{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,24]],"date-time":"2026-04-24T17:50:45Z","timestamp":1777053045058,"version":"3.51.4"},"reference-count":35,"publisher":"Public Library of Science (PLoS)","issue":"4","license":[{"start":{"date-parts":[[2026,4,24]],"date-time":"2026-04-24T00:00:00Z","timestamp":1776988800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Grant-in-Aid for Scientific Research (KAKENHI) from the Japan Society for the Promotion of Science","award":["19K08731"],"award-info":[{"award-number":["19K08731"]}]},{"name":"Grant-in-Aid for Scientific Research (KAKENHI) from the Japan Society for the Promotion of Science","award":["24K11435"],"award-info":[{"award-number":["24K11435"]}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:p>\n                    Chronic kidney disease progression involves phenotypic changes in tubular epithelial cells, and recent studies have highlighted the relationship between such pro-fibrotic phenotypes and cell cycle arrest in injured tubular epithelial cells undergoing repair. We investigated these processes using a mouse unilateral ischemia\u2013reperfusion injury model with \u03b3GT.Fucci2aR mice, which express fluorescent cell cycle markers, and\n                    <jats:italic>in vitro<\/jats:italic>\n                    experiments with human kidney-2 cells and public single-cell RNA sequencing data. In the unilateral ischemia\u2013reperfusion injury model, mVenus-positive cells (S\/G2\/M phases) in the \u03b3GT.Fucci2aR mice peaked on Day 3 post-injury, then rapidly declined. In kidneys with progressive tubular atrophy and interstitial fibrosis between 7\u201312 days post-injury, S\/G2\/M phase cells were limited. These findings were corroborated by analysis using public single-cell RNA sequencing data from the same mouse models, which confirmed dynamic cell cycle changes in the acute phase post-injury but no significant G2\/M phase cells in the chronic phase.\n                    <jats:italic>In vitro<\/jats:italic>\n                    experiments with human kidney-2 cells demonstrated that cellular communication network factor 2 and transforming growth factor-\u03b2 expression increased significantly as proliferating cells reached confluence and cell cycle progression slowed. Pro-fibrotic phenotypes in tubular epithelial cells were not exclusively acquired by G2\/M-arrested cells, as reported in previous studies, but can also be acquired by cells in the quiescent G0\/G1 phase during normal cell cycling. To develop novel therapeutics for chronic kidney disease, regulating pro-fibrotic gene expression in injured tubular epithelial cells, independent of specific cell cycle phases, appears to be crucial.\n                  <\/jats:p>","DOI":"10.1371\/journal.pone.0347835","type":"journal-article","created":{"date-parts":[[2026,4,24]],"date-time":"2026-04-24T17:32:33Z","timestamp":1777051953000},"page":"e0347835","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":0,"title":["Reappraisal of profibrotic phenotype and cell-cycle state of renal tubular epithelium after ischemia\u2013reperfusion 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Inoue","year":"2015","journal-title":"Kidney Int"},{"issue":"3","key":"pone.0347835.ref022","doi-asserted-by":"crossref","first-page":"350","DOI":"10.1007\/s10157-014-1011-0","article-title":"Effects of cell-type-specific expression of a pan-caspase inhibitor on renal fibrogenesis","volume":"19","author":"T Inoue","year":"2015","journal-title":"Clin Exp Nephrol"},{"issue":"9","key":"pone.0347835.ref023","doi-asserted-by":"crossref","first-page":"989","DOI":"10.1038\/nm.3901","article-title":"Snail1-induced partial epithelial-to-mesenchymal transition drives renal fibrosis in mice and can be targeted to reverse established disease","volume":"21","author":"MT Grande","year":"2015","journal-title":"Nat Med"},{"key":"pone.0347835.ref024","doi-asserted-by":"crossref","first-page":"2","DOI":"10.1186\/1471-2121-12-2","article-title":"Drug-induced cell cycle modulation leading to cell-cycle arrest, nuclear mis-segregation, or endoreplication","volume":"12","author":"A Sakaue-Sawano","year":"2011","journal-title":"BMC Cell Biol"},{"issue":"3","key":"pone.0347835.ref025","doi-asserted-by":"crossref","first-page":"487","DOI":"10.1016\/j.cell.2007.12.033","article-title":"Visualizing spatiotemporal dynamics of multicellular cell-cycle progression","volume":"132","author":"A Sakaue-Sawano","year":"2008","journal-title":"Cell"},{"issue":"1","key":"pone.0347835.ref026","doi-asserted-by":"crossref","first-page":"5805","DOI":"10.1038\/s41467-022-33110-5","article-title":"Tubular cell polyploidy protects from lethal acute kidney injury but promotes consequent chronic kidney disease","volume":"13","author":"L De Chiara","year":"2022","journal-title":"Nat Commun"},{"issue":"1","key":"pone.0347835.ref027","doi-asserted-by":"crossref","first-page":"1344","DOI":"10.1038\/s41467-018-03753-4","article-title":"Endocycle-related tubular cell hypertrophy and progenitor proliferation recover renal function after acute kidney injury","volume":"9","author":"E Lazzeri","year":"2018","journal-title":"Nat Commun"},{"key":"pone.0347835.ref028","first-page":"536","article-title":"Kidney tubular cell regeneration starts in the deep cortex after ischemia","volume":"27","author":"JU Kim","year":"2008","journal-title":"Kidney Research and Clinical Practice"},{"issue":"8","key":"pone.0347835.ref029","doi-asserted-by":"crossref","DOI":"10.1152\/ajprenal.00576.2013","article-title":"Remote conditioning or erythropoietin before surgery primes kidneys to clear ischemia-reperfusion-damaged cells: a renoprotective mechanism?","volume":"306","author":"DS Gardner","year":"2014","journal-title":"Am J Physiol Renal Physiol"},{"issue":"21","key":"pone.0347835.ref030","doi-asserted-by":"crossref","first-page":"12373","DOI":"10.1074\/jbc.270.21.12373","article-title":"A human keratinocyte cell line produces two autocrine growth inhibitors, transforming growth factor-beta and insulin-like growth factor binding protein-6, in a calcium- and cell density-dependent manner","volume":"270","author":"M Kato","year":"1995","journal-title":"J Biol Chem"},{"issue":"1","key":"pone.0347835.ref031","doi-asserted-by":"crossref","first-page":"61","DOI":"10.1038\/s41392-024-01764-w","article-title":"TGF-\u03b2 signaling in health, disease, and therapeutics","volume":"9","author":"Z Deng","year":"2024","journal-title":"Signal Transduct Target Ther"},{"issue":"4","key":"pone.0347835.ref032","doi-asserted-by":"crossref","first-page":"512","DOI":"10.1002\/(SICI)1097-0215(19991112)83:4<512::AID-IJC13>3.0.CO;2-Z","article-title":"TGF-beta-induced cell-cycle arrest through the p21(WAF1\/CIP1)-G1 cyclin\/Cdks-p130 pathway in gastric-carcinoma cells","volume":"83","author":"YD Yoo","year":"1999","journal-title":"Int J Cancer"},{"issue":"1","key":"pone.0347835.ref033","doi-asserted-by":"crossref","first-page":"119","DOI":"10.1016\/j.jdsr.2020.07.001","article-title":"Roles of CCN2 as a mechano-sensing regulator of chondrocyte differentiation","volume":"56","author":"T Nishida","year":"2020","journal-title":"Jpn Dent Sci Rev"},{"issue":"11","key":"pone.0347835.ref034","doi-asserted-by":"crossref","first-page":"587","DOI":"10.1302\/2046-3758.711.BJR-2018-0057.R1","article-title":"Mechanical stress contributes to osteoarthritis development through the activation of transforming growth factor beta 1 (TGF-\u03b21)","volume":"7","author":"R-K Zhang","year":"2018","journal-title":"Bone Joint Res"},{"issue":"1","key":"pone.0347835.ref035","doi-asserted-by":"crossref","first-page":"111029","DOI":"10.1016\/j.jbc.2025.111029","article-title":"PKM2 inhibitor suppresses kidney fibrogenesis by disrupting YAP-TEAD-CCN2 transcriptional signaling following ischemia-reperfusion injury","volume":"302","author":"W Kosakai","year":"2026","journal-title":"J Biol Chem"}],"container-title":["PLOS 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