisher","first-page":"1683","DOI":"10.1002\/ijc.27788","article-title":"Phytoestrogen intake from foods, during adolescence and adulthood, and risk of breast cancer by estrogen and progesterone receptor tumor subgroup among Ontario women.","volume":"132","author":"Anderson","year":"2013","journal-title":"Int. J. Cancer"},{"key":"B4","doi-asserted-by":"publisher","DOI":"10.3389\/fcvm.2018.00183","article-title":"The interplay of SIRT1 and Wnt signaling in vascular calcification.","volume":"5","author":"Bartoli-Leonard","year":"2018","journal-title":"Front. Cardiovasc. Med."},{"key":"B5","doi-asserted-by":"crossref","DOI":"10.1056\/NEJMcp1513724","article-title":"Postmenopausal osteoporosis.","volume":"374","author":"Black","year":"2016","journal-title":"N. Engl. J. Med."},{"key":"B6","doi-asserted-by":"publisher","first-page":"465","DOI":"10.1359\/jbmr.061113","article-title":"Incidence and economic burden of osteoporosis-related fractures in the United States, 2005-2025.","volume":"22","author":"Burge","year":"2007","journal-title":"J. Bone Miner. Res."},{"key":"B7","doi-asserted-by":"publisher","first-page":"471","DOI":"10.1016\/j.spinee.2011.04.023","article-title":"A critical review of recombinant human bone morphogenetic protein-2 trials in spinal surgery: emerging safety concerns and lessons learned.","volume":"11","author":"Carragee","year":"2011","journal-title":"Spine J."},{"key":"B8","doi-asserted-by":"publisher","DOI":"10.3389\/fcell.2020.00197","article-title":"Human obesity induces dysfunction and early senescence in adipose tissue-derived Mesenchymal stromal\/stem cells.","volume":"8","author":"Conley","year":"2020","journal-title":"Front. Cell Dev. Biol."},{"key":"B9","doi-asserted-by":"publisher","first-page":"1277","DOI":"10.1007\/s00198-011-1601-6","article-title":"Secular trends in the incidence of hip and other osteoporotic fractures.","volume":"22","author":"Cooper","year":"2011","journal-title":"Osteoporos. Int."},{"key":"B10","doi-asserted-by":"publisher","first-page":"853","DOI":"10.1359\/jbmr.040120","article-title":"The balance between concurrent activation of ERs and PPARs determines daidzein-induced Osteogenesis and Adipogenesis.","volume":"19","author":"Dang","year":"2004","journal-title":"J. Bone. Miner. Res."},{"key":"B11","doi-asserted-by":"publisher","first-page":"207","DOI":"10.1016\/j.tem.2005.05.001","article-title":"Dose-dependent effects of phytoestrogens on bone.","volume":"16","author":"Dang","year":"2005","journal-title":"Trends Endocrinol. Metab."},{"key":"B12","doi-asserted-by":"publisher","first-page":"4223","DOI":"10.1074\/jbc.m110709200","article-title":"Mapping the ligand-binding sites and disease-associated mutations on the most abundant protein in the human, type I collagen.","volume":"277","author":"Di Lullo","year":"2002","journal-title":"J. Biol. Chem."},{"key":"B13","doi-asserted-by":"publisher","first-page":"747","DOI":"10.1016\/s0092-8674(00)80257-3","article-title":"Osf2\/Cbfa1: a transcriptional activator of Osteoblast differentiation.","volume":"89","author":"Ducy","year":"1997","journal-title":"Cell"},{"key":"B14","first-page":"431","article-title":"Molecular mechanisms of selective estrogen receptor modulator (SERM) action.","volume":"295","author":"Dutertre","year":"2000","journal-title":"J. Pharmacol. Exp. Ther."},{"key":"B15","doi-asserted-by":"publisher","first-page":"58","DOI":"10.7150\/ijbs.25783","article-title":"Pyrroloquinoline Quinone prevents estrogen deficiency-induced osteoporosis by inhibiting oxidative stress and osteocyte senescence.","volume":"15","author":"Geng","year":"2019","journal-title":"Int. J. Biol. Sci."},{"key":"B16","doi-asserted-by":"publisher","first-page":"77","DOI":"10.1016\/j.ab.2004.02.002","article-title":"An Alizarin red-based assay of mineralization by adherent cells in culture: comparison with cetylpyridinium chloride extraction.","volume":"329","author":"Gregory","year":"2004","journal-title":"Anal. Biochem."},{"key":"B17","doi-asserted-by":"publisher","first-page":"755","DOI":"10.1074\/jbc.r114.619957","article-title":"Transcriptional regulation of adipocyte differentiation: a central role for CCAAT\/enhancer-binding protein (C\/EBP) beta.","volume":"290","author":"Guo","year":"2015","journal-title":"J. Biol. Chem."},{"key":"B18","doi-asserted-by":"crossref","first-page":"693","DOI":"10.1016\/j.berh.2008.04.002","article-title":"Osteoporosis in developing countries.","volume":"22","author":"Handa","year":"2008","journal-title":"Best Pract. Res. Clin. Rheumatol."},{"key":"B19","doi-asserted-by":"publisher","first-page":"2155","DOI":"10.1007\/s00198-019-05104-5","article-title":"Determinants, consequences and potential solutions to poor adherence to anti-osteoporosis treatment: results of an expert group meeting organized by the European society for clinical and economic aspects of osteoporosis, osteoarthritis and musculoskeletal diseases (ESCEO) and the International Osteoporosis Foundation (IOF).","volume":"30","author":"Hiligsmann","year":"2019","journal-title":"Osteoporos. Int."},{"key":"B20","doi-asserted-by":"publisher","first-page":"727","DOI":"10.1016\/j.devcel.2005.02.013","article-title":"Canonical Wnt\/beta-catenin signaling prevents Osteoblasts from differentiating into Chondrocytes.","volume":"8","author":"Hill","year":"2005","journal-title":"Dev. Cell"},{"key":"B21","doi-asserted-by":"publisher","first-page":"1895","DOI":"10.1089\/ars.2012.4692","article-title":"H2O2 accumulation mediates differentiation capacity alteration, but not proliferative decline, in senescent human fetal mesenchymal stem cells.","volume":"18","author":"Ho","year":"2013","journal-title":"Antioxid. Redox Signal."},{"key":"B22","doi-asserted-by":"publisher","first-page":"375","DOI":"10.1016\/j.bbrc.2008.08.167","article-title":"Development of high-throughput screening system for osteogenic drugs using a cell-based sensor.","volume":"376","author":"Hojo","year":"2008","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"B23","doi-asserted-by":"publisher","first-page":"1158","DOI":"10.1093\/jnci\/djg015","article-title":"Phytoestrogen intake and endometrial cancer risk.","volume":"95","author":"Horn-Ross","year":"2003","journal-title":"J. Natl. Cancer Inst."},{"key":"B24","doi-asserted-by":"publisher","first-page":"605","DOI":"10.1001\/jama.280.7.605","article-title":"Randomized trial of estrogen plus progestin for secondary prevention of coronary heart disease in postmenopausal women. Heart and Estrogen\/progestin Replacement Study (HERS) Research Group.","volume":"280","author":"Hulley","year":"1998","journal-title":"JAMA"},{"key":"B25","doi-asserted-by":"publisher","first-page":"300","DOI":"10.1111\/joim.12586","article-title":"Excess mortality after hip fracture in elderly persons from Europe and the USA: the CHANCES project.","volume":"281","author":"Katsoulis","year":"2017","journal-title":"J. Intern. Med."},{"key":"B26","doi-asserted-by":"publisher","first-page":"755","DOI":"10.1016\/s0092-8674(00)80258-5","article-title":"Targeted disruption of Cbfa1 results in a complete lack of bone formation owing to maturational arrest of osteoblasts.","volume":"89","author":"Komori","year":"1997","journal-title":"Cell"},{"key":"B27","doi-asserted-by":"publisher","DOI":"10.3389\/fcell.2020.00316","article-title":"Limited potential or unfavorable manipulations? strategies toward efficient Mesenchymal Stem\/stromal cell applications.","volume":"8","author":"Lavrentieva","year":"2020","journal-title":"Front. Cell Dev. Biol."},{"key":"B28","doi-asserted-by":"publisher","first-page":"8783","DOI":"10.1128\/mcb.20.23.8783-8792.2000","article-title":"Runx2 is a common target of transforming growth factor beta1 and bone morphogenetic protein 2, and cooperation between Runx2 and Smad5 induces osteoblast-specific gene expression in the pluripotent mesenchymal precursor cell line C2C12.","volume":"20","author":"Lee","year":"2000","journal-title":"Mol. Cell. Biol."},{"key":"B29","doi-asserted-by":"publisher","first-page":"1251","DOI":"10.1177\/1087057109346446","article-title":"Identification of upregulators of BMP2 expression via high-throughput screening of a synthetic and natural compound library.","volume":"14","author":"Li","year":"2009","journal-title":"J. Biomol. Screen."},{"key":"B30","doi-asserted-by":"publisher","DOI":"10.3389\/fcell.2020.00258","article-title":"Senescence in Mesenchymal stem cells: functional alterations, molecular mechanisms, and rejuvenation strategies.","volume":"8","author":"Liu","year":"2020","journal-title":"Front. Cell Dev. Biol."},{"key":"B31","doi-asserted-by":"publisher","first-page":"2387","DOI":"10.7150\/thno.23620","article-title":"Resveratrol counteracts bone loss via mitofilin-mediated osteogenic improvement of mesenchymal stem cells in senescence-accelerated mice.","volume":"8","author":"Lv","year":"2018","journal-title":"Theranostics"},{"key":"B32","doi-asserted-by":"publisher","first-page":"1478","DOI":"10.1056\/nejmp1400104","article-title":"Comparative effectiveness questions in oncology.","volume":"370","author":"Mailankody","year":"2014","journal-title":"N. Engl. J. Med."},{"key":"B33","doi-asserted-by":"publisher","first-page":"43121","DOI":"10.1074\/jbc.m306531200","article-title":"Runx2 integrates estrogen activity in Osteoblasts.","volume":"278","author":"McCarthy","year":"2003","journal-title":"J. Biol. Chem."},{"key":"B34","doi-asserted-by":"publisher","first-page":"379","DOI":"10.1111\/j.1474-9728.2004.00127.x","article-title":"Aging activates adipogenic and suppresses osteogenic programs in mesenchymal marrow stroma\/stem cells: the role of PPAR-gamma2 transcription factor and TGF-beta\/BMP signaling pathways.","volume":"3","author":"Moerman","year":"2004","journal-title":"Aging Cell"},{"key":"B35","doi-asserted-by":"publisher","first-page":"177","DOI":"10.1016\/s8756-3282(00)00317-3","article-title":"Is there a therapeutic opportunity to either prevent or treat osteopenic disorders by inhibiting marrow adipogenesis?","volume":"27","author":"Nuttall","year":"2000","journal-title":"Bone"},{"key":"B36","doi-asserted-by":"publisher","first-page":"249","DOI":"10.1182\/blood-2014-04-572255","article-title":"Epigenetic and in vivo comparison of diverse MSC sources reveals an endochondral signature for human hematopoietic niche formation.","volume":"125","author":"Reinisch","year":"2015","journal-title":"Blood"},{"key":"B37","doi-asserted-by":"crossref","first-page":"595","DOI":"10.1056\/NEJMcp043801","article-title":"Clinical practice. Postmenopausal osteoporosis.","volume":"353","author":"Rosen","year":"2005","journal-title":"N. Engl. J. Med."},{"key":"B38","doi-asserted-by":"publisher","first-page":"321","DOI":"10.1001\/jama.288.3.321","article-title":"Risks and benefits of estrogen plus progestin in healthy postmenopausal women: principal results From the Women\u2019s Health Initiative randomized controlled trial.","volume":"288","author":"Rossouw","year":"2002","journal-title":"JAMA"},{"key":"B39","doi-asserted-by":"publisher","first-page":"252","DOI":"10.1016\/j.jsbmb.2012.12.006","article-title":"Effects of phytoestrogens and other plant-derived compounds on mesenchymal stem cells, bone maintenance and regeneration.","volume":"139","author":"Schilling","year":"2014","journal-title":"J. Steroid Biochem. Mol. Biol."},{"key":"B40","doi-asserted-by":"publisher","first-page":"2815","DOI":"10.1001\/jama.286.22.2815","article-title":"Identification and fracture outcomes of undiagnosed low bone mineral density in postmenopausal women: results from the national osteoporosis risk assessment.","volume":"286","author":"Siris","year":"2001","journal-title":"JAMA"},{"key":"B41","doi-asserted-by":"publisher","first-page":"771","DOI":"10.1016\/0092-8674(79)90317-9","article-title":"Multiple new phenotypes induced in 10T1\/2 and 3T3 cells treated with 5-azacytidine.","volume":"17","author":"Taylor","year":"1979","journal-title":"Cell"},{"key":"B42","doi-asserted-by":"publisher","first-page":"2552","DOI":"10.1002\/jbmr.460","article-title":"Resveratrol promotes osteogenesis of human mesenchymal stem cells by upregulating RUNX2 gene expression via the SIRT1\/FOXO3A axis.","volume":"26","author":"Tseng","year":"2011","journal-title":"J. Bone Miner. Res."},{"key":"B43","doi-asserted-by":"publisher","first-page":"5713","DOI":"10.1038\/sj.onc.1208919","article-title":"The disparate role of BMP in stem cell biology.","volume":"24","author":"Varga","year":"2005","journal-title":"Oncogene"},{"key":"B44","doi-asserted-by":"publisher","first-page":"15","DOI":"10.1007\/s13318-012-0112-y","article-title":"Isoflavones: estrogenic activity, biological effect and bioavailability.","volume":"38","author":"Vitale","year":"2013","journal-title":"Eur. J. Drug. Metab. Pharmacokinet."},{"key":"B45","doi-asserted-by":"publisher","first-page":"356","DOI":"10.1097\/bor.0b013e32832c6aa4","article-title":"Adherence to osteoporosis treatments: room for improvement.","volume":"21","author":"Warriner","year":"2009","journal-title":"Curr. Opin. Rheumatol."},{"key":"B46","doi-asserted-by":"publisher","DOI":"10.3390\/nu11122979","article-title":"Herbal medicines attenuate PD-L1 expression to induce anti-proliferation in obesity-related cancers.","volume":"11","author":"Yang","year":"2019","journal-title":"Nutrients"},{"key":"B47","doi-asserted-by":"publisher","first-page":"1529","DOI":"10.3727\/096368910x564067","article-title":"Efficient derivation and concise gene expression profiling of human embryonic stem cell-derived mesenchymal progenitors (EMPs).","volume":"20","author":"Yen","year":"2011","journal-title":"Cell Transpl."},{"key":"B48","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0178520","article-title":"SIRT1 is a positive regulator of the master osteoblast transcription factor, RUNX2.","volume":"12","author":"Zainabadi","year":"2017","journal-title":"PLoS One"},{"key":"B49","doi-asserted-by":"crossref","DOI":"10.1038\/s41368-019-0060-3","article-title":"Stem cell-based bone and dental regeneration: a view of microenvironmental modulation.","volume":"11","author":"Zheng","year":"2019","journal-title":"Int. J. Oral Sci."},{"key":"B50","doi-asserted-by":"publisher","first-page":"5482","DOI":"10.7150\/thno.28315","article-title":"Development of an accurate and proactive immunomodulatory strategy to improve bone substitute material-mediated Osteogenesis and Angiogenesis.","volume":"8","author":"Zheng","year":"2018","journal-title":"Theranostics"},{"key":"B51","doi-asserted-by":"publisher","first-page":"309","DOI":"10.7150\/ijbs.37162","article-title":"Fangchinoline protects against bone loss in OVX mice via inhibiting osteoclast formation, bone resorption and RANKL-induced signaling.","volume":"16","author":"Zhou","year":"2020","journal-title":"Int. J. Biol. Sci."}],"container-title":["Frontiers in Cell and Developmental Biology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fcell.2020.607383\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2021,1,18]],"date-time":"2021-01-18T16:24:20Z","timestamp":1610987060000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fcell.2020.607383\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,1,8]]},"references-count":51,"alternative-id":["10.3389\/fcell.2020.607383"],"URL":"https:\/\/doi.org\/10.3389\/fcell.2020.607383","relation":{},"ISSN":["2296-634X"],"issn-type":[{"value":"2296-634X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,1,8]]},"article-number":"607383"}}