一种新的成年大鼠成肌细胞体外批量扩增培养方法

A Novel Method for Adult Rat Myoblasts Isolation and Amplification in Batches

目的:建立快速高效的成年大鼠成肌细胞分离、体外批量扩增培养的实验方法。方法:通过混合酶一步消化法从少量成年骨骼肌样品中分离成肌细胞;采用差速贴壁筛选法纯化分离并批量扩增成肌细胞。结果:混合酶一步消化法需要的时间为2.1±0.52h,较之于传统方法所需要的6.8±0.67h,分离细胞所需时间显著缩短(P<0.01);采用差速贴壁筛选法纯化分离的成肌细胞myf5/desmin阳性率>97%,同传统分离扩增方法相比,体外批量扩增培养的这些成肌细胞具有典型的成肌细胞生长特性,在生长液中具有更好的增殖潜能,而在分化液中分化为肌管的特性无显著性差异(P>0.05)。结论:通过采用新建立的成年成肌细胞分离培养和批量扩增的方法,能够快速高效地获取高纯度的成肌细胞。

Objective： To establish a method for rapid isolation and amplification in batches of myoblasts from adult rats. Methods： Myoblasts were isolated from adult skeletal muscle by method of one step digestion with mixed enzyme we previously founded; then, they were purified and amplified in batches with the novel method which was named as purification with diverse adherence method. Results： In the new isolation method, the cell isolation time was 2.1±0.52 h, which was remarkably reduced compared with 6.8±0.67 h for the old isolation method（P〈0.05）. The myf5/desmin positive rate of isolated myoblasts got by means of purification with diverse adherence method was over 97%, and the characteristics of which had a preferable proliferation potency and were not different from that by the old method in regard to differentiate into myotubes in differentiation medium（P〉0.05）. Conclusion： The novel method could be used to rapidly isolate and amplify highly purified myoblasts.