Zhu Haijing, Zheng Liming, Wang Long, Tang Furong, Arisha Ahmed H, Zhou Hongchao, Hua Jinlian
College of Veterinary Medicine, Shaanxi Centre of Stem Cells Engineering & Technology, Northwest A&F University, Yangling, China.
Shaanxi Province Engineering and Technology Research Center of Cashmere Goat, Research Center of Life Science in Yulin University, Yulin, China.
Reprod Domest Anim. 2020 Mar;55(3):405-417. doi: 10.1111/rda.13645. Epub 2020 Feb 17.
Male germline stem cells (mGSCs) can transmit genetic materials to the next generation and dedifferentiate into pluripotent stem cells. However, in livestock, mGSC lines are difficult to establish, because of the factors that affect their isolation and culture. The extracellular matrix serves as a substrate for attachment and affects the fate of these stem cells. Poly-L-lysine (PL), an extracellular matrix of choice, inhibits and/or kills cancer cells, and promotes the attachment of stem cells in culture. However, how it affects the characteristics and potentials of these stem cells in culture needs to be elucidated. Here, we isolated, enriched and cultured dairy goat mGSCs on five types of extracellular matrices. To explore the best extracellular matrix to use for culturing them, the characteristics and proliferation ability of the cells were determined. Results showed that the cells shared several characteristics with previously reported mGSCs, including the poor effect of PL on their proliferative and colony-forming abilities. Further examination showed upregulation of p53 expression in these cells, which could be inhibiting their proliferation. When a p53 inhibitor was included in the culture medium, it was confirmed to be responsible for the inhibition of proliferation in mGSCs. Optimal concentration of the inhibitor in the culture of these cells was 5 µM. Furthermore, addition of the p53 inhibitor increased the expression of the markers of self-renewal and cell cycle in goat mGSCs. In summary, suppressing p53 is beneficial for the proliferation of dairy goat mGSCs, cultured on PL.
雄性生殖系干细胞（mGSCs）能够将遗传物质传递给下一代，并去分化为多能干细胞。然而，在牲畜中，由于影响其分离和培养的因素，mGSC系很难建立。细胞外基质作为细胞附着的底物，会影响这些干细胞的命运。聚-L-赖氨酸（PL）是一种常用的细胞外基质，它能抑制和/或杀死癌细胞，并促进培养中的干细胞附着。然而，它如何影响培养中这些干细胞的特性和潜能仍有待阐明。在此，我们在五种类型的细胞外基质上分离、富集并培养了奶山羊mGSCs。为了探索用于培养它们的最佳细胞外基质，我们测定了细胞的特性和增殖能力。结果表明，这些细胞与先前报道的mGSCs具有一些共同特征，包括PL对其增殖和集落形成能力的影响较差。进一步检测显示这些细胞中p53表达上调，这可能抑制了它们的增殖。当在培养基中加入p53抑制剂时，证实它是mGSCs增殖受抑制的原因。该抑制剂在这些细胞培养中的最佳浓度为5 μM。此外，添加p53抑制剂增加了山羊mGSCs自我更新和细胞周期标志物的表达。总之，抑制p53有利于在PL上培养的奶山羊mGSCs的增殖。
