鸡永生化骨骼肌卫星细胞系的建立及分析

doi:10.1016/j.jia.2024.01.034

Journal of Integrative Agriculture ›› 2025, Vol. 24 ›› Issue (11): 4370-4378.DOI: 10.1016/j.jia.2024.01.034

鸡永生化骨骼肌卫星细胞系的建立及分析

收稿日期:2023-09-10 修回日期:2024-01-24 接受日期:2023-11-28 出版日期:2025-11-20 发布日期:2025-10-17

Establishment and analysis of a chicken skeletal muscle satellite cell line using TERT

Yanxing Wang¹, Haigang Ji¹, Liyang He¹, Yufang Niu¹, Yushi Zhang¹, Yang Liu^1,, Yadong Tian^1,2, Xiaojun Liu^1,2, Hong Li^1,2, Xiangtao Kang^1,2, Yanling Gao^3#, Zhuanjian Li^1,2#

¹College of Animal Science and Technology, Henan Agricultural University, Zhengzhou 450046, China
²Key Laboratory of Livestock and Poultry Resources (Poultry) Evaluation and Utilization, Ministry of Agriculture and Rural Affairs, Zhengzhou 450046, China
³Henan Vocational College of Agriculture, Zhengzhou 451450, China

Received:2023-09-10 Revised:2024-01-24 Accepted:2023-11-28 Online:2025-11-20 Published:2025-10-17
About author:Yanxing Wang, E-mail: m18700258066@163.com; #Correspondence Zhuanjian Li, E-mail: lizhuanjian@163.com; Yanling Gao, E-mail: hnivdc@163.com
Supported by:
This work was supported by the grants from the National Natural Science Foundation of China (32372873, 32441084 and 32172720), the Program for Science & Technology Innovation Talents in Universities of Henan Province, China (22HASTIT038) and the Zhongyuan Youth Talent Support Program of China (ZYYCYU202012156).

摘要/Abstract

摘要：

骨骼肌卫星细胞作为肌源性干细胞，具有增殖、分化和自我更新的能力。然而，原代骨骼肌卫星细胞在体外的增殖能力有限、分离步骤繁琐等因素限制了其在骨骼肌发育研究中的应用。细胞永生化能够使原代细胞越过细胞衰老期获得持续增殖的能力，永生化鸡骨骼肌卫星细胞系（ICMS）的建立将为鸡骨骼肌相关基因的功能研究提供可靠的细胞模型。在本研究中，我们利用慢病毒包装鸡端粒酶逆转录酶（chTERT）感染原代细胞，筛选出慢病毒的最适MOI和杀稻瘟菌素的最适浓度；随后，对感染的细胞进行药物筛选，获得阳性细胞后连续传代培养，通过端粒酶活性的重建以实现鸡骨骼肌卫星细胞的永生化。研究结果显示，传代至第10代、第15代、第22代的永生化鸡骨骼肌卫星（P10 ICMS、P15 ICMS、P22 ICMS）细胞形态与鸡原代骨骼肌卫星细胞（CPMSCs）保持一致，均呈纤维样状；与CPMSCs相比，P10 ICMS的生长曲线与CPMSCs相似；P10 ICMS具有正常的细胞增殖周期，且处于S期的细胞数量极显著高于CPMSCs；P10 ICMS复苏后的细胞活性较好；当血清浓度增加到15%时，P10、P22 ICMS可以增殖，说明ICMS具有一定的血清依赖性；软琼脂试验表明，培养14 d后，P10 ICMS依旧为单独的细胞，没有细胞克隆团的形成，可见ICMS没有发生恶性转化；经2%孕马血清的诱导，P22 ICMS的增殖能力较强、分化能力较弱。综上所述，本研究首次建立了鸡永生化骨骼肌卫星细胞系，其保持了与原代细胞相似的生物学特性，无致瘤性转化。鸡永生化骨骼肌卫星细胞系的建立为chTERT构建家禽细胞的永生化提供了参考，为家禽骨骼肌发育的研究提供了理想的体外细胞模型。

Abstract:

Skeletal muscle satellite cells are stem cells characterized by their multipotency and capacity for in vitro proliferation. However, primary skeletal muscle satellite cells demonstrate limited proliferative capacity in vitro, which impedes their investigation in poultry skeletal muscle research. Cell immortalization techniques have emerged as valuable tools to address this limitation and facilitate the study of skeletal muscle satellite cell functions. This study achieved the immortalization of chicken skeletal muscle satellite cells through the transduction of primary cells with TERT (telomerase reverse transcriptase) amplified from chicken (chTERT) using a lentiviral vector via telomerase activity reconstitution. While the cells successfully overcame replicative senescence, complete immortalization was not achieved. Initial functional characterization revealed that the proliferative properties and cell cycle characteristics of the immortalized chicken skeletal muscle satellite cell lines (ICMS) closely resembled those of chicken primary muscle satellite cells (CPMSCs). Serum dependency analysis and soft agar assays confirmed that ICMS did not undergo malignant transformation. Furthermore, induced differentiation experiments demonstrated preserved differentiation capacity in ICMS. The established cell lines provide a fundamental framework for developing immortalized poultry cell lines and offer a cellular model for investigating poultry skeletal muscle-related functional genes.

Key words: chicken , skeletal muscle satellite cell , immortalization , chTERT , proliferation , differentiation

Yanxing Wang, Haigang Ji, Liyang He, Yufang Niu, Yushi Zhang, Yang Liu, Yadong Tian, Xiaojun Liu, Hong Li, Xiangtao Kang, Yanling Gao, Zhuanjian Li. 鸡永生化骨骼肌卫星细胞系的建立及分析[J]. Journal of Integrative Agriculture, 2025, 24(11): 4370-4378.

Yanxing Wang, Haigang Ji, Liyang He, Yufang Niu, Yushi Zhang, Yang Liu, Yadong Tian, Xiaojun Liu, Hong Li, Xiangtao Kang, Yanling Gao, Zhuanjian Li. Establishment and analysis of a chicken skeletal muscle satellite cell line using TERT[J]. Journal of Integrative Agriculture, 2025, 24(11): 4370-4378.

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鸡永生化骨骼肌卫星细胞系的建立及分析

Establishment and analysis of a chicken skeletal muscle satellite cell line using TERT

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