{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,12,13]],"date-time":"2025-12-13T05:22:35Z","timestamp":1765603355538,"version":"3.48.0"},"reference-count":56,"publisher":"MDPI AG","issue":"12","license":[{"start":{"date-parts":[[2025,12,11]],"date-time":"2025-12-11T00:00:00Z","timestamp":1765411200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100003725","name":"National Research Foundation of Korea","doi-asserted-by":"crossref","id":[{"id":"10.13039\/501100003725","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/501100003725","name":"Ministry of Education","doi-asserted-by":"publisher","award":["2022R1A6A1A03053343"],"award-info":[{"award-number":["2022R1A6A1A03053343"]}],"id":[{"id":"10.13039\/501100003725","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003725","name":"Korea government","doi-asserted-by":"publisher","award":["RS-2025-24683056"],"award-info":[{"award-number":["RS-2025-24683056"]}],"id":[{"id":"10.13039\/501100003725","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003725","name":"Korea government","doi-asserted-by":"publisher","award":["RS-2025-02273000"],"award-info":[{"award-number":["RS-2025-02273000"]}],"id":[{"id":"10.13039\/501100003725","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Antioxidants"],"abstract":"<jats:p>Most respiratory diseases are driven by excessive airway inflammation and oxidative stress, yet current therapies often lack durable efficacy or are unsafe. Host-defense peptides, commonly enriched in animal venoms, offer diverse, target-selective scaffolds for new therapeutics. In this study, we aimed to discover a novel bioactive peptide with therapeutic potential on respiratory tract damage by utilizing Nephila clavata venom gland transcriptome. Using in silico analysis and machine learning-based functional prediction, we designed a peptide, NC-CV, expected to have dual anti-inflammatory and antioxidant activities with low cytotoxicity. In experimental validation, NC-CV improved human bronchial epithelial BEAS-2B cell viability under lipopolysaccharide (LPS) exposure while reducing LPS-induced pro-inflammatory cytokine expression and intracellular reactive oxygen species (ROS) generation. Mechanistic studies and molecular docking simulations indicated that NC-CV prevents toll-like receptor 4 signaling activation, suppressing nuclear factor \u03baB and mitogen-activated protein kinase pathways. Moreover, the antioxidant activity of NC-CV was primarily based on direct intracellular ROS scavenging rather than the induction of endogenous antioxidant enzymes. Collectively, these findings demonstrated that the venom-derived peptide NC-CV disrupts the self-reinforcing cycle involving inflammatory signaling and oxidative stress in airway epithelium, highlighting its promise as a therapeutic candidate for respiratory disease.<\/jats:p>","DOI":"10.3390\/antiox14121485","type":"journal-article","created":{"date-parts":[[2025,12,11]],"date-time":"2025-12-11T11:17:27Z","timestamp":1765451847000},"page":"1485","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Spider Venom-Derived Peptide Exhibits Dual Anti-Inflammatory and Antioxidative Activities in LPS-Stimulated BEAS-2B Cells"],"prefix":"10.3390","volume":"14","author":[{"given":"Jin Wook","family":"Oh","sequence":"first","affiliation":[{"name":"Department of Life Science, Dongguk University-Seoul, Goyang 10326, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Min Kyoung","family":"Shin","sequence":"additional","affiliation":[{"name":"Department of Life Science, Dongguk University-Seoul, Goyang 10326, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0007-3574-2046","authenticated-orcid":false,"given":"Hye-Ran","family":"Park","sequence":"additional","affiliation":[{"name":"Department of Life Science, Dongguk University-Seoul, Goyang 10326, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sukin","family":"Jeong","sequence":"additional","affiliation":[{"name":"Department of Life Science, Dongguk University-Seoul, Goyang 10326, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0168-9546","authenticated-orcid":false,"given":"Minho","family":"Lee","sequence":"additional","affiliation":[{"name":"Department of Life Science, Dongguk University-Seoul, Goyang 10326, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4848-3764","authenticated-orcid":false,"given":"Ji Hyuk","family":"Ko","sequence":"additional","affiliation":[{"name":"Department of Life Science, Dongguk University-Seoul, Goyang 10326, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jae Young","family":"Lee","sequence":"additional","affiliation":[{"name":"Department of Life Science, Dongguk University-Seoul, Goyang 10326, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Seung-Cheol","family":"Jee","sequence":"additional","affiliation":[{"name":"Department of Life Science, Dongguk University-Seoul, Goyang 10326, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3145-8458","authenticated-orcid":false,"given":"Jung-Suk","family":"Sung","sequence":"additional","affiliation":[{"name":"Department of Life Science, Dongguk University-Seoul, Goyang 10326, Republic of Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,12,11]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"404","DOI":"10.1513\/AnnalsATS.201311-405PS","article-title":"The global burden of respiratory disease","volume":"11","author":"Ferkol","year":"2014","journal-title":"Ann. Am. Thorac. Soc."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"27","DOI":"10.1111\/j.1440-1843.2008.01447.x","article-title":"Impact of oxidative stress on lung diseases","volume":"14","author":"Park","year":"2009","journal-title":"Respirology"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"S150","DOI":"10.1513\/AnnalsATS.201305-116AW","article-title":"Oxidative stress in airway diseases","volume":"10","author":"Holguin","year":"2013","journal-title":"Ann. Am. Thorac. Soc."},{"doi-asserted-by":"crossref","unstructured":"Alshabanat, A., Zafari, Z., Albanyan, O., Dairi, M., and FitzGerald, J. (2015). Asthma and COPD overlap syndrome (ACOS): A systematic review and meta analysis. PLoS ONE, 10.","key":"ref_4","DOI":"10.1371\/journal.pone.0136065"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"22580","DOI":"10.1038\/s41598-025-07197-x","article-title":"Incidence, risk factors, and outcomes of respiratory related hospitalizations in older patients with COPD, asthma, and bronchiectasis","volume":"15","author":"Chae","year":"2025","journal-title":"Sci. Rep."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"289","DOI":"10.1097\/00004836-200110000-00006","article-title":"Side effects of corticosteroid therapy","volume":"33","author":"Buchman","year":"2001","journal-title":"J. Clin. Gastroenterol."},{"key":"ref_7","first-page":"5484","article-title":"LPS-induced proinflammatory cytokine expression in human airway epithelial cells and macrophages via NF-\u03baB, STAT3 or AP-1 activation","volume":"17","author":"Liu","year":"2018","journal-title":"Mol. Med. Rep."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"294","DOI":"10.1046\/j.1365-3083.2002.01137.x","article-title":"Differences in LPS-induced activation of bronchial epithelial cells (BEAS-2B) and type II-like pneumocytes (A-549)","volume":"56","author":"Schulz","year":"2002","journal-title":"Scand. J. Immunol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"145","DOI":"10.1016\/j.cyto.2008.01.006","article-title":"LPS\/TLR4 signal transduction pathway","volume":"42","author":"Lu","year":"2008","journal-title":"Cytokine"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"7421","DOI":"10.1073\/pnas.1201193109","article-title":"Structural basis of species-specific endotoxin sensing by innate immune receptor TLR4\/MD-2","volume":"109","author":"Ohto","year":"2012","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"107272","DOI":"10.1016\/j.intimp.2020.107272","article-title":"Effect of gut microbiota on LPS-induced acute lung injury by regulating the TLR4\/NF-kB signaling pathway","volume":"91","author":"Tang","year":"2021","journal-title":"Int. Immunopharmacol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"191","DOI":"10.1016\/j.neulet.2014.10.016","article-title":"Mitochondrial ROS govern the LPS-induced pro-inflammatory response in microglia cells by regulating MAPK and NF-\u03baB pathways","volume":"584","author":"Park","year":"2015","journal-title":"Neurosci. Lett."},{"doi-asserted-by":"crossref","unstructured":"Sul, O.-J., and Ra, S.W. (2021). Quercetin Prevents LPS-Induced Oxidative Stress and Inflammation by Modulating NOX2\/ROS\/NF-kB in Lung Epithelial Cells. Molecules, 26.","key":"ref_13","DOI":"10.3390\/molecules26226949"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"447","DOI":"10.1016\/j.cardiores.2006.09.012","article-title":"Role of NADPH oxidase 4 in lipopolysaccharide-induced proinflammatory responses by human aortic endothelial cells","volume":"72","author":"Park","year":"2006","journal-title":"Cardiovasc. Res."},{"unstructured":"Oslowski, C.M. (2015). Cross Talk Between ER Stress, Oxidative Stress, and Inflammation in Health and Disease. Stress Responses: Methods and Protocols, Springer.","key":"ref_15"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"29","DOI":"10.1093\/fqs\/fyx006","article-title":"Bioactive peptides: A review","volume":"1","year":"2017","journal-title":"Food Qual. Saf."},{"doi-asserted-by":"crossref","unstructured":"Daliri, E.B.-M., Oh, D.H., and Lee, B.H. (2017). Bioactive Peptides. Foods, 6.","key":"ref_17","DOI":"10.3390\/foods6050032"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"125","DOI":"10.1007\/s00011-018-1208-x","article-title":"Antiinflammatory peptides: Current knowledge and promising prospects","volume":"68","author":"Dadar","year":"2019","journal-title":"Inflamm. Res."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"139797","DOI":"10.1016\/j.foodchem.2024.139797","article-title":"Purification, identification and molecular docking studies of antioxidant and anti-inflammatory peptides from Edible Bird\u2019s Nest","volume":"454","author":"Lee","year":"2024","journal-title":"Food Chem."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"44","DOI":"10.1016\/j.tifs.2020.02.012","article-title":"Advances on the antioxidant peptides from edible plant sources","volume":"99","author":"Wong","year":"2020","journal-title":"Trends Food Sci. Technol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"1551","DOI":"10.1038\/nbt1267","article-title":"Antimicrobial and host-defense peptides as new anti-infective therapeutic strategies","volume":"24","author":"Hancock","year":"2006","journal-title":"Nat. Biotechnol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"5377","DOI":"10.1007\/s13197-015-1731-5","article-title":"Bioactive peptides of animal origin: A review","volume":"52","author":"Bhat","year":"2015","journal-title":"J. Food Sci. Technol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"3137","DOI":"10.1093\/bioinformatics\/btx373","article-title":"FQC Dashboard: Integrates FastQC results into a web-based, interactive, and extensible FASTQ quality control tool","volume":"33","author":"Brown","year":"2017","journal-title":"Bioinformatics"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"2114","DOI":"10.1093\/bioinformatics\/btu170","article-title":"Trimmomatic: A flexible trimmer for Illumina sequence data","volume":"30","author":"Bolger","year":"2014","journal-title":"Bioinformatics"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"644","DOI":"10.1038\/nbt.1883","article-title":"Full-length transcriptome assembly from RNA-Seq data without a reference genome","volume":"29","author":"Grabherr","year":"2011","journal-title":"Nat. Biotechnol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"4647","DOI":"10.1093\/molbev\/msab199","article-title":"BUSCO update: Novel and streamlined workflows along with broader and deeper phylogenetic coverage for scoring of eukaryotic, prokaryotic, and viral genomes","volume":"38","author":"Manni","year":"2021","journal-title":"Mol. Biol. Evol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"417","DOI":"10.1038\/nmeth.4197","article-title":"Salmon provides fast and bias-aware quantification of transcript expression","volume":"14","author":"Patro","year":"2017","journal-title":"Nat. Methods"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"139","DOI":"10.1093\/bioinformatics\/btp616","article-title":"edgeR: A Bioconductor package for differential expression analysis of digital gene expression data","volume":"26","author":"Robinson","year":"2010","journal-title":"Bioinformatics"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1494","DOI":"10.1038\/nprot.2013.084","article-title":"De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis","volume":"8","author":"Haas","year":"2013","journal-title":"Nat. Protoc."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"420","DOI":"10.1038\/s41587-019-0036-z","article-title":"SignalP 5.0 improves signal peptide predictions using deep neural networks","volume":"37","author":"Tsirigos","year":"2019","journal-title":"Nat. Biotechnol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"3403","DOI":"10.1093\/bioinformatics\/btm477","article-title":"XtalPred: A web server for prediction of protein crystallizability","volume":"23","author":"Slabinski","year":"2007","journal-title":"Bioinformatics"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"W432","DOI":"10.1093\/nar\/gkad376","article-title":"PEP-FOLD4: A pH-dependent force field for peptide structure prediction in aqueous solution","volume":"51","author":"Rey","year":"2023","journal-title":"Nucleic Acids Res."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"2101","DOI":"10.1093\/bioinformatics\/btn392","article-title":"HELIQUEST: A web server to screen sequences with specific \u03b1-helical properties","volume":"24","author":"Gautier","year":"2008","journal-title":"Bioinformatics"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"21471","DOI":"10.1038\/s41598-020-78319-w","article-title":"AnOxPePred: Using deep learning for the prediction of antioxidative properties of peptides","volume":"10","author":"Olsen","year":"2020","journal-title":"Sci. Rep."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"1198","DOI":"10.1038\/s42003-024-06911-1","article-title":"PepNet: An interpretable neural network for anti-inflammatory and antimicrobial peptides prediction using a pre-trained protein language model","volume":"7","author":"Han","year":"2024","journal-title":"Commun. Biol."},{"doi-asserted-by":"crossref","unstructured":"Khatun, M.S., Hasan, M.M., and Kurata, H. (2019). PreAIP: Computational prediction of anti-inflammatory peptides by integrating multiple complementary features. Front. Genet., 10.","key":"ref_36","DOI":"10.3389\/fgene.2019.00129"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"D288","DOI":"10.1093\/nar\/gkaa991","article-title":"DBAASP v3: Database of antimicrobial\/cytotoxic activity and structure of peptides as a resource for development of new therapeutics","volume":"49","author":"Pirtskhalava","year":"2021","journal-title":"Nucleic Acids Res."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"2546","DOI":"10.1021\/acs.jcim.2c01317","article-title":"Serverless prediction of peptide properties with recurrent neural networks","volume":"63","author":"Ansari","year":"2023","journal-title":"J. Chem. Inf. Model."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"1191","DOI":"10.1038\/nature07830","article-title":"The structural basis of lipopolysaccharide recognition by the TLR4\u2013MD-2 complex","volume":"458","author":"Park","year":"2009","journal-title":"Nature"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"2785","DOI":"10.1002\/jcc.21256","article-title":"AutoDock4 and AutoDockTools4: Automated docking with selective receptor flexibility","volume":"30","author":"Morris","year":"2009","journal-title":"J. Comput. Chem."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"679","DOI":"10.1038\/s41592-022-01488-1","article-title":"ColabFold: Making protein folding accessible to all","volume":"19","author":"Mirdita","year":"2022","journal-title":"Nat. Methods"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"3891","DOI":"10.1021\/acs.jcim.1c00203","article-title":"AutoDock Vina 1.2. 0: New docking methods, expanded force field, and python bindings","volume":"61","author":"Eberhardt","year":"2021","journal-title":"J. Chem. Inf. Model."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"774","DOI":"10.1016\/j.jmb.2007.05.022","article-title":"Inference of macromolecular assemblies from crystalline state","volume":"372","author":"Krissinel","year":"2007","journal-title":"J. Mol. Biol."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"1167","DOI":"10.1016\/j.toxicon.2008.02.001","article-title":"Intersexual variations in Northern (Missulena pruinosa) and Eastern (M. bradleyi) mouse spider venom","volume":"51","author":"Herzig","year":"2008","journal-title":"Toxicon"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"e4691","DOI":"10.7717\/peerj.4691","article-title":"Sexually dimorphic venom proteins in long-jawed orb-weaving spiders (Tetragnatha) comprise novel gene families","volume":"6","author":"Bulger","year":"2018","journal-title":"PeerJ"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"750","DOI":"10.1016\/j.foodchem.2017.11.119","article-title":"Designing antioxidant peptides based on the antioxidant properties of the amino acid side-chains","volume":"245","author":"Matsui","year":"2018","journal-title":"Food Chem."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"1789","DOI":"10.1016\/j.foodchem.2012.06.016","article-title":"Antioxidant and metal chelating activities of peptide fractions from phaseolin and bean protein hydrolysates","volume":"135","author":"Alaiz","year":"2012","journal-title":"Food Chem."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"401","DOI":"10.1146\/annurev-pharmtox-011112-140320","article-title":"Role of nrf2 in oxidative stress and toxicity","volume":"53","author":"Ma","year":"2013","journal-title":"Annu. Rev. Pharmacol. Toxicol."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"203","DOI":"10.1016\/j.molimm.2015.08.008","article-title":"Studies of the TLR4-associated protein MD-2 using yeast-display and mutational analyses","volume":"68","author":"Mattis","year":"2015","journal-title":"Mol. Immunol."},{"doi-asserted-by":"crossref","unstructured":"Cui, L., Li, W., He, S., Guo, L., Liu, K., Dong, J., Li, J., and Wang, H. (2025). Inhibition of NOX4-Mediated ROS Production Contributes to Selenomethionine\u2019s Anti-Inflammatory Effect in LPS-Stimulated Bovine Endometrial Epithelial Cells. Vet. Sci., 12.","key":"ref_50","DOI":"10.3390\/vetsci12090789"},{"doi-asserted-by":"crossref","unstructured":"Zhang, Y., Li, Y., Quan, Z., Xiao, P., and Duan, J.-A. (2024). New insights into antioxidant peptides: An overview of efficient screening, evaluation models, molecular mechanisms, and applications. Antioxidants, 13.","key":"ref_51","DOI":"10.3390\/antiox13020203"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"509","DOI":"10.1038\/nchem.1062","article-title":"Contemporary strategies for peptide macrocyclization","volume":"3","author":"White","year":"2011","journal-title":"Nat. Chem."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"296","DOI":"10.1038\/342296a0","article-title":"Capping and \u03b1-helix stability","volume":"342","author":"Serrano","year":"1989","journal-title":"Nature"},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"3185","DOI":"10.2174\/138161210793292555","article-title":"Chemical modifications designed to improve peptide stability: Incorporation of non-natural amino acids, pseudo-peptide bonds, and cyclization","volume":"16","author":"Gentilucci","year":"2010","journal-title":"Curr. Pharm. Des."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"2692","DOI":"10.1021\/bi048046i","article-title":"Binding of oligoarginine to membrane lipids and heparan sulfate: Structural and thermodynamic characterization of a cell-penetrating peptide","volume":"44","author":"Kitas","year":"2005","journal-title":"Biochemistry"},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"1130","DOI":"10.1021\/acs.accounts.6b00074","article-title":"How membrane-active peptides get into lipid membranes","volume":"49","author":"Sani","year":"2016","journal-title":"Acc. Chem. Res."}],"container-title":["Antioxidants"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-3921\/14\/12\/1485\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,12,13]],"date-time":"2025-12-13T05:18:29Z","timestamp":1765603109000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-3921\/14\/12\/1485"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,12,11]]},"references-count":56,"journal-issue":{"issue":"12","published-online":{"date-parts":[[2025,12]]}},"alternative-id":["antiox14121485"],"URL":"https:\/\/doi.org\/10.3390\/antiox14121485","relation":{},"ISSN":["2076-3921"],"issn-type":[{"type":"electronic","value":"2076-3921"}],"subject":[],"published":{"date-parts":[[2025,12,11]]}}}