首页|Characterization of long-term ex vivo expansion of tree shrew spermatogonial stem cells

Characterization of long-term ex vivo expansion of tree shrew spermatogonial stem cells

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Tree shrews(Tupaia belangeri chinensis)share a close relationship to primates and have been widely used in biomedical research.We previously established a spermatogonial stem cell(SSC)-based gene editing platform to generate transgenic tree shrews.However,the influences of long-term expansion on tree shrew SSC spermatogenesis potential remain unclear.Here,we examined the in vivo spermatogenesis potential of tree shrew SSCs cultured across different passages.We found that SSCs lost spermatogenesis ability after long-term expansion(＞50 passages),as indicated by the failure to colonize the seminiferous epithelium and generate donor spermatogonia(SPG)-derived spermatocytes or spermatids marking spermatogenesis.RNA sequencing(RNA-seq)analysis of undifferentiated SPGs across different passages revealed significant gene expression changes after sub-culturing primary SPG lines for more than 40 passages on feeder layers.Specifically,DNA damage response and repair genes(e.g.,MRE11,SMC3,BLM,and GEN1)were down-regulated,whereas genes associated with mitochondrial function(e.g.,NDUFA9,NDUFA8,NDUFA13,and NDUFB8)were up-regulated after expansion.The DNA damage accumulation and mitochondrial dysfunction were experimentally validated in high-passage cells.Supplementation with nicotinamide adenine dinucleotide(NAD+)precursor nicotinamide riboside(NR)exhibited beneficial effects by reducing DNA damage accumulation and mitochondrial dysfunction in SPG elicited by long-term culture.Our research presents a comprehensive analysis of the genetic and physiological attributes critical for the sustained expansion of undifferentiated SSCs in tree shrews and proposes an effective strategy for extended in vitro maintenance.

Tree shrewsSpermatogonial stem cellsNicotinamide ribosideDNA damageMitochondrial dysfunction

Cong Li、Rui Bi、Lin Wang、Yu-Hua Ma、Yong-Gang Yao、Ping Zheng

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State Key Laboratory of Genetic Resources and Evolution,Kunming Institute of Zoology,Chinese Academy of Sciences,Kunming,Yunnan 650201,China

Key Laboratory of Animal Models and Human Disease Mechanisms of Yunnan Province,Kunming Institute of Zoology,Chinese Academy of Sciences,Kunming,Yunnan 650201,China

Kunming College of Life Science,University of Chinese Academy of Sciences,Kunming,Yunnan 650204,China

National Resource Center for Non-Human Primates,National Research Facility for Phenotypic & Genetic Analysis of Model Animals(Primate Facility),Kunming Institute of Zoology,Chinese Academy of Sciences,Kunming,Yunnan 650107,China

KIZ/CUHK Joint L

National Resource Center for Non-Human Primates,National Research Facility for Phenotypic & Genetic Anal

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Ministry of Science and Technology of ChinaMinistry of Science and Technology of ChinaNational Natural Science Foundation of ChinaNational Natural Science Foundation of ChinaYunnan Province

2021YFF0702700STI2030-Major Project 2021ZD0200900U2102202U1702284202305AH340006

2023

10.24272/j.issn.2095-8137.2023.317

动物学研究
中国科学院昆明动物研究所 中国动物学会

动物学研究

CSTPCDCSCD
影响因子：0.582
ISSN：0254-5853
年,卷(期)：2023.44(6)
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