{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,15]],"date-time":"2026-07-15T05:07:35Z","timestamp":1784092055518,"version":"3.55.0"},"reference-count":43,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,7,15]],"date-time":"2026-07-15T00:00:00Z","timestamp":1784073600000},"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. Immunol."],"abstract":"<jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>Airborne pollutants comprise biological agents such as lipopolysaccharide (LPS) and engineered nanomaterials, including carbon nanotubes (CNTs), which are increasingly prevalent in industrial and consumer applications. Although the pulmonary toxicity of high-dose CNT exposure is established, the inflammatory consequences of low-dose, non-cytotoxic CNT exposure, particularly in combination with other aerosols, remain poorly understood.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>We investigated cytotoxicity and proinflammatory responses to multi-walled CNTs in three-dimensional A549 human epithelial spheroids, compared with A549 monolayers and MM6 monocytes. Cells were exposed to two CNTs differing in diameter (CNT1: 7\u201315 nm; CNT2: 110\u2013170 nm), alone or combined with LPS. Cell viability was assessed by WST-1 assay and F-actin junction staining, while IL-8 and IL-6 production was quantified by ELISA and qPCR.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>A549 spheroids were more susceptible to CNT-induced cytotoxicity than monolayers, with CNT1 inducing greater cytotoxicity than CNT2. At non-cytotoxic concentrations, CNTs alone did not elicit cytokine release; however, co-exposure with LPS significantly enhanced cytokine secretion in spheroids, particularly in the presence of CNT2. This synergistic inflammatory response was not observed in MM6 monocytes.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusion<\/jats:title>\n                    <jats:p>Our study demonstrates that non-cytotoxic CNTs can potentiate LPS-driven inflammation in A549 epithelial spheroids, suggesting potential respiratory risks from combined exposure to environmental endotoxins and nanomaterials, even at low doses.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.3389\/fimmu.2026.1794151","type":"journal-article","created":{"date-parts":[[2026,7,15]],"date-time":"2026-07-15T04:44:18Z","timestamp":1784090658000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":0,"title":["Low-dose multi-walled carbon nanotubes are non-inflammatory alone but amplify lipopolysaccharide-induced cytokine responses in A549 lung epithelial spheroids"],"prefix":"10.3389","volume":"17","author":[{"given":"Jian","family":"Liu","sequence":"first","affiliation":[{"name":"School of Biochemistry and Biomedical Sciences, Faculty of Health and Life Sciences, University of Bristol","place":["Bristol, United Kingdom"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Li","family":"Zeng","sequence":"additional","affiliation":[{"name":"Shanxi Academy of Medical Sciences","place":["Taiyuan, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wondwossen","family":"Abate","sequence":"additional","affiliation":[{"name":"Clinical and Biomedical Sciences, University of Exeter Medical School","place":["Exeter, United Kingdom"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Simon K.","family":"Jackson","sequence":"additional","affiliation":[{"name":"School of Biomedical Science, Faculty of Health, University of Plymouth","place":["Plymouth, United Kingdom"]},{"name":"Molendotech Ltd, Brixham Laboratory","place":["Brixham, Devon, United Kingdom"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2026,7,15]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","DOI":"10.1111\/bcp.12546","article-title":"Characterization of the inflammatory response to inhaled lipopolysaccharide in mild to moderate chronic obstructive pulmonary disease","volume":"79","author":"Gupta","year":"2015","journal-title":"Br J Clin Pharmacol"},{"key":"B2","doi-asserted-by":"publisher","DOI":"10.1126\/scisignal.abd2533","article-title":"The suppressive effects of Mer inhibition on inflammatory responses in the pathogenesis of LPS-induced ALI\/ARDS","volume":"15","author":"Fukatsu","year":"2022","journal-title":"Sci Signal"},{"key":"B3","doi-asserted-by":"publisher","DOI":"10.1186\/1465-9921-10-126","article-title":"Inhibiting toll-like receptor 4 signaling ameliorates pulmonary fibrosis during acute lung injury induced by lipopolysaccharide: an experimental study","volume":"10","author":"He","year":"2009","journal-title":"Respir Res"},{"key":"B4","doi-asserted-by":"publisher","DOI":"10.1189\/jlb.3MR0316-118RR","article-title":"Molecular mechanisms of innate memory and tolerance to LPS","volume":"101","author":"Seeley","year":"2017","journal-title":"J Leukoc Biol"},{"key":"B5","doi-asserted-by":"publisher","DOI":"10.1038\/s12276-020-0394-0","article-title":"Mechanisms of ultrafine particle-induced respiratory health effects","volume":"52","author":"Leikauf","year":"2020","journal-title":"Exp Mol Med"},{"key":"B6","doi-asserted-by":"publisher","DOI":"10.1097\/SHK.0b013e3181aa9690","article-title":"Chronic pulmonary LPS tolerance induces selective immunosuppression while maintaining the neutrophilic response","volume":"33","author":"Natarajan","year":"2010","journal-title":"Shock"},{"key":"B7","doi-asserted-by":"publisher","DOI":"10.1080\/15376516.2024.2431687","article-title":"Mechanistic paradigms of immunotoxicity, triggered by nanoparticles - a review","volume":"35","author":"Rana","year":"2025","journal-title":"Toxicol Mech Methods"},{"key":"B8","doi-asserted-by":"publisher","DOI":"10.1002\/tox.22248","article-title":"Effects of lipopolysaccharide, multiwalled carbon nantoubes, and the combination on lung alveolar epithelial cells","volume":"32","author":"Pacurari","year":"2017","journal-title":"Environ Toxicol"},{"key":"B9","doi-asserted-by":"publisher","DOI":"10.3402\/nano.v4i0.21521","article-title":"A concise review of carbon nanotube's toxicology","volume":"4","author":"Madani","year":"2013","journal-title":"Nano Rev"},{"key":"B10","doi-asserted-by":"publisher","DOI":"10.1021\/acsanm.4c02721","article-title":"Review of carbon nanotube research and development: materials and emerging applications","volume":"7","author":"Hughes","year":"2024","journal-title":"ACS Appl Nano Mater"},{"key":"B11","doi-asserted-by":"publisher","first-page":"1059","DOI":"10.3390\/nano13061059","article-title":"Single-walled vs. Multi-walled carbon nanotubes: influence of physico-chemical properties on toxicogenomics responses in mouse lungs","volume":"13","author":"Solorio-Rodriguez","year":"2023","journal-title":"Nanomaterials (Basel)"},{"key":"B12","doi-asserted-by":"publisher","DOI":"10.1039\/d2ra02519a","article-title":"In vitro toxicity of carbon nanotubes: a systematic review","volume":"12","author":"Chetyrkina","year":"2022","journal-title":"RSC Adv"},{"key":"B13","doi-asserted-by":"publisher","DOI":"10.1021\/acs.est.5b01841","article-title":"Lipopolysaccharide density and structure govern the extent and distance of nanoparticle interaction with actual and model bacterial outer membranes","volume":"49","author":"Jacobson","year":"2015","journal-title":"Environ Sci Technol"},{"key":"B14","doi-asserted-by":"publisher","DOI":"10.1289\/ehp.8903","article-title":"Effects of airway exposure to nanoparticles on lung inflammation induced by bacterial endotoxin in mice","volume":"114","author":"Inoue","year":"2006","journal-title":"Environ Health Perspect"},{"key":"B15","doi-asserted-by":"publisher","DOI":"10.1016\/j.jaerosci.2021.105771","article-title":"Translating nanoparticle dosimetry from conventional in vitro systems to occupational inhalation exposures","volume":"155","author":"Smith","year":"2021","journal-title":"J Aerosol Sci"},{"key":"B16","doi-asserted-by":"publisher","first-page":"745","DOI":"10.3390\/nano11030745","article-title":"Occupational exposure to carbon nanotubes and carbon nanofibres: more than a cobweb","volume":"11","author":"Bergamaschi","year":"2021","journal-title":"Nanomaterials (Basel)"},{"key":"B17","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.73.2.1129-1140.2005","article-title":"A549 lung epithelial cells grown as three-dimensional aggregates: alternative tissue culture model for Pseudomonas aeruginosa pathogenesis","volume":"73","author":"Carterson","year":"2005","journal-title":"Infect Immun"},{"key":"B18","doi-asserted-by":"publisher","DOI":"10.1016\/j.jiph.2020.03.011","article-title":"A three-dimensional A549 cell culture model to study respiratory syncytial virus infections","volume":"13","author":"Saleh","year":"2020","journal-title":"J Infect Public Health"},{"key":"B19","doi-asserted-by":"publisher","DOI":"10.1177\/1753425910365733","article-title":"Three-dimensional spheroid cultures of A549 and HepG2 cells exhibit different lipopolysaccharide (LPS) receptor expression and LPS-induced cytokine response compared with monolayer cultures","volume":"17","author":"Liu","year":"2011","journal-title":"Innate Immun"},{"key":"B20","doi-asserted-by":"publisher","first-page":"6621","DOI":"10.1038\/s41598-021-86037-0","article-title":"Long-term evolution of the epithelial cell secretome in preclinical 3D models of the human bronchial epithelium","volume":"11","author":"Sanchez-Guzman","year":"2021","journal-title":"Sci Rep"},{"key":"B21","doi-asserted-by":"publisher","DOI":"10.1002\/cbin.12075","article-title":"Significance of cell culture phases for utilizing 3D cultures of A549 cells for in vitro studies","volume":"47","author":"N G","year":"2023","journal-title":"Cell Biol Int"},{"key":"B22","doi-asserted-by":"publisher","DOI":"10.1002\/jcb.21345","article-title":"Functional three-dimensional HepG2 aggregate cultures generated from an ultrasound trap: comparison with HepG2 spheroids","volume":"102","author":"Liu","year":"2007","journal-title":"J Cell Biochem"},{"key":"B23","doi-asserted-by":"publisher","first-page":"192","DOI":"10.1016\/j.jconrel.2012.04.045","article-title":"Opportunities and challenges for use of tumor spheroids as models to test drug delivery and efficacy","volume":"164","author":"Mehta","year":"2012","journal-title":"J Control Release"},{"key":"B24","doi-asserted-by":"publisher","DOI":"10.1155\/2016\/2159537","article-title":"Effects of multiwalled carbon nanotube surface modification and purification on bovine serum albumin binding and biological responses","volume":"2016","author":"Bai","year":"2016","journal-title":"J Nanomater"},{"key":"B25","doi-asserted-by":"publisher","DOI":"10.1021\/nn102112b","article-title":"Quantitative techniques for assessing and controlling the dispersion and biological effects of multiwalled carbon nanotubes in mammalian tissue culture cells","volume":"4","author":"Wang","year":"2010","journal-title":"ACS Nano"},{"key":"B26","doi-asserted-by":"publisher","DOI":"10.1186\/1754-1611-6-22","article-title":"Multi-walled carbon nanotube-induced inflammatory response and oxidative stress in a dynamic cell growth environment","volume":"6","author":"Patel","year":"2012","journal-title":"J Biol Eng"},{"key":"B27","doi-asserted-by":"publisher","DOI":"10.1039\/d0nr03330h","article-title":"Rapid and sensitive quantification of cell-associated multi-walled carbon nanotubes","volume":"12","author":"Steinmetz","year":"2020","journal-title":"Nanoscale"},{"key":"B28","doi-asserted-by":"publisher","DOI":"10.2147\/IJN.S26573","article-title":"Effect of dispersants of multi-walled carbon nanotubes on cellular uptake and biological responses","volume":"6","author":"Haniu","year":"2011","journal-title":"Int J Nanomedicine"},{"key":"B29","doi-asserted-by":"publisher","DOI":"10.1186\/s44331-026-00010-0","article-title":"Discrete, oxidized multi-walled carbon nanotubes for efficient plasmid DNA transfection","volume":"2","author":"Quinlan","year":"2026","journal-title":"Bioscience Nanotechnology"},{"key":"B30","doi-asserted-by":"publisher","first-page":"96","DOI":"10.3390\/mi12010096","article-title":"Recent advances in three-dimensional multicellular spheroid culture and future development","volume":"12","author":"Shen","year":"2021","journal-title":"Micromachines (Basel)"},{"key":"B31","doi-asserted-by":"publisher","DOI":"10.3389\/fbioe.2023.1299166","article-title":"Carbon nanotube nanocomposite scaffolds: advances in fabrication and applications for tissue regeneration and cancer therapy","volume":"11","author":"Shar","year":"2023","journal-title":"Front Bioeng Biotechnol"},{"key":"B32","doi-asserted-by":"publisher","DOI":"10.3390\/foods13040554","article-title":"Understanding the interaction between nanomaterials originated from high-temperature processed starch\/myristic acid and human monocyte cells","volume":"13","author":"Periasamy","year":"2024","journal-title":"Foods"},{"key":"B33","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.00373-09","article-title":"Use of toll-like receptor assays to detect and identify microbial contaminants in biological products","volume":"47","author":"Huang","year":"2009","journal-title":"J Clin Microbiol"},{"key":"B34","doi-asserted-by":"publisher","first-page":"19","DOI":"10.1007\/BF03257159","article-title":"The role of interleukin-8 and its receptors in inflammatory lung disease: implications for therapy","volume":"1","author":"Pease","year":"2002","journal-title":"Am J Respir Med"},{"key":"B35","doi-asserted-by":"publisher","DOI":"10.1186\/1465-9921-6-116","article-title":"Respiratory epithelial cells require Toll-like receptor 4 for induction of human beta-defensin 2 by lipopolysaccharide","volume":"6","author":"MacRedmond","year":"2005","journal-title":"Respir Res"},{"key":"B36","doi-asserted-by":"publisher","first-page":"294","DOI":"10.1177\/17534259211013887","article-title":"Baicalin attenuates LPS-induced alveolar type II epithelial cell A549 injury by attenuation of the FSTL1 signaling pathway via increasing miR-200b-3p expression","volume":"27","author":"Duan","year":"2021","journal-title":"Innate Immun"},{"key":"B37","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0021827","article-title":"Innate immune responses to bacterial ligands in the peripheral human lung--role of alveolar epithelial TLR expression and signalling","volume":"6","author":"Thorley","year":"2011","journal-title":"PloS One"},{"key":"B38","doi-asserted-by":"publisher","DOI":"10.1080\/17435390.2017.1401142","article-title":"Bacterial endotoxin (lipopolysaccharide) binds to the surface of gold nanoparticles, interferes with biocorona formation and induces human monocyte inflammatory activation","volume":"11","author":"Li","year":"2017","journal-title":"Nanotoxicology"},{"key":"B39","doi-asserted-by":"publisher","DOI":"10.1186\/s12989-016-0127-7","article-title":"Multi-walled carbon nanotube induces nitrative DNA damage in human lung epithelial cells via HMGB1-RAGE interaction and Toll-like receptor 9 activation","volume":"13","author":"Hiraku","year":"2016","journal-title":"Part Fibre Toxicol"},{"key":"B40","doi-asserted-by":"publisher","DOI":"10.1007\/s00210-023-02541-2","article-title":"Harnessing three-dimensional (3D) cell culture models for pulmonary infections: State of the art and future directions","volume":"396","author":"Shah","year":"2023","journal-title":"Naunyn Schmiedebergs Arch Pharmacol"},{"key":"B41","doi-asserted-by":"publisher","DOI":"10.1165\/rcmb.2009-0113OC","article-title":"Bacterial lipopolysaccharide enhances PDGF signaling and pulmonary fibrosis in rats exposed to carbon nanotubes","volume":"43","author":"Cesta","year":"2010","journal-title":"Am J Respir Cell Mol Biol"},{"key":"B42","doi-asserted-by":"publisher","first-page":"62","DOI":"10.1016\/j.ejpb.2023.01.013","article-title":"Air-liquid interface (ALI) impact on different respiratory cell cultures","volume":"184","author":"Silva","year":"2023","journal-title":"Eur J Pharm Biopharm"},{"key":"B43","doi-asserted-by":"publisher","first-page":"231","DOI":"10.1186\/s12931-024-02855-6","article-title":"Precision cut lung slices: an integrated ex vivo model for studying lung physiology, pharmacology, disease pathogenesis and drug discovery","volume":"25","author":"Koziol-White","year":"2024","journal-title":"Respir Res"}],"container-title":["Frontiers in Immunology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fimmu.2026.1794151\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,7,15]],"date-time":"2026-07-15T04:44:29Z","timestamp":1784090669000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fimmu.2026.1794151\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,7,15]]},"references-count":43,"alternative-id":["10.3389\/fimmu.2026.1794151"],"URL":"https:\/\/doi.org\/10.3389\/fimmu.2026.1794151","relation":{},"ISSN":["1664-3224"],"issn-type":[{"value":"1664-3224","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,7,15]]},"article-number":"1794151"}}