{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,18]],"date-time":"2026-03-18T20:47:53Z","timestamp":1773866873844,"version":"3.50.1"},"reference-count":60,"publisher":"MDPI AG","issue":"7","license":[{"start":{"date-parts":[[2021,7,3]],"date-time":"2021-07-03T00:00:00Z","timestamp":1625270400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"FCT (Funda\u00e7\u00e3o para a Ci\u00eancia e a Tecnologia) and MCTES through national funds and \u201cPrograma Operacional Competitividade e Internacionaliza\u00e7\u00e3o\u201d (COMPETE)","award":["PTDC\/MED-QUI\/29243\/2017- POCI-01-0145-FEDER-029243"],"award-info":[{"award-number":["PTDC\/MED-QUI\/29243\/2017- POCI-01-0145-FEDER-029243"]}]},{"name":"FCT\/MCTES, Funda\u00e7\u00e3o para a Ci\u00eancia e Tecnologia and Minist\u00e9rio da Ci\u00eancia, Tecnologia e Ensino Superior","award":["UIDB\/50006\/2020"],"award-info":[{"award-number":["UIDB\/50006\/2020"]}]},{"name":"FCT\/MCTES","award":["UIDP\/50017\/2020"],"award-info":[{"award-number":["UIDP\/50017\/2020"]}]},{"name":"FCT\/MCTES","award":["UIDP\/50017\/2020"],"award-info":[{"award-number":["UIDP\/50017\/2020"]}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e a Tecnologia","doi-asserted-by":"publisher","award":["CEECIND\/04050\/2017"],"award-info":[{"award-number":["CEECIND\/04050\/2017"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e a Tecnologia","doi-asserted-by":"publisher","award":["SFRH\/BPD\/74868\/2010"],"award-info":[{"award-number":["SFRH\/BPD\/74868\/2010"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pharmaceuticals"],"abstract":"<jats:p>New agents are demanded to increase the therapeutic options for osteosarcoma (OS). Although OS is the most common bone cancer in children and adolescents, it is considered a rare disorder. Therefore, finding adjuvant drugs has potential to advance therapy for this disease. In this study, 3\u2032,4\u2032-dihydroxyflavonol (DiOHF) was investigated to assess the effects in OS cellular models in combination with doxorubicin (Dox). MG-63 and U2OS human OS cells were exposed to DiOHF and Dox and tested for cell viability and growth. To elucidate the inhibitory effects of DiOHF, additional studies were conducted to assess apoptosis and cell cycle distribution, gene expression quantification of cell cycle regulators, and cytokinesis-block cytome assay to determine nuclear division rate. DiOHF decreased OS cell growth and viability in a concentration-dependent manner. Its combination with Dox enabled Dox dose reduction in both cell lines, with synergistic interactions in U2OS cells. Although no significant apoptotic effects were detected at low concentrations, cytostatic effects were demonstrated in both cell lines. Incubation with DiOHF altered cell cycle dynamics and resulted in differential cyclin and cyclin-dependent kinase expression. Overall, this study presents an antiproliferative action of DiOHF in OS combination therapy via modulation of the cell cycle and nuclear division.<\/jats:p>","DOI":"10.3390\/ph14070640","type":"journal-article","created":{"date-parts":[[2021,7,4]],"date-time":"2021-07-04T22:34:36Z","timestamp":1625438076000},"page":"640","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":6,"title":["3\u2032,4\u2032-Dihydroxyflavonol Modulates the Cell Cycle in Cancer Cells: Implication as a Potential Combination Drug in Osteosarcoma"],"prefix":"10.3390","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3883-0806","authenticated-orcid":false,"given":"Jos\u00e9 Miguel P.","family":"Ferreira de Oliveira","sequence":"first","affiliation":[{"name":"LAQV, REQUIMTE, Laboratory of Applied Chemistry, Department of Chemical Sciences, Faculty of Pharmacy, University of Porto, 4050-313 Porto, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2325-5439","authenticated-orcid":false,"given":"Joana Filipa D.","family":"Almeida","sequence":"additional","affiliation":[{"name":"Department of Biology, Faculty of Sciences, University of Porto, 4169-007 Porto, Portugal"}]},{"given":"Maria","family":"Martins","sequence":"additional","affiliation":[{"name":"Department of Biology, Faculty of Sciences, University of Porto, 4169-007 Porto, Portugal"}]},{"given":"Carina","family":"Proen\u00e7a","sequence":"additional","affiliation":[{"name":"LAQV, REQUIMTE, Laboratory of Applied Chemistry, Department of Chemical Sciences, Faculty of Pharmacy, University of Porto, 4050-313 Porto, Portugal"}]},{"given":"Helena","family":"Oliveira","sequence":"additional","affiliation":[{"name":"Department of Biology & CESAM, University of Aveiro, 3810-193 Aveiro, Portugal"}]},{"given":"Eduarda","family":"Fernandes","sequence":"additional","affiliation":[{"name":"LAQV, REQUIMTE, Laboratory of Applied Chemistry, Department of Chemical Sciences, Faculty of Pharmacy, University of Porto, 4050-313 Porto, Portugal"}]},{"given":"Concei\u00e7\u00e3o","family":"Santos","sequence":"additional","affiliation":[{"name":"Department of Biology, Faculty of Sciences, University of Porto, 4169-007 Porto, Portugal"},{"name":"LAQV, REQUIMTE, Faculty of Sciences, University of Porto, 4169-007 Porto, Portugal"}]}],"member":"1968","published-online":{"date-parts":[[2021,7,3]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"480","DOI":"10.1038\/nrendo.2017.16","article-title":"Germline and somatic genetics of osteosarcoma\u2014Connecting aetiology, biology and therapy","volume":"13","author":"Gianferante","year":"2017","journal-title":"Nat. Rev. Endocrinol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.anndiagpath.2020.151654","article-title":"Osteosarcoma: A comprehensive review of management and treatment strategies","volume":"49","author":"Jafari","year":"2020","journal-title":"Ann. Diagn. Pathol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1186\/s12938-021-00860-0","article-title":"Osteosarcoma: A review of current and future therapeutic approaches","volume":"20","author":"Zhao","year":"2021","journal-title":"Biomed. Eng. Online"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"221","DOI":"10.1007\/s40744-016-0046-y","article-title":"Review of osteosarcoma and current management","volume":"3","author":"Durfee","year":"2016","journal-title":"Rheumatol. Ther."},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Gazouli, I., Kyriazoglou, A., Kotsantis, I., Anastasiou, M., Pantazopoulos, A., Prevezanou, M., Chatzidakis, I., Kavourakis, G., Economopoulou, P., and Kontogeorgakos, V. (2021). Systematic review of recurrent osteosarcoma systemic therapy. Cancers, 13.","DOI":"10.3390\/cancers13081757"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"7","DOI":"10.3322\/caac.21654","article-title":"Cancer statistics, 2021","volume":"71","author":"Siegel","year":"2021","journal-title":"CA Cancer J. Clin."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"495","DOI":"10.1517\/14728214.2015.1051965","article-title":"Advances in emerging drugs for osteosarcoma","volume":"20","author":"Hattinger","year":"2015","journal-title":"Exp. Opin. Emerg. Drugs"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"106","DOI":"10.1002\/med.21280","article-title":"Doxorubicin-induced cardiotoxicity: From bioenergetic failure and cell death to cardiomyopathy","volume":"34","author":"Carvalho","year":"2014","journal-title":"Med. Res. Rev."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"38022","DOI":"10.18632\/oncotarget.16723","article-title":"Combination therapy in combating cancer","volume":"8","author":"Homayouni","year":"2017","journal-title":"Oncotarget"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"298","DOI":"10.1016\/j.drudis.2011.02.016","article-title":"In silico repositioning of approved drugs for rare and neglected diseases","volume":"16","author":"Ekins","year":"2011","journal-title":"Drug Discov. Today"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"41","DOI":"10.1038\/nrd.2018.168","article-title":"Drug repurposing: Progress, challenges and recommendations","volume":"18","author":"Pushpakom","year":"2019","journal-title":"Nat. Rev. Drug Discov."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"513","DOI":"10.1080\/21678707.2018.1500690","article-title":"Omics-driven drug repurposing as a source of innovative therapies in rare cancers","volume":"6","author":"Pantziarka","year":"2018","journal-title":"Exp. Opin. Orphan Drugs"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1183","DOI":"10.1007\/s11101-017-9529-x","article-title":"Molecular signaling mechanisms behind polyphenol-induced bone anabolism","volume":"16","author":"Torre","year":"2017","journal-title":"Phytochem. Rev."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"357","DOI":"10.1007\/s10616-014-9694-3","article-title":"Apigenin inhibits osteoblastogenesis and osteoclastogenesis and prevents bone loss in ovariectomized mice","volume":"67","author":"Goto","year":"2015","journal-title":"Cytotechnology"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"8","DOI":"10.1016\/j.jnutbio.2009.11.002","article-title":"The effects of luteolin on osteoclast differentiation, function in vitro and ovariectomy-induced bone loss","volume":"22","author":"Kim","year":"2011","journal-title":"J. Nutr. Biochem."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1239","DOI":"10.1002\/mnfr.201300836","article-title":"The flavonoid fisetin promotes osteoblasts differentiation through Runx2 transcriptional activity","volume":"58","author":"Davicco","year":"2014","journal-title":"Mol. Nutr. Food Res."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"59","DOI":"10.1016\/j.bcp.2014.10.019","article-title":"Myricetin prevents titanium particle-induced osteolysis in vivo and inhibits RANKL-induced osteoclastogenesis in vitro","volume":"93","author":"Wu","year":"2015","journal-title":"Biochem. Pharmacol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1993","DOI":"10.2174\/1381612823666170214115718","article-title":"Flavonoids active against osteosarcoma: A review of the molecular mechanisms involved","volume":"23","author":"Liu","year":"2017","journal-title":"Curr. Pharm. Des."},{"key":"ref_19","first-page":"3239","article-title":"Plant flavonoid taxifolin inhibits the growth, migration and invasion of human osteosarcoma cells","volume":"17","author":"Chen","year":"2018","journal-title":"Mol. Med. Rep."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1155\/2018\/4296985","article-title":"Citrus aurantium naringenin prevents osteosarcoma progression and recurrence in the patients who underwent osteosarcoma surgery by improving antioxidant capability","volume":"2018","author":"Zhang","year":"2018","journal-title":"Oxid. Med. Cell. Longev."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.biopha.2019.108839","article-title":"Quercetin suppresses the proliferation and metastasis of metastatic osteosarcoma cells by inhibiting parathyroid hormone receptor 1","volume":"114","author":"Li","year":"2019","journal-title":"Biomed. Pharmacother."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"5077","DOI":"10.2147\/DDDT.S275148","article-title":"Acacetin induces apoptosis in human osteosarcoma cells by modulation of ROS\/JNK activation","volume":"14","author":"Wang","year":"2020","journal-title":"Drug Des. Devel. Ther."},{"key":"ref_23","first-page":"1","article-title":"Flavonoid compound breviscapine suppresses human osteosarcoma Saos-2 progression property and induces apoptosis by regulating mitochondria-dependent pathway","volume":"35","author":"Wang","year":"2021","journal-title":"J. Biochem. Mol. Toxicol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1425","DOI":"10.1016\/j.fct.2010.04.007","article-title":"New advances in molecular mechanisms and the prevention of adriamycin toxicity by antioxidant nutrients","volume":"48","author":"Quiles","year":"2010","journal-title":"Food Chem. Toxicol."},{"key":"ref_25","first-page":"15190","article-title":"The flavonoid luteolin enhances doxorubicin-induced autophagy in human osteosarcoma U2OS cells","volume":"8","author":"Zhang","year":"2015","journal-title":"Int. J. Clin. Exp. Med."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1017\/jns.2016.41","article-title":"Flavonoids: An overview","volume":"5","author":"Panche","year":"2016","journal-title":"J. Nutr. Sci."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"786","DOI":"10.1111\/j.1440-1681.2004.04072.x","article-title":"Vascular and anti-oxidant actions of flavonols and flavones","volume":"31","author":"Woodman","year":"2004","journal-title":"Clin. Exp. Pharmacol. Physiol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"39","DOI":"10.4103\/ijp.IJP_727_20","article-title":"Increased apoptosis, tumor necrosis factor-\u03b1, and DNA damage attenuated by 3\u2019,4\u2019-dihydroxyflavonol in rats with brain \u0130schemia-reperfusion","volume":"53","author":"Dasdelen","year":"2021","journal-title":"Indian J. Pharmacol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"350","DOI":"10.1002\/biof.111","article-title":"Effects of resveratrol and flavonols on cardiovascular function: Physiological mechanisms","volume":"36","author":"Yap","year":"2010","journal-title":"Biofactors"},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"(2017). Rationalizing combination therapies. Nat. Med., 23, 1113.","DOI":"10.1038\/nm.4426"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"803","DOI":"10.1111\/j.1440-1681.2010.05384.x","article-title":"Effects of 3\u2019,4\u2019-dihydroxyflavonol on vascular contractions of rat aortic rings","volume":"37","author":"Song","year":"2010","journal-title":"Clin. Exp. Pharmacol. Physiol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1371\/journal.pone.0020813","article-title":"3\u2032,4\u2032-Dihydroxyflavonol reduces superoxide and improves nitric oxide function in diabetic rat mesenteric arteries","volume":"6","author":"Leo","year":"2011","journal-title":"PLoS ONE"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"378","DOI":"10.1002\/biof.1033","article-title":"Trihydroxyflavones with antioxidant and anti-inflammatory efficacy","volume":"38","author":"Gomes","year":"2012","journal-title":"Biofactors"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"673","DOI":"10.2217\/fon-2016-0519","article-title":"Doxorubicin-resistant osteosarcoma: Novel therapeutic approaches in sight?","volume":"13","author":"Hattinger","year":"2017","journal-title":"Future Oncol."},{"key":"ref_35","doi-asserted-by":"crossref","unstructured":"Prudowsky, Z.D., and Yustein, J.T. (2020). Recent insights into therapy resistance in osteosarcoma. Cancers, 13.","DOI":"10.3390\/cancers13010083"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1186\/s13046-018-0689-3","article-title":"SOX2OT variant 7 contributes to the synergistic interaction between EGCG and Doxorubicin to kill osteosarcoma via autophagy and stemness inhibition","volume":"37","author":"Wang","year":"2018","journal-title":"J. Exp. Clin. Cancer Res."},{"key":"ref_37","first-page":"1","article-title":"The kinetics of G2 and M transitions regulated by B cyclins","volume":"8","author":"Huang","year":"2013","journal-title":"PLoS ONE"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1186\/1747-1028-5-11","article-title":"An overview of Cdk1-controlled targets and processes","volume":"5","author":"Enserink","year":"2010","journal-title":"Cell Div."},{"key":"ref_39","first-page":"1365","article-title":"Effect of quercetin on cell cycle and cyclin expression in ovarian carcinoma and osteosarcoma cell lines","volume":"10","author":"Catanzaro","year":"2015","journal-title":"Nat. Prod. Commun."},{"key":"ref_40","first-page":"1297","article-title":"Evaluation of quercetin as a potential drug in osteosarcoma treatment","volume":"33","author":"Berndt","year":"2013","journal-title":"Anticancer Res."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"435","DOI":"10.3892\/or.2016.5224","article-title":"Galangin suppresses human osteosarcoma cells: An exploration of its underlying mechanism","volume":"37","author":"Yang","year":"2017","journal-title":"Oncol. Rep."},{"key":"ref_42","first-page":"185","article-title":"Apigenin induces cell cycle arrest and p21\/WAF1 expression in a p53-independent pathway","volume":"26","author":"Takagaki","year":"2005","journal-title":"Int. J. Oncol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1177\/1010428317705745","article-title":"Icarisid II inhibits the proliferation of human osteosarcoma cells by inducing apoptosis and cell cycle arrest","volume":"39","author":"Tang","year":"2017","journal-title":"Tumor Biol."},{"key":"ref_44","first-page":"735","article-title":"Pelargonidin induces antitumor effects in human osteosarcoma cells via autophagy induction, loss of mitochondrial membrane potential, G2\/M cell cycle arrest and downregulation of PI3K\/AKT signalling pathway","volume":"23","author":"Chen","year":"2018","journal-title":"J. BUON"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"7617","DOI":"10.3892\/mmr.2017.7506","article-title":"Investigation of crucial genes and microRNAs in conventional osteosarcoma using gene expression profiling analysis","volume":"16","author":"Peng","year":"2017","journal-title":"Mol. Med. Rep."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"1479","DOI":"10.1007\/s10147-019-01500-3","article-title":"CDC20 and its downstream genes: Potential prognosis factors of osteosarcoma","volume":"24","author":"Wu","year":"2019","journal-title":"Int. J. Clin. Oncol."},{"key":"ref_47","first-page":"7538","article-title":"Exploration and validation of downregulated microRNA-199a-3p, downstream messenger RNA targets and transcriptional regulation in osteosarcoma","volume":"11","author":"Huang","year":"2019","journal-title":"Am. J. Transl. Res."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"844","DOI":"10.1159\/000487178","article-title":"LncRNA GAS5 Represses osteosarcoma cells growth and metastasis via sponging MiR-203a","volume":"45","author":"Wang","year":"2018","journal-title":"Cell. Physiol. Biochem."},{"key":"ref_49","doi-asserted-by":"crossref","unstructured":"Chen, C.Y., Chen, C.C., Shieh, T.M., Hsueh, C., Wang, S.H., Leu, Y.L., Lian, J.H., and Wang, T.H. (2018). Corylin suppresses hepatocellular carcinoma progression via the inhibition of epithelial-mesenchymal transition, mediated by long noncoding RNA GAS5. Int. J. Mol. Sci., 19.","DOI":"10.3390\/ijms19020380"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"713","DOI":"10.3892\/mmr.2015.4597","article-title":"Fibroblast growth factor receptor 1 promotes MG63 cell proliferation and is associated with increased expression of cyclin-dependent kinase 1 in osteosarcoma","volume":"13","author":"Zhou","year":"2016","journal-title":"Mol. Med. Rep."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"1713","DOI":"10.1016\/j.cellsig.2012.04.011","article-title":"The anti-tumor role and mechanism of integrated and truncated PDCD5 proteins in osteosarcoma cells","volume":"24","author":"Han","year":"2012","journal-title":"Cell. Signal."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"937","DOI":"10.1158\/1541-7786.MCR-07-2115","article-title":"Cell cycle regulator gene CDC5L, a potential target for 6p12-p21 amplicon in osteosarcoma","volume":"6","author":"Lu","year":"2008","journal-title":"Mol. Cancer Res."},{"key":"ref_53","first-page":"10451","article-title":"CDC5L contributes to malignant cell proliferation in human osteosarcoma via cell cycle regulation","volume":"9","author":"Wang","year":"2016","journal-title":"Int. J. Clin. Exp. Pathol."},{"key":"ref_54","first-page":"1","article-title":"Paclitaxel and the dietary flavonoid fisetin: A synergistic combination that induces mitotic catastrophe and autophagic cell death in A549 non-small cell lung cancer cells","volume":"16","author":"Tadrowski","year":"2016","journal-title":"Cancer Cell Int."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"1032","DOI":"10.1093\/carcin\/bgp101","article-title":"Dietary flavonoid fisetin induces a forced exit from mitosis by targeting the mitotic spindle checkpoint","volume":"30","author":"Salmela","year":"2009","journal-title":"Carcinogenesis"},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"36","DOI":"10.1016\/j.dnarep.2016.12.006","article-title":"Hesperetin-etoposide combinations induce cytotoxicity in U2OS cells: Implications on therapeutic developments for osteosarcoma","volume":"50","author":"Coutinho","year":"2017","journal-title":"DNA Repair"},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"621","DOI":"10.1124\/pr.58.3.10","article-title":"Theoretical basis, experimental design, and computerized simulation of synergism and antagonism in drug combination studies","volume":"58","author":"Chou","year":"2006","journal-title":"Pharmacol. Rev."},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"1084","DOI":"10.1038\/nprot.2007.77","article-title":"Cytokinesis-block micronucleus cytome assay","volume":"2","author":"Fenech","year":"2007","journal-title":"Nat. Protoc."},{"key":"ref_59","first-page":"365","article-title":"Primer3 on the WWW for general users and for biologist programmers","volume":"132","author":"Rozen","year":"2000","journal-title":"Methods Mol. Biol."},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"402","DOI":"10.1006\/meth.2001.1262","article-title":"Analysis of relative gene expression data using real-time quantitative PCR and the 2(-delta delta C(T)) method","volume":"25","author":"Livak","year":"2001","journal-title":"Methods"}],"container-title":["Pharmaceuticals"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1424-8247\/14\/7\/640\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T06:25:32Z","timestamp":1760163932000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1424-8247\/14\/7\/640"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,7,3]]},"references-count":60,"journal-issue":{"issue":"7","published-online":{"date-parts":[[2021,7]]}},"alternative-id":["ph14070640"],"URL":"https:\/\/doi.org\/10.3390\/ph14070640","relation":{},"ISSN":["1424-8247"],"issn-type":[{"value":"1424-8247","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,7,3]]}}}