{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,23]],"date-time":"2026-10-23T20:50:13Z","timestamp":1792788613272,"version":"4.5.0"},"reference-count":58,"publisher":"Public Library of Science (PLoS)","issue":"11","license":[{"start":{"date-parts":[[2024,11,15]],"date-time":"2024-11-15T00:00:00Z","timestamp":1731628800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001691","name":"Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["21K15460"],"award-info":[{"award-number":["21K15460"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["21H02891"],"award-info":[{"award-number":["21H02891"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["22K19120"],"award-info":[{"award-number":["22K19120"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Program on Open Innovation Platforms for Industry-academia Co-creation","award":["JPMJPF2108"],"award-info":[{"award-number":["JPMJPF2108"]}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS ONE"],"abstract":"<jats:p>Live imaging visualizes the structure, dynamics, and function of cells and tissues to reveal the molecular mechanisms, and has contributed to the advancement of life science. In live imaging, it has been well known that there is a trade-off between higher-resolution analysis and cell damage caused by light illumination, i.e., phototoxicity. However, despite the risk of unknowingly distorting experimental results, phototoxicity is an unresolved issue in live imaging because overall consequences occurring inside cells due to phototoxicity remains unknown. Here, we determined the molecular process of phototoxicity-induced cell damage systematically under low- and high-dose light illumination conditions by analyzing differential gene expression using RNA-sequencing in a three-dimensional organoid of small intestinal epithelial cells, enteroid. The low-dose light illumination already induced various abnormalities in functional molecules involved in the response to reactive oxygen species generated by the excitation of fluorescent dyes, intracellular metabolism, mitosis, immune responses, etc., at mRNA expression level. Together with the behavior toward apoptosis caused by high-dose light illumination, the light dose-dependent progression of intracellular damage was revealed. About visible impairment of intestinal epithelial function, failures in both the structure-forming ability of enteroids and Paneth cell granule secretion were observed under high-dose light illumination, while the drug efflux was not disturbed despite abnormal drug efflux transporter mRNA expression. Based on the gene expression profiles, we comprehensively clarified phenomena in the cells at mRNA level that cannot be recognized both morphologically and functionally during live imaging, further providing a new insight into the risk of phototoxicity. This study warns from the aspect of mRNA expression that awareness of phototoxic artifacts is needed when analyzing cellular function and the mechanism in live imaging.<\/jats:p>","DOI":"10.1371\/journal.pone.0313213","type":"journal-article","created":{"date-parts":[[2024,11,15]],"date-time":"2024-11-15T13:31:08Z","timestamp":1731677468000},"page":"e0313213","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":15,"title":["Potential consequences of phototoxicity on cell function during live imaging of intestinal organoids"],"prefix":"10.1371","volume":"19","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-8077-3200","authenticated-orcid":true,"given":"Yuki","family":"Yokoi","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ryu","family":"Nakamura","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4326-9707","authenticated-orcid":true,"given":"Shuya","family":"Ohira","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shota","family":"Takemi","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tokiyoshi","family":"Ayabe","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2138-1733","authenticated-orcid":true,"given":"Kiminori","family":"Nakamura","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2024,11,15]]},"reference":[{"key":"pone.0313213.ref001","first-page":"10","article-title":"Circumventing photodamage in live-cell microscopy","volume":"114","author":"V Magidson","year":"2013","journal-title":"Methods Cell Biol"},{"key":"pone.0313213.ref002","doi-asserted-by":"crossref","first-page":"1700003","DOI":"10.1002\/bies.201700003","article-title":"Phototoxicity in live fluorescence microscopy, and how to avoid it","volume":"39","author":"J Icha","year":"2017","journal-title":"BioEssays"},{"key":"pone.0313213.ref003","doi-asserted-by":"crossref","first-page":"751","DOI":"10.1038\/s41592-018-0170-4","article-title":"Phototoxicity revisited","volume":"15","year":"2018","journal-title":"Nat Methods"},{"key":"pone.0313213.ref004","doi-asserted-by":"crossref","first-page":"280","DOI":"10.1046\/j.1365-313X.2003.01868.x","article-title":"Cell damage and reactive oxygen species production induced by fluorescence microscopy: effect on mitosis and guidelines for non-invasive fluorescence microscopy","volume":"36","author":"R Dixit","year":"2003","journal-title":"Plant J"},{"key":"pone.0313213.ref005","doi-asserted-by":"crossref","first-page":"657","DOI":"10.1038\/nmeth.4344","article-title":"Assessing phototoxicity in live fluorescence imaging","volume":"14","author":"PP Laissue","year":"2017","journal-title":"Nat Methods"},{"key":"pone.0313213.ref006","doi-asserted-by":"crossref","first-page":"15348","DOI":"10.1038\/srep15348","article-title":"Light-induced cell damage in live-cell super-resolution microscopy","volume":"5","author":"S W\u00e4ldchen","year":"2015","journal-title":"Sci Rep"},{"key":"pone.0313213.ref007","doi-asserted-by":"crossref","first-page":"2461","DOI":"10.1002\/jcp.25588","article-title":"Live Cell Imaging: Assessing the Phototoxicity of 488 and 546 nm Light and Methods to Alleviate it","volume":"232","author":"S Douthwright","year":"2017","journal-title":"J Cell Physiol"},{"key":"pone.0313213.ref008","doi-asserted-by":"crossref","DOI":"10.3389\/fcell.2021.738786","article-title":"Assessing Phototoxicity in a Mammalian Cell Line: How Low Levels of Blue Light Affect Motility in PC3 Cells","volume":"9","author":"RA Alghamdi","year":"2021","journal-title":"Front Cell Dev Biol"},{"key":"pone.0313213.ref009","doi-asserted-by":"crossref","first-page":"e0128898","DOI":"10.1371\/journal.pone.0128898","article-title":"Reliable Screening of Dye Phototoxicity by Using a Caenorhabditis elegans Fast Bioassay","volume":"10","author":"JI Bianchi","year":"2015","journal-title":"PLOS ONE"},{"key":"pone.0313213.ref010","doi-asserted-by":"crossref","first-page":"1586","DOI":"10.1016\/j.cell.2016.05.082","article-title":"Modeling Development and Disease with Organoids","volume":"165","author":"H. 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