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In vitro models have been widely used to investigate ENP toxicity. Air\u2013liquid interface (ALI) cell cultures have been emerging as a valuable alternative to submerged cultures as they are more representative of the inhalation exposure to airborne nano-sized particles. We compared the in vitro toxicity of four ENP used as raw materials in the advanced ceramics sector in human alveolar epithelial-like cells cultured under submerged or ALI conditions. Submerged cultures were exposed to ENP liquid suspensions or to aerosolised ENP at ALI. Toxicity was assessed by determining LDH release, WST-1 metabolisation and DNA damage. Overall, cells were more sensitive to ENP cytotoxic effects when cultured and exposed under ALI. No significant cytotoxicity was observed after 24 h exposure to ENP liquid suspensions, although aerosolised ENP clearly affected cell viability and LDH release. In general, all ENP increased primary DNA damage regardless of the exposure mode, where an increase in DNA strand-breaks was only detected under submerged conditions. Our data show that at relevant occupational concentrations, the selected ENP exert mild toxicity to alveolar epithelial cells and exposure at ALI might be the most suitable choice when assessing ENP toxicity in respiratory models under realistic exposure conditions.<\/jats:p>","DOI":"10.3390\/nano11123225","type":"journal-article","created":{"date-parts":[[2021,11,30]],"date-time":"2021-11-30T23:22:56Z","timestamp":1638314576000},"page":"3225","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":21,"title":["In Vitro Toxicity of Industrially Relevant Engineered Nanoparticles in Human Alveolar Epithelial Cells: Air\u2013Liquid Interface versus Submerged Cultures"],"prefix":"10.3390","volume":"11","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-5357-6167","authenticated-orcid":false,"given":"Maria Jo\u00e3o","family":"Bessa","sequence":"first","affiliation":[{"name":"Department of Environmental Health, National Institute of Health Dr. Ricardo Jorge, 4000-053 Porto, Portugal"},{"name":"EPIUnit-Instituto de Sa\u00fade P\u00fablica, Universidade do Porto, 4050-091 Porto, Portugal"},{"name":"Laborat\u00f3rio para a Investiga\u00e7\u00e3o Integrativa e Translacional em Sa\u00fade Populacional (ITR), 4050-091 Porto, Portugal"},{"name":"Instituto de Ci\u00eancias Biom\u00e9dicas Abel Salazar (ICBAS), Universidade do Porto, 4050-313 Porto, Portugal"}]},{"given":"F\u00e1tima","family":"Brand\u00e3o","sequence":"additional","affiliation":[{"name":"Department of Environmental Health, National Institute of Health Dr. Ricardo Jorge, 4000-053 Porto, Portugal"},{"name":"EPIUnit-Instituto de Sa\u00fade P\u00fablica, Universidade do Porto, 4050-091 Porto, Portugal"},{"name":"Laborat\u00f3rio para a Investiga\u00e7\u00e3o Integrativa e Translacional em Sa\u00fade Populacional (ITR), 4050-091 Porto, Portugal"},{"name":"Instituto de Ci\u00eancias Biom\u00e9dicas Abel Salazar (ICBAS), Universidade do Porto, 4050-313 Porto, Portugal"}]},{"given":"Paul H. B.","family":"Fokkens","sequence":"additional","affiliation":[{"name":"National Institute for Public Health and Environment (RIVM), 3721 Bilthoven, The Netherlands"}]},{"given":"Daan L. A. C.","family":"Leseman","sequence":"additional","affiliation":[{"name":"National Institute for Public Health and Environment (RIVM), 3721 Bilthoven, The Netherlands"}]},{"given":"A. John F.","family":"Boere","sequence":"additional","affiliation":[{"name":"National Institute for Public Health and Environment (RIVM), 3721 Bilthoven, The Netherlands"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9958-8630","authenticated-orcid":false,"given":"Flemming R.","family":"Cassee","sequence":"additional","affiliation":[{"name":"National Institute for Public Health and Environment (RIVM), 3721 Bilthoven, The Netherlands"},{"name":"Institute for Risk Assessment Sciences (IRAS), Utrecht University, 3584 Utrecht, The Netherlands"}]},{"given":"Apostolos","family":"Salmatonidis","sequence":"additional","affiliation":[{"name":"Institute of Environmental Assessment and Water Research, Spanish Research Council (IDAEA-CSIC), 08034 Barcelona, Spain"},{"name":"LEITAT Technological Center, C\/de la Innovaci\u00f3 2, 08225 Terrassa, Spain"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4073-3802","authenticated-orcid":false,"given":"Mar","family":"Viana","sequence":"additional","affiliation":[{"name":"Institute of Environmental Assessment and Water Research, Spanish Research Council (IDAEA-CSIC), 08034 Barcelona, Spain"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5334-4980","authenticated-orcid":false,"given":"Adriana","family":"Vulpoi","sequence":"additional","affiliation":[{"name":"Nanostructured Materials and Bio-Nano-Interfaces Center, Interdisciplinary Research Institute on Bio-Nano-Sciences, Babes-Bolyai University, 400271 Cluj-Napoca, Romania"}]},{"given":"Simion","family":"Simon","sequence":"additional","affiliation":[{"name":"Nanostructured Materials and Bio-Nano-Interfaces Center, Interdisciplinary Research Institute on Bio-Nano-Sciences, Babes-Bolyai University, 400271 Cluj-Napoca, Romania"}]},{"given":"Eliseo","family":"Monfort","sequence":"additional","affiliation":[{"name":"Institute of Ceramic Technology (ITC), Universitat Jaume I, 12006 Castell\u00f3n, Spain"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8693-5250","authenticated-orcid":false,"given":"Jo\u00e3o Paulo","family":"Teixeira","sequence":"additional","affiliation":[{"name":"Department of Environmental Health, National Institute of Health Dr. Ricardo Jorge, 4000-053 Porto, Portugal"},{"name":"EPIUnit-Instituto de Sa\u00fade P\u00fablica, Universidade do Porto, 4050-091 Porto, Portugal"},{"name":"Laborat\u00f3rio para a Investiga\u00e7\u00e3o Integrativa e Translacional em Sa\u00fade Populacional (ITR), 4050-091 Porto, Portugal"}]},{"given":"S\u00f3nia","family":"Fraga","sequence":"additional","affiliation":[{"name":"Department of Environmental Health, National Institute of Health Dr. Ricardo Jorge, 4000-053 Porto, Portugal"},{"name":"EPIUnit-Instituto de Sa\u00fade P\u00fablica, Universidade do Porto, 4050-091 Porto, Portugal"},{"name":"Laborat\u00f3rio para a Investiga\u00e7\u00e3o Integrativa e Translacional em Sa\u00fade Populacional (ITR), 4050-091 Porto, Portugal"}]}],"member":"1968","published-online":{"date-parts":[[2021,11,27]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Mohajerani, A., Burnett, L., Smith, J.V., Kurmus, H., Milas, J., Arulrajah, A., Horpibulsuk, S., and Kadir, A.A. 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