Long-term propagation of tree shrew spermatogonial stem cells in culture and successful generation of transgenic offspring View Full Text


Ontology type: schema:ScholarlyArticle      Open Access: True


Article Info

DATE

2017-02

AUTHORS

Chao-Hui Li, Lan-Zhen Yan, Wen-Zan Ban, Qiu Tu, Yong Wu, Lin Wang, Rui Bi, Shuang Ji, Yu-Hua Ma, Wen-Hui Nie, Long-Bao Lv, Yong-Gang Yao, Xu-Dong Zhao, Ping Zheng

ABSTRACT

Tree shrews have a close relationship to primates and have many advantages over rodents in biomedical research. However, the lack of gene manipulation methods has hindered the wider use of this animal. Spermatogonial stem cells (SSCs) have been successfully expanded in culture to permit sophisticated gene editing in the mouse and rat. Here, we describe a culture system for the long-term expansion of tree shrew SSCs without the loss of stem cell properties. In our study, thymus cell antigen 1 was used to enrich tree shrew SSCs. RNA-sequencing analysis revealed that the Wnt/β-catenin signaling pathway was active in undifferentiated SSCs, but was downregulated upon the initiation of SSC differentiation. Exposure of tree shrew primary SSCs to recombinant Wnt3a protein during the initial passages of culture enhanced the survival of SSCs. Use of tree shrew Sertoli cells, but not mouse embryonic fibroblasts, as feeder was found to be necessary for tree shrew SSC proliferation, leading to a robust cell expansion and long-term culture. The expanded tree shrew SSCs were transfected with enhanced green fluorescent protein (EGFP)-expressing lentiviral vectors. After transplantation into sterilized adult male tree shrew's testes, the EGFP-tagged SSCs were able to restore spermatogenesis and successfully generate transgenic offspring. Moreover, these SSCs were suitable for the CRISPR/Cas9-mediated gene modification. The development of a culture system to expand tree shrew SSCs in combination with a gene editing approach paves the way for precise genome manipulation using the tree shrew. More... »

PAGES

241

Identifiers

URI

http://scigraph.springernature.com/pub.10.1038/cr.2016.156

DOI

http://dx.doi.org/10.1038/cr.2016.156

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1004507618

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/28008926


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