Compositional editing of extracellular matrices by CRISPR/Cas9 engineering of human mesenchymal stem cell lines.

IF 6.4 1区 生物学 Q1 BIOLOGY
eLife Pub Date : 2025-03-28 DOI:10.7554/eLife.96941
Sujeethkumar Prithiviraj, Alejandro Garcia Garcia, Karin Linderfalk, Bai Yiguang, Sonia Ferveur, Ludvig Nilsén Falck, Agatheeswaran Subramaniam, Sofie Mohlin, David Hidalgo Gil, Steven J Dupard, Dimitra Zacharaki, Deepak Bushan Raina, Paul E Bourgine
{"title":"Compositional editing of extracellular matrices by CRISPR/Cas9 engineering of human mesenchymal stem cell lines.","authors":"Sujeethkumar Prithiviraj, Alejandro Garcia Garcia, Karin Linderfalk, Bai Yiguang, Sonia Ferveur, Ludvig Nilsén Falck, Agatheeswaran Subramaniam, Sofie Mohlin, David Hidalgo Gil, Steven J Dupard, Dimitra Zacharaki, Deepak Bushan Raina, Paul E Bourgine","doi":"10.7554/eLife.96941","DOIUrl":null,"url":null,"abstract":"<p><p>Tissue engineering strategies predominantly rely on the production of living substitutes, whereby implanted cells actively participate in the regenerative process. Beyond cost and delayed graft availability, the patient-specific performance of engineered tissues poses serious concerns on their clinical translation ability. A more exciting paradigm consists in exploiting cell-laid, engineered extracellular matrices (eECMs), which can be used as off-the-shelf materials. Here, the regenerative capacity solely relies on the preservation of the eECM structure and embedded signals to instruct an endogenous repair. We recently described the possibility to exploit custom human stem cell lines for eECM manufacturing. In addition to the conferred standardization, the availability of such cell lines opened avenues for the design of tailored eECMs by applying dedicated genetic tools. In this study, we demonstrated the exploitation of CRISPR/Cas9 as a high precision system for editing the composition and function of eECMs. Human mesenchymal stromal/stem cell (hMSC) lines were modified to knock out vascular endothelial growth factor (VEGF) and Runt-related transcription factor 2 (RUNX2) and assessed for their capacity to generate osteoinductive cartilage matrices. We report the successful editing of hMSCs, subsequently leading to targeted VEGF and RUNX2-knockout cartilage eECMs. Despite the absence of VEGF, eECMs retained full capacity to instruct ectopic endochondral ossification. Conversely, RUNX2-edited eECMs exhibited impaired hypertrophy, reduced ectopic ossification, and superior cartilage repair in a rat osteochondral defect. In summary, our approach can be harnessed to identify the necessary eECM factors driving endogenous repair. Our work paves the road toward the compositional eECMs editing and their exploitation in broad regenerative contexts.</p>","PeriodicalId":11640,"journal":{"name":"eLife","volume":"13 ","pages":""},"PeriodicalIF":6.4000,"publicationDate":"2025-03-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11952750/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"eLife","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.7554/eLife.96941","RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOLOGY","Score":null,"Total":0}
引用次数: 0

Abstract

Tissue engineering strategies predominantly rely on the production of living substitutes, whereby implanted cells actively participate in the regenerative process. Beyond cost and delayed graft availability, the patient-specific performance of engineered tissues poses serious concerns on their clinical translation ability. A more exciting paradigm consists in exploiting cell-laid, engineered extracellular matrices (eECMs), which can be used as off-the-shelf materials. Here, the regenerative capacity solely relies on the preservation of the eECM structure and embedded signals to instruct an endogenous repair. We recently described the possibility to exploit custom human stem cell lines for eECM manufacturing. In addition to the conferred standardization, the availability of such cell lines opened avenues for the design of tailored eECMs by applying dedicated genetic tools. In this study, we demonstrated the exploitation of CRISPR/Cas9 as a high precision system for editing the composition and function of eECMs. Human mesenchymal stromal/stem cell (hMSC) lines were modified to knock out vascular endothelial growth factor (VEGF) and Runt-related transcription factor 2 (RUNX2) and assessed for their capacity to generate osteoinductive cartilage matrices. We report the successful editing of hMSCs, subsequently leading to targeted VEGF and RUNX2-knockout cartilage eECMs. Despite the absence of VEGF, eECMs retained full capacity to instruct ectopic endochondral ossification. Conversely, RUNX2-edited eECMs exhibited impaired hypertrophy, reduced ectopic ossification, and superior cartilage repair in a rat osteochondral defect. In summary, our approach can be harnessed to identify the necessary eECM factors driving endogenous repair. Our work paves the road toward the compositional eECMs editing and their exploitation in broad regenerative contexts.

求助全文
约1分钟内获得全文 求助全文
来源期刊
eLife
eLife BIOLOGY-
CiteScore
12.90
自引率
3.90%
发文量
3122
审稿时长
17 weeks
期刊介绍: eLife is a distinguished, not-for-profit, peer-reviewed open access scientific journal that specializes in the fields of biomedical and life sciences. eLife is known for its selective publication process, which includes a variety of article types such as: Research Articles: Detailed reports of original research findings. Short Reports: Concise presentations of significant findings that do not warrant a full-length research article. Tools and Resources: Descriptions of new tools, technologies, or resources that facilitate scientific research. Research Advances: Brief reports on significant scientific advancements that have immediate implications for the field. Scientific Correspondence: Short communications that comment on or provide additional information related to published articles. Review Articles: Comprehensive overviews of a specific topic or field within the life sciences.
×
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
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
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 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信