{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,14]],"date-time":"2026-05-14T02:44:40Z","timestamp":1778726680917,"version":"3.51.4"},"reference-count":33,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2024,1,30]],"date-time":"2024-01-30T00:00:00Z","timestamp":1706572800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2024,1,30]],"date-time":"2024-01-30T00:00:00Z","timestamp":1706572800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["82070021"],"award-info":[{"award-number":["82070021"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Quanzhou High-level Talents Major Project","award":["2020C001R"],"award-info":[{"award-number":["2020C001R"]}]},{"name":"Doctoral Miaopu project of the Second Affiliated Hospital of Fujian Medical University","award":["BS202306"],"award-info":[{"award-number":["BS202306"]}]},{"name":"the Startup Fund for Scientific Research of Fujian Medical University","award":["2019QH2040"],"award-info":[{"award-number":["2019QH2040"]}]},{"name":"the Startup Fund for Scientific Research of Fujian Medical University","award":["2022QH1107"],"award-info":[{"award-number":["2022QH1107"]}]}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Stem Cell Res Ther"],"abstract":"<jats:title>Abstract<\/jats:title><jats:sec>\n                <jats:title>Background<\/jats:title>\n                <jats:p>The stem cell characteristic makes basal cells desirable for ex vivo modeling of airway diseases. However, to date, approaches allowing them extensively in vitro serial expansion and maintaining bona fide stem cell property are still awaiting to be established. This study aims to develop a feeder-free culture system of mouse airway basal stem cells (ABSCs) that sustain their stem cell potential in vitro, providing an experimental basis for further in-depth research and mechanism exploration.<\/jats:p>\n              <\/jats:sec><jats:sec>\n                <jats:title>Methods<\/jats:title>\n                <jats:p>We used ROCK inhibitor Y-27632-containing 3T3-CM, MEF-CM, and RbEF-CM to determine the proper feeder-free culture system that could maintain\u00a0in vitro stem cell morphology of mouse ABSCs. Immunocytofluorescence was used to identify the basal cell markers of obtained cells. Serial propagation was carried out to observe whether the stem cell morphology and basal cell markers could be preserved in this cultivation system. Next, we examined the\u00a0in vitro\u00a0expansion and self-renewal ability by evaluating population doubling time and colony-forming efficiency. Moreover, the differentiation potential was detected by an\u00a0in vitro\u00a0differentiation culture and a 3D tracheosphere assay.<\/jats:p>\n              <\/jats:sec><jats:sec>\n                <jats:title>Results<\/jats:title>\n                <jats:p>When the mouse ABSCs were cultured using 3T3-CM containing ROCK inhibitor Y-27632 in combination with Matrigel-coated culture dishes, they could stably expand and maintain stem cell-like clones. We confirmed that the obtained clones comprised p63\/Krt5 double-positive ABSCs. In continuous passage and maintenance culture, we found that it could be subculture to at least 15 passages in vitro, stably maintaining its stem cell morphology, basal cell markers, and in vitro expansion and self-renewal capabilities. Meanwhile, through in vitro differentiation culture and 3D tracheosphere culture, we found that in addition to maintaining self-renewal, mouse ABSCs could differentiate into other airway epithelial cells such as acetylated tubulin (Act-Tub)\u2009+\u2009ciliated and MUC5AC\u2009+\u2009mucus-secreting cells. However, they failed to differentiate into alveoli epithelial cells, including alveolar type I and alveolar type II.<\/jats:p>\n              <\/jats:sec><jats:sec>\n                <jats:title>Conclusion<\/jats:title>\n                <jats:p>We established an\u00a0in vitro\u00a0feeder-free culture system that allows mouse ABSCs to maintain their stem cell characteristics, including self-renewal and airway epithelium differentiation potential, while keeping up\u00a0in vitro\u00a0expansion stability.<\/jats:p>\n              <\/jats:sec>","DOI":"10.1186\/s13287-024-03642-2","type":"journal-article","created":{"date-parts":[[2024,1,30]],"date-time":"2024-01-30T01:02:51Z","timestamp":1706576571000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Culture and expansion of murine proximal airway basal stem cells"],"prefix":"10.1186","volume":"15","author":[{"given":"Meirong","family":"Wu","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xiaojing","family":"Zhang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yanjuan","family":"Tu","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Wenzhao","family":"Cheng","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5635-3006","authenticated-orcid":false,"given":"Yiming","family":"Zeng","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2024,1,30]]},"reference":[{"issue":"4","key":"3642_CR1","doi-asserted-by":"publisher","first-page":"L643","DOI":"10.1152\/ajplung.00155.2003","volume":"286","author":"KU Hong","year":"2004","unstructured":"Hong KU, Reynolds SD, Watkins S, et al. 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The title of the approved project is as follows: Maintenance of the stem cell properties of airway basal cells and preclinical research on their application in lung regenerative medicine.","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Ethics approval and consent to participate"}},{"value":"Not applicable.","order":3,"name":"Ethics","group":{"name":"EthicsHeading","label":"Consent for publication"}},{"value":"The authors declare that they have no competing interests.","order":4,"name":"Ethics","group":{"name":"EthicsHeading","label":"Competing interests"}}],"article-number":"26"}}