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| Vacuum Low-temperature Preservation Combined with bFGF-supplemented Medium Enhances Somatic Cell Line Establishment Efficiency in Apeijiaza Cattle (Bos taurus) |
| ZHANG Yu-Rong1,2, WANG Jing2, WANG Qing-Wei3, GU Shi-Gang2, HUANG Yong-Ye4, YU Da-Wei2,*, XU Ye-Fen1,* |
1 College of Animal Science, Xizang Agricultural and Animal Husbandry University, Linzhi 860000, China;
2 Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100080, China;
3 College of Animal Science, Guizhou University, Guiyang 550000, China;
4 College of Life and Health Science, Northeastern University, Shenyang 110000, China |
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Abstract The Tibet Autonomous Region boasts abundant and unique animal genetic resources, serving as a critical national reserve for germplasm resources. Restricted by plateau geography and transportation conditions, animal tissue samples collected in the wild cannot be promptly transported to laboratories for processing, which severely impedes the establishment of primary fibroblast cell lines. To improve the efficiency of somatic cell line establishment from endangered livestock breeds in remote plateau regions such as Tibet, a vacuum low-temperature preservation method capable of stably maintaining ear marginal tissue at 0~4 ℃ during transport, as well as a corresponding cell line establishment system, was developed in this study. The feasibility of this approach was first validated in the indigenous Bama pig (Sus scrofa domestica). Ear marginal tissues were collected, subjected to vacuum treatment, and preserved at 4 ℃ for 0, 7, and 14 d prior to cell line initiation, and the success rate was 98%. Subsequently, this vacuum low-temperature preservation method was applied to ear marginal tissues of the Tibet-endangered Apeijiaza cattle (Bos taurus) following long-distance transportation and off-site cell line establishment. Compared with the conventional PBS-immersion method, the vacuum low-temperature preservation protocol yielded a greater number of viable cells at initiation, with cell viability consistently exceeding 85%. The success rate of primary fibroblast establishment from Apeijiaza cattle ear tissues was significantly increased by 37.5% relative to the control group (P<0.05). Furthermore, supplementation of the culture medium with 10 ng/mL basic fibroblast growth factor (bFGF) effectively retarded the senescence progression of Apeijiaza cattle somatic cells during serial passaging in vitro. Under bFGF-supplemented conditions, cells maintained high proliferative activity up to passage 20. There was no obvious apoptosis and senescence morphology, accompanied by significant downregulation of senescence-associated genes, including tumor protein p53 (TP53), insulin-like growth factor-binding protein 2 (IGFBP2), and thrombospondin-1 (THBS1). Collectively, the findings demonstrated that the combined strategy of vacuum low-temperature preservation and bFGF supplementation in culture medium effectively overcame the long-standing bottleneck associated with prolonged sample preservation of livestock genetic resources in remote areas, thereby providing a robust and reliable technological platform for the off-site establishment of somatic cell genetic resources from endangered species.
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Received: 29 January 2026
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| Fund:国家重点研发计划(2024YFD1200700); 西藏自治区重点研发计划(XZ202501ZY0031; XZ202301ZY0003N03); 中国农业科学院科技创新工程(CAAS-CSAB-202402) |
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Corresponding Authors:
*xzlzxyf@163.com; ydw023@163.com
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