Life-long functional regeneration of in vivo airway epithelium by the engraftment of airway basal stem cells.

IF 13.1 1区 生物学 Q1 BIOCHEMICAL RESEARCH METHODS Nature Protocols Pub Date : 2024-11-05 DOI:10.1038/s41596-024-01067-y
Liang Ma, Bibek R Thapa, Jake A Le Suer, Andrew Tilston-Lünel, Michael J Herriges, Feiya Wang, Pushpinder S Bawa, Xaralabos Varelas, Finn J Hawkins, Darrell N Kotton
{"title":"Life-long functional regeneration of in vivo airway epithelium by the engraftment of airway basal stem cells.","authors":"Liang Ma, Bibek R Thapa, Jake A Le Suer, Andrew Tilston-Lünel, Michael J Herriges, Feiya Wang, Pushpinder S Bawa, Xaralabos Varelas, Finn J Hawkins, Darrell N Kotton","doi":"10.1038/s41596-024-01067-y","DOIUrl":null,"url":null,"abstract":"<p><p>Durable and functional regeneration of the airway epithelium in vivo with transplanted stem cells has the potential to reconstitute healthy tissue in diseased airways, such as in cystic fibrosis or primary ciliary dyskinesia. Here, we present detailed protocols for the preparation and culture expansion of murine primary and induced pluripotent stem cell-derived airway basal stem cells (iBCs) and methods for their intra-airway transplantation into polidocanol-conditioned murine recipients to achieve durable in vivo airway regeneration. Reconstitution of the airway tissue resident epithelial stem cell compartment of immunocompetent mice with syngeneic donor cells leverages the extensive self-renewal and multipotent differentiation properties of basal stem cells (BCs) to durably generate a broad diversity of mature airway epithelial lineages in vivo. Engrafted donor-derived cells re-establish planar cell polarity as well as functional ciliary transport. By using this same approach, human primary BCs or iBCs transplanted into NOD-SCID gamma recipient mice similarly display engraftment and multilineage airway epithelial differentiation in vivo. The time to generate mouse or human iBCs takes ~60 d, which can be reduced to ~20 d if previously differentiated cells are thawed from cryopreserved iBC archives. The tracheal conditioning regimen and cell transplantation procedure is completed in 1 d. A competent graduate student or postdoctoral trainee should be able to perform the procedures listed in this protocol.</p>","PeriodicalId":18901,"journal":{"name":"Nature Protocols","volume":null,"pages":null},"PeriodicalIF":13.1000,"publicationDate":"2024-11-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Nature Protocols","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1038/s41596-024-01067-y","RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
引用次数: 0

Abstract

Durable and functional regeneration of the airway epithelium in vivo with transplanted stem cells has the potential to reconstitute healthy tissue in diseased airways, such as in cystic fibrosis or primary ciliary dyskinesia. Here, we present detailed protocols for the preparation and culture expansion of murine primary and induced pluripotent stem cell-derived airway basal stem cells (iBCs) and methods for their intra-airway transplantation into polidocanol-conditioned murine recipients to achieve durable in vivo airway regeneration. Reconstitution of the airway tissue resident epithelial stem cell compartment of immunocompetent mice with syngeneic donor cells leverages the extensive self-renewal and multipotent differentiation properties of basal stem cells (BCs) to durably generate a broad diversity of mature airway epithelial lineages in vivo. Engrafted donor-derived cells re-establish planar cell polarity as well as functional ciliary transport. By using this same approach, human primary BCs or iBCs transplanted into NOD-SCID gamma recipient mice similarly display engraftment and multilineage airway epithelial differentiation in vivo. The time to generate mouse or human iBCs takes ~60 d, which can be reduced to ~20 d if previously differentiated cells are thawed from cryopreserved iBC archives. The tracheal conditioning regimen and cell transplantation procedure is completed in 1 d. A competent graduate student or postdoctoral trainee should be able to perform the procedures listed in this protocol.

查看原文
分享 分享
微信好友 朋友圈 QQ好友 复制链接
本刊更多论文
通过气道基底干细胞的移植,实现体内气道上皮细胞的终身功能再生。
通过移植干细胞在体内实现气道上皮的持久和功能性再生,有可能在囊性纤维化或原发性纤毛运动障碍等疾病的气道中重建健康组织。在此,我们介绍了小鼠原代干细胞和诱导多能干细胞衍生气道基底干细胞(iBCs)的制备和培养扩增的详细方案,以及将其在气道内移植到经多聚糖醇调节的小鼠受体中以实现持久的体内气道再生的方法。用合成供体细胞重建免疫功能正常小鼠的气道组织常驻上皮干细胞区系,可利用基底干细胞(BCs)广泛的自我更新和多能分化特性,在体内持久产生多种成熟的气道上皮细胞系。移植的供体衍生细胞可重建平面细胞极性和功能性纤毛运输。采用同样的方法,将人类原代 BCs 或 iBCs 移植到 NOD-SCID γ 受体小鼠体内,同样会出现移植和多系气道上皮分化。生成小鼠或人类 iBCs 的时间约为 60 d,如果从低温保存的 iBC 档案中解冻先前分化的细胞,则时间可缩短至约 20 d。有能力的研究生或博士后受训者应能完成本方案中列出的程序。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 去求助
来源期刊
Nature Protocols
Nature Protocols 生物-生化研究方法
CiteScore
29.10
自引率
0.70%
发文量
128
审稿时长
4 months
期刊介绍: Nature Protocols focuses on publishing protocols used to address significant biological and biomedical science research questions, including methods grounded in physics and chemistry with practical applications to biological problems. The journal caters to a primary audience of research scientists and, as such, exclusively publishes protocols with research applications. Protocols primarily aimed at influencing patient management and treatment decisions are not featured. The specific techniques covered encompass a wide range, including but not limited to: Biochemistry, Cell biology, Cell culture, Chemical modification, Computational biology, Developmental biology, Epigenomics, Genetic analysis, Genetic modification, Genomics, Imaging, Immunology, Isolation, purification, and separation, Lipidomics, Metabolomics, Microbiology, Model organisms, Nanotechnology, Neuroscience, Nucleic-acid-based molecular biology, Pharmacology, Plant biology, Protein analysis, Proteomics, Spectroscopy, Structural biology, Synthetic chemistry, Tissue culture, Toxicology, and Virology.
期刊最新文献
Bio-orthogonal tuning of matrix properties during 3D cell culture to induce morphological and phenotypic changes. Life-long functional regeneration of in vivo airway epithelium by the engraftment of airway basal stem cells. Design, performance, processing, and validation of a pooled CRISPR perturbation screen for bacterial toxins. Seq-Scope: repurposing Illumina sequencing flow cells for high-resolution spatial transcriptomics. Outcome devaluation as a method for identifying goal-directed behaviors in rats.
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
现在去查看 取消
×
提示
确定
0
微信
客服QQ
Book学术公众号 扫码关注我们
反馈
×
意见反馈
请填写您的意见或建议
请填写您的手机或邮箱
已复制链接
已复制链接
快去分享给好友吧!
我知道了
×
扫码分享
扫码分享
Book学术官方微信
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术
文献互助 智能选刊 最新文献 互助须知 联系我们:info@booksci.cn
Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。
Copyright © 2023 Book学术 All rights reserved.
ghs 京公网安备 11010802042870号 京ICP备2023020795号-1