{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,2,21]],"date-time":"2025-02-21T18:53:06Z","timestamp":1740163986224,"version":"3.37.3"},"reference-count":36,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2024,2,19]],"date-time":"2024-02-19T00:00:00Z","timestamp":1708300800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2024,2,19]],"date-time":"2024-02-19T00:00:00Z","timestamp":1708300800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"funder":[{"DOI":"10.13039\/100020595","name":"National Science and Technology Council","doi-asserted-by":"publisher","award":["MOST 109-2313-B-020-005"],"award-info":[{"award-number":["MOST 109-2313-B-020-005"]}],"id":[{"id":"10.13039\/100020595","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Sci Rep"],"abstract":"<jats:title>Abstract<\/jats:title><jats:p>In the increasing demand for virus vaccines, large-scale production of safe, efficient, and economical viral antigens has become a significant challenge. High-cell-density manufacturing processes are the most commonly used to produce vaccine antigens and protein drugs. However, the cellular stress response in large-scale cell culture may directly affect host cell growth and metabolism, reducing antigen production and increasing production costs. This study provided a novel strategy of the antioxidant auxiliary system (AAS) to supply molecular hydrogen (H<jats:sub>2<\/jats:sub>) into the cell culture media via proton exchange membrane (PEM) electrolysis. Integrated with a high-density cell bioreactor, the AAS aims to alleviate cellular stress response and increase viral vaccine production. In the results, the AAS stably maintained H<jats:sub>2<\/jats:sub> concentration in media even in the high-air exposure tiding cell bioreactor. H<jats:sub>2<\/jats:sub> treatment was shown safe to cell culture and effectively alleviated oxidative stress. In two established virus cultures models, bovine epidemic fever virus (BEFV) and porcine circovirus virus type 2 (PCV-2), were employed to verify the efficacy of AAS. The virus yield was increased by 3.7 and 2.5 folds in BEFV and PCV-2 respectively. In conclusion, the AAS-connected bioreactor effectively alleviated cellular oxidative stress and enhanced virus production in high-density cell culture.<\/jats:p>","DOI":"10.1038\/s41598-024-54847-7","type":"journal-article","created":{"date-parts":[[2024,2,19]],"date-time":"2024-02-19T13:02:37Z","timestamp":1708347757000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["A strategy of novel molecular hydrogen-producing antioxidative auxiliary system improves virus production in cell bioreactor"],"prefix":"10.1038","volume":"14","author":[{"given":"Yu-Jing","family":"Zeng","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Min-Kung","family":"Hsu","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jia-Rong","family":"Cai","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hsian-Yu","family":"Wang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2024,2,19]]},"reference":[{"key":"54847_CR1","doi-asserted-by":"publisher","first-page":"2121","DOI":"10.1007\/s00253-015-7267-9","volume":"100","author":"F Tapia","year":"2016","unstructured":"Tapia, F., V\u00e1zquez-Ram\u00edrez, D., Genzel, Y. & Reichl, U. Bioreactors for high cell density and continuous multi-stage cultivations: Options for process intensification in cell culture-based viral vaccine production. Appl. Microbiol. Biotechnol. 100, 2121\u20132132 (2016).","journal-title":"Appl. Microbiol. Biotechnol."},{"key":"54847_CR2","doi-asserted-by":"publisher","first-page":"921755","DOI":"10.3389\/fbioe.2022.921755","volume":"10","author":"Z Fang","year":"2022","unstructured":"Fang, Z. et al. Application of bioreactor technology for cell culture-based viral vaccine production: Present status and future prospects. Front. Bioeng. Biotechnol. 10, 921755 (2022).","journal-title":"Front. Bioeng. Biotechnol."},{"key":"54847_CR3","doi-asserted-by":"publisher","first-page":"1181","DOI":"10.1586\/14760584.2015.1067144","volume":"14","author":"LE Gallo-Ram\u00edrez","year":"2015","unstructured":"Gallo-Ram\u00edrez, L. E., Nikolay, A., Genzel, Y. & Reichl, U. Bioreactor concepts for cell culture-based viral vaccine production. Expert Rev. Vaccines 14, 1181\u20131195 (2015).","journal-title":"Expert Rev. Vaccines"},{"key":"54847_CR4","doi-asserted-by":"publisher","first-page":"1172","DOI":"10.1002\/bit.27247","volume":"117","author":"V Chevallier","year":"2020","unstructured":"Chevallier, V., Andersen, M. R. & Malphettes, L. Oxidative stress-alleviating strategies to improve recombinant protein production in CHO cells. Biotechnol. Bioeng. 117, 1172\u20131186 (2020).","journal-title":"Biotechnol. Bioeng."},{"key":"54847_CR5","doi-asserted-by":"publisher","first-page":"e22000029","DOI":"10.1002\/biot.202200029","volume":"17","author":"D Komuczki","year":"2022","unstructured":"Komuczki, D. et al. High cysteine concentrations in cell culture media lead to oxidative stress and reduced bioprocess performance of recombinant CHO cells. Biotechnol. J. 17, e22000029 (2022).","journal-title":"Biotechnol. J."},{"key":"54847_CR6","doi-asserted-by":"publisher","first-page":"601","DOI":"10.3390\/vaccines9060601","volume":"9","author":"YJ Zeng","year":"2021","unstructured":"Zeng, Y. J. et al. A senescence-like cellular response inhibits bovine ephemeral fever virus proliferation. Vaccines (Basel) 9, 601 (2021).","journal-title":"Vaccines (Basel)"},{"key":"54847_CR7","doi-asserted-by":"publisher","first-page":"715","DOI":"10.1146\/annurev-biochem-061516-045037","volume":"86","author":"H Sies","year":"2017","unstructured":"Sies, H., Berndt, C. & Jones, D. P. Oxidative stress. Annu. Rev. Biochem. 86, 715\u2013748 (2017).","journal-title":"Annu. Rev. Biochem."},{"key":"54847_CR8","doi-asserted-by":"publisher","first-page":"122","DOI":"10.1016\/j.cotox.2018.01.002","volume":"7","author":"H Sies","year":"2018","unstructured":"Sies, H. On the history of oxidative stress: Concept and some aspects of current development. Curr. Opin. Toxicol. 7, 122\u2013126 (2018).","journal-title":"Curr. Opin. Toxicol."},{"key":"54847_CR9","doi-asserted-by":"publisher","first-page":"437","DOI":"10.3390\/oxygen2040030","volume":"2","author":"AK Aranda-Rivera","year":"2022","unstructured":"Aranda-Rivera, A. K., Cruz-Gregorio, A., Arancibia-Hern\u00e1ndez, Y. L., Hern\u00e1ndez-Cruz, E. Y. & Pedraza-Chaverri, J. RONS and oxidative stress: An overview of basic concepts. Oxygen 2, 437\u2013478 (2022).","journal-title":"Oxygen"},{"key":"54847_CR10","doi-asserted-by":"publisher","first-page":"651","DOI":"10.1038\/s42255-022-00591-z","volume":"4","author":"MP Murphy","year":"2022","unstructured":"Murphy, M. P. et al. Guidelines for measuring reactive oxygen species and oxidative damage in cells and in vivo. Nat. Metab. 4, 651\u2013662 (2022).","journal-title":"Nat. Metab."},{"key":"54847_CR11","doi-asserted-by":"publisher","first-page":"694","DOI":"10.3389\/fphys.2020.00694","volume":"11","author":"M Sharifi-Rad","year":"2020","unstructured":"Sharifi-Rad, M. et al. Lifestyle, oxidative stress, and antioxidants: back and forth in the pathophysiology of chronic diseases. Front. Physiol. 11, 694 (2020).","journal-title":"Front. Physiol."},{"key":"54847_CR12","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1099\/jgv.0.001596","volume":"102","author":"NA Khan","year":"2021","unstructured":"Khan, N. A. et al. Oxidative stress specifically inhibits replication of dengue virus. J. Gen. Virol. 102, 1\u201314 (2021).","journal-title":"J. Gen. Virol."},{"key":"54847_CR13","doi-asserted-by":"crossref","unstructured":"Poljsak, B., \u0160uput, D. & Milisav, I. Achieving the balance between ROS and antioxidants: When to use the synthetic antioxidants. Oxid. Med. Cell. Longevity (2013).","DOI":"10.1155\/2013\/956792"},{"key":"54847_CR14","first-page":"219","volume":"2","author":"K Rahman","year":"2007","unstructured":"Rahman, K. Studies on free radicals, antioxidants, and co-factors. Clin. Interv. Aging 2, 219\u2013236 (2007).","journal-title":"Clin. Interv. Aging"},{"key":"54847_CR15","doi-asserted-by":"publisher","first-page":"586","DOI":"10.1016\/j.bbagen.2011.05.006","volume":"1820","author":"S Ohta","year":"2012","unstructured":"Ohta, S. Molecular hydrogen is a novel antioxidant to efficiently reduce oxidative stress with potential for the improvement of mitochondrial diseases. Biochimica et Biophysica Acta Gen. Subj. 1820, 586\u2013594 (2012).","journal-title":"Biochimica et Biophysica Acta Gen. Subj."},{"key":"54847_CR16","doi-asserted-by":"publisher","first-page":"2241","DOI":"10.2174\/138161211797052664","volume":"17","author":"S Ohta","year":"2011","unstructured":"Ohta, S. Recent progress toward hydrogen medicine: Potential of molecular hydrogen for preventive and therapeutic applications. Curr. Pharm. Des. 17, 2241\u20132252 (2011).","journal-title":"Curr. Pharm. Des."},{"key":"54847_CR17","doi-asserted-by":"publisher","first-page":"114","DOI":"10.4103\/2045-9912.296041","volume":"10","author":"R Asada","year":"2020","unstructured":"Asada, R., Tazawa, K., Sato, S. & Miwa, N. Effects of hydrogen-rich water prepared by alternating-current-electrolysis on antioxidant activity, DNA oxidative injuries, and diabetes-related markers. Med. Gas Res. 10, 114\u2013121 (2020).","journal-title":"Med. Gas Res."},{"key":"54847_CR18","doi-asserted-by":"publisher","first-page":"72","DOI":"10.1007\/s13577-016-0150-x","volume":"30","author":"L Xiao","year":"2017","unstructured":"Xiao, L. & Miwa, N. Hydrogen-rich water achieves cytoprotection from oxidative stress injury in human gingival fibroblasts in culture or 3D-tissue equivalents, and wound-healing promotion, together with ROS-scavenging and relief from glutathione diminishment. Hum. Cell 30, 72\u201387 (2017).","journal-title":"Hum. Cell"},{"key":"54847_CR19","doi-asserted-by":"publisher","first-page":"24","DOI":"10.1016\/j.jphotobiol.2011.09.006","volume":"106","author":"S Kato","year":"2012","unstructured":"Kato, S., Saitoh, Y., Iwai, K. & Miwa, N. Hydrogen-rich electrolyzed warm water represses wrinkle formation against UVA ray together with type-I collagen production and oxidative-stress diminishment in fibroblasts and cell-injury prevention in keratinocytes. J. Photochem. Photobiol. B 106, 24\u201333 (2012).","journal-title":"J. Photochem. Photobiol. B"},{"key":"54847_CR20","doi-asserted-by":"publisher","first-page":"00420","DOI":"10.3389\/fbioe.2019.00420","volume":"7","author":"NK Tripathi","year":"2019","unstructured":"Tripathi, N. K. & Shrivastava, A. Recent developments in bioprocessing of recombinant proteins: Expression hosts and process development. Front. Bioeng. Biotechnol. 7, 00420 (2019).","journal-title":"Front. Bioeng. Biotechnol."},{"key":"54847_CR21","doi-asserted-by":"publisher","first-page":"1063","DOI":"10.1007\/s00253-022-12342-x","volume":"107","author":"WJ Xu","year":"2023","unstructured":"Xu, W. J., Lin, Y., Mi, C. L., Pang, J. Y. & Wang, T. Y. Progress in fed-batch culture for recombinant protein production in CHO cells. Appl. Microbiol. Biotechnol. 107, 1063\u20131075 (2023).","journal-title":"Appl. Microbiol. Biotechnol."},{"key":"54847_CR22","doi-asserted-by":"publisher","first-page":"75","DOI":"10.1038\/s41580-020-00314-w","volume":"22","author":"R Di Micco","year":"2021","unstructured":"Di Micco, R., Krizhanovsky, V., Baker, D. & d\u2019Adda di Fagagna, F. Cellular senescence in ageing: from mechanisms to therapeutic opportunities. Nat. Rev. Mol. Cell Biol. 22, 75\u201395 (2021).","journal-title":"Nat. Rev. Mol. Cell Biol."},{"key":"54847_CR23","doi-asserted-by":"publisher","first-page":"37007","DOI":"10.1038\/srep37007","volume":"6","author":"M Baz-Mart\u00ednez","year":"2016","unstructured":"Baz-Mart\u00ednez, M. et al. Cell senescence is an antiviral defense mechanism. Sci. Rep. 6, 37007 (2016).","journal-title":"Sci. Rep."},{"key":"54847_CR24","doi-asserted-by":"publisher","first-page":"1","DOI":"10.3390\/biology9120455","volume":"9","author":"R Seoane","year":"2020","unstructured":"Seoane, R., Vidal, S., Bouzaher, Y. H., El Motiam, A. & Rivas, C. The interaction of viruses with the cellular senescence response. Biology 9, 1\u201316 (2020).","journal-title":"Biology"},{"key":"54847_CR25","doi-asserted-by":"publisher","first-page":"e00238","DOI":"10.1128\/JVI.00238-20","volume":"94","author":"R Sun","year":"2020","unstructured":"Sun, R. et al. PCV2 induces reactive oxygen species to promote nucleocytoplasmic translocation of the viral DNA binding protein HMGB1 to enhance its replication. J. Virol. 94, e00238-e320 (2020).","journal-title":"J. Virol."},{"key":"54847_CR26","doi-asserted-by":"publisher","first-page":"289","DOI":"10.3390\/v12030289","volume":"12","author":"Y Zhang","year":"2020","unstructured":"Zhang, Y., Sun, R., Li, X. & Fang, W. Porcine circovirus 2 induction of ROS is responsible for mitophagy in PK-15 cells via activation of Drp1 phosphorylation. Viruses 12, 289 (2020).","journal-title":"Viruses"},{"key":"54847_CR27","doi-asserted-by":"publisher","first-page":"186","DOI":"10.1179\/1351000213Y.0000000058","volume":"18","author":"X Chen","year":"2013","unstructured":"Chen, X. et al. Interaction of porcine circovirus type 2 replication with intracellular redox status in vitro. Redox Rep. 18, 186\u2013192 (2013).","journal-title":"Redox Rep."},{"key":"54847_CR28","doi-asserted-by":"publisher","first-page":"66","DOI":"10.1016\/j.virol.2012.01.023","volume":"426","author":"X Chen","year":"2012","unstructured":"Chen, X. et al. Reactive oxygen species regulate the replication of porcine circovirus type 2 via NF-\u03baB pathway. Virology 426, 66\u201372 (2012).","journal-title":"Virology"},{"key":"54847_CR29","doi-asserted-by":"publisher","first-page":"865","DOI":"10.1189\/jlb.1212662","volume":"93","author":"H Yang","year":"2013","unstructured":"Yang, H., Antoine, D. J., Andersson, U. & Tracey, K. J. The many faces of HMGB1: Molecular structure-functional activity in inflammation, apoptosis, and chemotaxis. J. Leukoc. Biol. 93, 865\u2013873 (2013).","journal-title":"J. Leukoc. Biol."},{"key":"54847_CR30","doi-asserted-by":"publisher","DOI":"10.1186\/1471-230X-14-12","author":"Y Liu","year":"2014","unstructured":"Liu, Y. et al. Protective effects of hydrogen enriched saline on liver ischemia reperfusion injury by reducing oxidative stress and HMGB1 release. BMC Gastroenterol. https:\/\/doi.org\/10.1186\/1471-230X-14-12 (2014).","journal-title":"BMC Gastroenterol."},{"key":"54847_CR31","doi-asserted-by":"publisher","first-page":"11","DOI":"10.1016\/j.intimp.2019.01.022","volume":"69","author":"Y Yu","year":"2019","unstructured":"Yu, Y. et al. Hydrogen gas reduces HMGB1 release in lung tissues of septic mice in an Nrf2\/HO-1-dependent pathway. Int. Immunopharmacol. 69, 11\u201318 (2019).","journal-title":"Int. Immunopharmacol."},{"key":"54847_CR32","doi-asserted-by":"publisher","first-page":"54","DOI":"10.1016\/j.cytogfr.2021.08.003","volume":"62","author":"X Ding","year":"2021","unstructured":"Ding, X., Li, S. & Zhu, L. Potential effects of HMGB1 on viral replication and virus infection-induced inflammatory responses: A promising therapeutic target for virus infection-induced inflammatory diseases. Cytokine Growth Factor Rev. 62, 54\u201361 (2021).","journal-title":"Cytokine Growth Factor Rev."},{"key":"54847_CR33","doi-asserted-by":"publisher","DOI":"10.3389\/fphys.2021.789507","author":"Y Tian","year":"2021","unstructured":"Tian, Y. et al. Hydrogen, a novel therapeutic molecule, regulates oxidative stress, inflammation, and apoptosis. Front. Physiol. https:\/\/doi.org\/10.3389\/fphys.2021.789507 (2021).","journal-title":"Front. Physiol."},{"key":"54847_CR34","doi-asserted-by":"publisher","first-page":"219","DOI":"10.4103\/2045-9912.196904","volume":"6","author":"L Huang","year":"2016","unstructured":"Huang, L. Molecular hydrogen: A therapeutic antioxidant and beyond. Med. Gas Res. 6, 219\u2013222 (2016).","journal-title":"Med. Gas Res."},{"key":"54847_CR35","doi-asserted-by":"publisher","first-page":"313","DOI":"10.1186\/s12917-019-2059-6","volume":"15","author":"LT Cheng","year":"2019","unstructured":"Cheng, L. T., Zeng, Y. J., Chu, C. Y. & Wang, H. Y. Development of a quick dot blot assay for the titering of bovine ephemeral fever virus. BMC Vet. Res. 15, 313 (2019).","journal-title":"BMC Vet. Res."},{"key":"54847_CR36","doi-asserted-by":"publisher","first-page":"543","DOI":"10.1292\/jvms.68.543","volume":"68","author":"Y-C Hsieh","year":"2006","unstructured":"Hsieh, Y.-C. et al. DNA sequence analysis of glycoprotein g gene of bovine ephemeral fever virus and development of a double oil emulsion vaccine against bovine ephemeral fever. Vet. Med. Sci. 68, 543\u2013548 (2006).","journal-title":"Vet. Med. Sci."}],"container-title":["Scientific Reports"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.nature.com\/articles\/s41598-024-54847-7.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/www.nature.com\/articles\/s41598-024-54847-7","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/www.nature.com\/articles\/s41598-024-54847-7.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2024,2,19]],"date-time":"2024-02-19T13:13:33Z","timestamp":1708348413000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.nature.com\/articles\/s41598-024-54847-7"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,2,19]]},"references-count":36,"journal-issue":{"issue":"1","published-online":{"date-parts":[[2024,12]]}},"alternative-id":["54847"],"URL":"https:\/\/doi.org\/10.1038\/s41598-024-54847-7","relation":{},"ISSN":["2045-2322"],"issn-type":[{"type":"electronic","value":"2045-2322"}],"subject":[],"published":{"date-parts":[[2024,2,19]]},"assertion":[{"value":"16 November 2023","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"17 February 2024","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"19 February 2024","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"The authors declare no competing interests.","order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Competing interests"}}],"article-number":"4092"}}