Department of Animal Sciences, University of Florida, Gainesville 32611, USA.
J Anim Sci. 2011 Jun;89(6):1751-7. doi: 10.2527/jas.2010-3568. Epub 2011 Feb 4.
Satellite cells are a heterogeneous population of myogenic precursors responsible for muscle growth and repair in mammals. The objectives of the experiment were to examine the growth rates and degree of heterogeneity within bovine satellite cells (BSC) isolated from young and adult animals. The BSC were harvested from the semimembranosus of young (4.3 ± 0.5 d) and adult (estimated 24 to 27 mo) cattle and cultured en masse. Young animal BSC re-enter the cell cycle sooner and reach maximal 5-ethynyl-2'-deoxyuridine (EdU) incorporation earlier (P < 0.05) than adult contemporaries. Adult BSC contain fewer (P < 0.05) MyoD and myogenin immunopositive nuclei than BSC isolated from young animals after 3, 4, and 5 d in culture. These results indicate that BSC from young animals activate, proliferate, and differentiate sooner than isolates from adult animals. Lineage heterogeneity within BSC was examined using antibodies specific for Pax7 and Myf5, lineage markers of satellite cells, and myoblasts. Immunocytochemistry revealed the majority of Pax7-expressing BSC also express Myf5; a minor population (~5%) fails to exhibit Myf5 immunoreactivity. The percentage of Pax7:Myf5 BSC from young animals decreases sooner (P < 0.05) in culture than adult BSC, indicating a more rapid rate of muscle fiber formation. A subpopulation immunopositive for Myf5 only was identified in both ages of BSC isolates. The growth kinetics and heterogeneity of young BSC was further evaluated by clonal analysis. Single cell clones were established and analyzed after 10 d. Colonies segregated into 2 groups based upon population doubling time. Immunostaining of the slow-growing colonies (population doubling time ≥ 3 d) revealed that a portion exhibited asymmetric distribution of the lineage markers Pax7 and Myf5, similar to self-renewable mouse muscle stem cells. In summary, these results offer insight into the heterogeneity of BSC and provide evidence for subtle differences between rodent and bovine myogenic precursors.
卫星细胞是一种具有异质性的肌源性前体细胞群，负责哺乳动物的肌肉生长和修复。本实验的目的是研究从年轻和成年动物分离的牛卫星细胞（BSC）的生长速度和异质性程度。BSC 从年轻（4.3 ± 0.5 d）和成年（估计 24 至 27 月龄）牛的半膜肌中收获，并进行大规模培养。年轻动物的 BSC 更早地重新进入细胞周期，更早地达到最大 5-乙炔基-2'-脱氧尿苷（EdU）掺入（P < 0.05）。与从年轻动物分离的 BSC 相比，培养 3、4 和 5 d 后，成年 BSC 中的 MyoD 和 myogenin 免疫阳性核较少（P < 0.05）。这些结果表明，来自年轻动物的 BSC 比来自成年动物的 BSC 更早地激活、增殖和分化。使用专门针对 Pax7 和 Myf5 的抗体（卫星细胞和成肌细胞的谱系标记物）检查 BSC 中的谱系异质性。免疫细胞化学显示，大多数表达 Pax7 的 BSC 也表达 Myf5；一小部分（~5%）未能表现出 Myf5 免疫反应性。与成年 BSC 相比，来自年轻动物的 Pax7:Myf5 BSC 的百分比在培养过程中更快（P < 0.05）下降，表明更快的肌纤维形成速度。在两种年龄的 BSC 分离物中都鉴定出仅对 Myf5 免疫阳性的亚群。通过克隆分析进一步评估了年轻 BSC 的生长动力学和异质性。单细胞克隆建立并在 10 d 后进行分析。根据群体倍增时间，克隆集落分为 2 组。对生长缓慢的克隆（群体倍增时间≥3 d）进行免疫染色显示，一部分表现出谱系标记物 Pax7 和 Myf5 的不对称分布，类似于具有自我更新能力的小鼠肌肉干细胞。总之，这些结果深入了解了 BSC 的异质性，并为啮齿动物和牛肌源性前体之间的细微差异提供了证据。
