{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,13]],"date-time":"2026-07-13T14:11:08Z","timestamp":1783951868916,"version":"3.55.0"},"reference-count":42,"publisher":"MDPI AG","issue":"19","license":[{"start":{"date-parts":[[2024,10,4]],"date-time":"2024-10-04T00:00:00Z","timestamp":1728000000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Singapore Ministry of Education","award":["A-8000983-01-00"],"award-info":[{"award-number":["A-8000983-01-00"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>5,6-dihydroxyflavone (5,6-DHF), a flavonoid that possesses potential anti-inflammatory and antioxidant activities owing to its special catechol motif on the A ring. However, its function and mechanism of action against inflammation and cellular oxidative stress have not been elucidated. In the current study, 5,6-DHF was observed inhibiting lipopolysaccharide (LPS)-induced nitric oxide (NO) and cytoplasmic reactive oxygen species (ROS) production with the IC50 of 11.55 \u00b1 0.64 \u03bcM and 0.8310 \u00b1 0.633 \u03bcM in murine macrophages, respectively. Meanwhile, 5,6-DHF suppressed the overexpression of pro-inflammatory mediators such as proteins and cytokines and eradicated the accumulation of mitochondrial ROS (mtROS). The blockage of the activation of cell surface toll-like receptor 4 (TLR4), impediment of the phosphorylation of c-Jun N-terminal kinase (JNK) and p38 from the mitogen-activated protein kinases (MAPK) pathway, Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) from the JAK-STAT pathway, and p65 from nuclear factor-\u03baB (NF-\u03baB) pathways were involved in the process of 5,6-DHF suppressing inflammation. Furthermore, 5,6-DHF acted as a cellular ROS scavenger and heme-oxygenase 1 (HO-1) inducer in relieving cellular oxidative stress. Importantly, 5,6-DHF exerted more potent anti-inflammatory activity than its close structural relatives, such as baicalein and chrysin. Overall, our findings pave the road for further research on 5,6-DHF in animal models.<\/jats:p>","DOI":"10.3390\/ijms251910694","type":"journal-article","created":{"date-parts":[[2024,10,4]],"date-time":"2024-10-04T06:00:57Z","timestamp":1728021657000},"page":"10694","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":7,"title":["Molecular Mechanism of 5,6-Dihydroxyflavone in Suppressing LPS-Induced Inflammation and Oxidative Stress"],"prefix":"10.3390","volume":"25","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-9984-3080","authenticated-orcid":false,"given":"Yujia","family":"Cao","sequence":"first","affiliation":[{"name":"Department of Food Science and Technology, National University of Singapore, Singapore 117542, Singapore"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2278-4497","authenticated-orcid":false,"given":"Yee-Joo","family":"Tan","sequence":"additional","affiliation":[{"name":"Infectious Diseases Translational Research Program, Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117545, Singapore"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2305-3960","authenticated-orcid":false,"given":"Dejian","family":"Huang","sequence":"additional","affiliation":[{"name":"Department of Food Science and Technology, National University of Singapore, Singapore 117542, Singapore"},{"name":"National University of Singapore (Suzhou) Research Institute, 377 Linquan Street, Suzhou 215123, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2024,10,4]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Gusev, E., and Zhuravleva, Y. (2022). Inflammation: A new look at an old problem. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms23094596"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"203","DOI":"10.1515\/hsz-2013-0241","article-title":"The role of oxidative stress during inflammatory processes","volume":"395","author":"Lugrin","year":"2014","journal-title":"Biol. Chem."},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"Leyane, T.S., Jere, S.W., and Houreld, N.N. (2022). Oxidative stress in ageing and chronic degenerative pathologies: Molecular mechanisms involved in counteracting oxidative stress and chronic inflammation. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms23137273"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1984742","DOI":"10.1155\/2022\/1984742","article-title":"Oxidative stress and intervertebral disc degenersation: Pathophysiology, signaling pathway, and therapy","volume":"2022","author":"Li","year":"2022","journal-title":"Oxid. Med. Cell. Longev."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"e47","DOI":"10.1017\/jns.2016.41","article-title":"Flavonoids: An overview","volume":"5","author":"Panche","year":"2016","journal-title":"J. Nutr. Sci."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"125124","DOI":"10.1016\/j.foodchem.2019.125124","article-title":"Anti-inflammatory effects of flavonoids","volume":"299","author":"Maleki","year":"2019","journal-title":"Food Chem."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Al-Khayri, J.M., Sahana, G.R., Nagella, P., Joseph, B.V., Alessa, F.M., and Al-Mssallem, M.Q. (2022). Flavonoids as potential anti-inflammatory molecules: A review. Molecules, 27.","DOI":"10.3390\/molecules27092901"},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Rakha, A., Umar, N., Rabail, R., Butt, M.S., Kieliszek, M., Hassoun, A., and Aadil, R.M. (2022). Anti-inflammatory and anti-allergic potential of dietary flavonoids: A review. Biomed. Pharmacother., 156.","DOI":"10.1016\/j.biopha.2022.113945"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"8788","DOI":"10.1021\/acs.jafc.2c03050","article-title":"Structure\u2013activity relationship (SAR) of flavones on their anti-inflammatory activity in murine macrophages in culture through the NF-\u03baB pathway and c-Src kinase receptor","volume":"70","author":"Wang","year":"2022","journal-title":"J. Agric. Food Chem."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"132531","DOI":"10.1016\/j.foodchem.2022.132531","article-title":"Plant flavonoids: Classification, distribution, biosynthesis, and antioxidant activity","volume":"383","author":"Shen","year":"2022","journal-title":"Food Chem."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"114671","DOI":"10.1016\/j.ejmech.2022.114671","article-title":"What about the progress in the synthesis of flavonoid from 2020?","volume":"243","author":"Liu","year":"2022","journal-title":"Eur. J. Med. Chem."},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Baker, W. (1939). 204. The synthesis of 5: 6-dihydroxyflavone and the structure of primetin. J. Am. Chem. Soc., 956\u2013961.","DOI":"10.1039\/jr9390000956"},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Fan, H., Liu, S., Shen, W., Kang, A., Tan, J., Li, L., Liu, X., Xu, C., Xu, X., and Lai, Y. (2020). Identification of the absorbed components and metabolites of Xiao-Ai-Jie-Du decoction and their distribution in rats using ultra high-performance liquid chromatography\/quadrupole time-of-flight mass spectrometry. J. Pharm. Biomed. Anal., 179.","DOI":"10.1016\/j.jpba.2019.112984"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"9293","DOI":"10.1039\/D3SC00752A","article-title":"Designing multi-target-directed flavonoids: A strategic approach to Alzheimer\u2019s disease","volume":"14","author":"Park","year":"2023","journal-title":"Chem. Sci."},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Lombardo, E., Sabellico, C., H\u00e1jek, J., Sta\u0148kov\u00e1, V., Filipsk\u00fd, T., Balducci, V., De Vito, P., Leone, S., Bavavea, E.I., and Silvestri, I.P. (2013). Protection of cells against oxidative stress by nanomolar levels of hydroxyflavones indicates a new type of intracellular antioxidant mechanism. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0060796"},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Behl, T., Rana, T., Alotaibi, G.H., Shamsuzzaman, M., Naqvi, M., Sehgal, A., Singh, S., Sharma, N., Almoshari, Y., and Abdellatif, A.A. (2022). Polyphenols inhibiting MAPK signalling pathway mediated oxidative stress and inflammation in depression. Biomed. Pharmacother., 146.","DOI":"10.1016\/j.biopha.2021.112545"},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Wang, X., Cao, Y., Chen, S., Yang, X., Bian, J., and Huang, D. (2023). Antioxidant and anti-inflammatory effects of 6, 3\u2032, 4\u2032-and 7, 3\u2032, 4\u2032-Trihydroxyflavone on 2D and 3D RAW264. 7 Models. Antioxidants, 12.","DOI":"10.3390\/antiox12010204"},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Li, G., Ding, K., Qiao, Y., Zhang, L., Zheng, L., Pan, T., and Zhang, L. (2020). Flavonoids regulate inflammation and oxidative stress in cancer. Molecules, 25.","DOI":"10.3390\/molecules25235628"},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Shamsudin, N.F., Ahmed, Q.U., Mahmood, S., Shah, S.A.A., Sarian, M.N., Khattak, M.M.A.K., Khatib, A., Sabere, A.S.M., Yusoff, Y.M., and Latip, J. (2022). Flavonoids as antidiabetic and anti-inflammatory agents: A review on structural activity relationship-based studies and meta-analysis. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms232012605"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"2853","DOI":"10.1016\/j.cell.2022.06.010","article-title":"Nitric oxide signaling in health and disease","volume":"185","author":"Lundberg","year":"2022","journal-title":"Cell"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"10","DOI":"10.1016\/j.brainresbull.2011.10.004","article-title":"Role of pro-inflammatory cytokines released from microglia in neurodegenerative diseases","volume":"87","author":"Smith","year":"2012","journal-title":"Brain Res. Bull."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Zhang, H., and Dhalla, N.S. (2024). The role of pro-Inflammatory cytokines in the pathogenesis of cardiovascular disease. Int. J. Mol. Sci., 25.","DOI":"10.3390\/ijms25021082"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"887","DOI":"10.1038\/nrc2507","article-title":"Cytokines and their relationship to the symptoms and outcome of cancer","volume":"8","author":"Seruga","year":"2008","journal-title":"Nat. Rev. Cancer."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1233","DOI":"10.1007\/s00018-020-03656-y","article-title":"TLR4 and CD14 trafficking and its influence on LPS-induced pro-inflammatory signaling","volume":"78","author":"Ciesielska","year":"2021","journal-title":"Cell. Mol. Life Sci."},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Behl, T., Upadhyay, T., Singh, S., Chigurupati, S., Alsubayiel, A.M., Mani, V., Vargas-De-La-Cruz, C., Uivarosan, D., Bustea, C., and Sava, C. (2021). Polyphenols targeting MAPK mediated oxidative stress and inflammation in rheumatoid arthritis. Molecules, 26.","DOI":"10.3390\/molecules26216570"},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Paracha, R.Z., Ahmad, J., Ali, A., Hussain, R., Niazi, U., Tareen, S.H.K., and Aslam, B. (2014). Formal modelling of toll like receptor 4 and JAK\/STAT signalling pathways: Insight into the roles of SOCS-1, interferon-\u03b2 and proinflammatory cytokines in sepsis. PLoS ONE, 9.","DOI":"10.1371\/journal.pone.0108466"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"287","DOI":"10.1016\/j.gendis.2020.06.005","article-title":"The Nuclear Factor Kappa B (NF-kB) signaling in cancer development and immune diseases","volume":"8","author":"Zinatizadeh","year":"2021","journal-title":"Genes Dis."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"271","DOI":"10.3892\/ijmm.2017.3036","article-title":"The crucial role of protein phosphorylation in cell signaling and its use as targeted therapy","volume":"40","author":"Ardito","year":"2017","journal-title":"Int. J. Mol. Med."},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Tirichen, H., Yaigoub, H., Xu, W., Wu, C., Li, R., and Li, Y. (2021). Mitochondrial reactive oxygen species and their contribution in chronic kidney disease progression through oxidative stress. Front. Physiol., 12.","DOI":"10.3389\/fphys.2021.627837"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"90","DOI":"10.1111\/j.2040-1124.2010.00013.x","article-title":"Oxidative stress: A cause and therapeutic target of diabetic complications","volume":"1","author":"Araki","year":"2010","journal-title":"J. Diabetes Investig."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"109","DOI":"10.1016\/j.freeradbiomed.2021.02.043","article-title":"Mitochondrial ROS and mitochondria-targeted antioxidants in the aged heart","volume":"167","author":"Kaludercic","year":"2021","journal-title":"Free Radic. Biol. Med."},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Casey, A.M., and Murphy, M.P. (2022). Uncovering the source of mitochondrial superoxide in pro-inflammatory macrophages: Insights from immunometabolism. Biochim. Biophys. Acta-Mol. Basis Dis, 1868.","DOI":"10.1016\/j.bbadis.2022.166481"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"909","DOI":"10.1152\/physrev.00026.2013","article-title":"Mitochondrial reactive oxygen species (ROS) and ROS-induced ROS release","volume":"94","author":"Zorov","year":"2014","journal-title":"Physiol. Rev."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"792639","DOI":"10.1155\/2011\/792639","article-title":"Mitogen-activated protein kinases and reactive oxygen species: How can ROS activate MAPK pathways?","volume":"2011","author":"Son","year":"2011","journal-title":"J. Signal Transduct."},{"key":"ref_35","doi-asserted-by":"crossref","unstructured":"Liu, J., Wang, F., and Luo, F. (2023). The role of JAK\/STAT pathway in fibrotic diseases: Molecular and cellular mechanisms. Biomolecules, 13.","DOI":"10.3390\/biom13010119"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"81","DOI":"10.1016\/j.cotox.2017.11.002","article-title":"NF-\u03baB in oxidative stress","volume":"7","author":"Lingappan","year":"2018","journal-title":"Curr. Opin. Toxicol."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"253","DOI":"10.1248\/bpb.28.253","article-title":"Cytotoxicity of flavonoids toward cultured normal human cells","volume":"28","author":"Matsuo","year":"2005","journal-title":"Biol. Pharm. Bull."},{"key":"ref_38","doi-asserted-by":"crossref","unstructured":"Zhai, K., Mazurakova, A., Koklesova, L., Kubatka, P., and B\u00fcsselberg, D. (2021). Flavonoids synergistically enhance the anti-glioblastoma effects of chemotherapeutic drugs. Biomolecules, 11.","DOI":"10.3390\/biom11121841"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"1446","DOI":"10.1080\/14786419.2017.1346637","article-title":"In vitro antioxidant\/prooxidant effects of combined use of flavonoids","volume":"32","author":"Ince","year":"2018","journal-title":"Nat. Prod. Res."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"289","DOI":"10.2174\/1568009621666210203111220","article-title":"Flavonoids overcome drug resistance to cancer chemotherapy by epigenetically modulating multiple mechanisms","volume":"21","author":"To","year":"2021","journal-title":"Curr. Cancer Drug Targets"},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"More, G.K., and Makola, R.T. (2020). In-vitro analysis of free radical scavenging activities and suppression of LPS-induced ROS production in macrophage cells by Solanum sisymbriifolium extracts. Sci. Rep., 10.","DOI":"10.1038\/s41598-020-63491-w"},{"key":"ref_42","doi-asserted-by":"crossref","unstructured":"Jiang, C., Zhang, J., Xie, H., Guan, H., Li, R., Chen, C., Dong, H., Zhou, Y., and Zhang, W. (2022). Baicalein suppresses lipopolysaccharide-induced acute lung injury by regulating Drp1-dependent mitochondrial fission of macrophages. Biomed. Pharmacother., 145.","DOI":"10.1016\/j.biopha.2021.112408"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/25\/19\/10694\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T16:10:31Z","timestamp":1760112631000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/25\/19\/10694"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,10,4]]},"references-count":42,"journal-issue":{"issue":"19","published-online":{"date-parts":[[2024,10]]}},"alternative-id":["ijms251910694"],"URL":"https:\/\/doi.org\/10.3390\/ijms251910694","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2024,10,4]]}}}