Hindi Lubna, McMillan Joseph D, Afroze Dil, Hindi Sajedah M, Kumar Ashok
Department of Anatomical Sciences and Neurobiology, University of Louisville School of Medicine, Louisville, USA.
Professor and Distinguished University Scholar, Department of Anatomical Sciences and Neurobiology, University of Louisville School of Medicine, Louisville, USA.
Bio Protoc. 2017 May 5;7(9). doi: 10.21769/BioProtoc.2248.
Myogenesis is a multi-step process that leads to the formation of skeletal muscle during embryonic development and repair of injured myofibers. In this process, myoblasts are the main effector cell type which fuse with each other or to injured myofibers leading to the formation of new myofibers or regeneration of skeletal muscle in adults. Many steps of myogenesis can be recapitulated through differentiation of myoblasts into myotubes. Most laboratories use immortalized myogenic cells lines that also differentiate into myotubes. Although these cell lines have been found quite useful to delineating the regulatory mechanisms of myogenesis, they often show a great degree of variability depending on the origin of the cells and culture conditions. Primary myoblasts have been suggested as the most physiologically relevant model for studying myogenesis . However, due to their low abundance in adult skeletal muscle, isolation of primary myoblasts is technically challenging. In this article, we describe an improved protocol for the isolation of primary myoblasts from adult skeletal muscle of mice. We also describe methods for their culturing and differentiation into myotubes.
成肌作用是一个多步骤过程，在胚胎发育和受损肌纤维修复过程中导致骨骼肌形成。在此过程中，成肌细胞是主要的效应细胞类型，它们相互融合或与受损肌纤维融合，从而在成体中形成新的肌纤维或实现骨骼肌再生。成肌作用的许多步骤可通过成肌细胞分化为肌管来重现。大多数实验室使用也能分化为肌管的永生化成肌细胞系。尽管已发现这些细胞系对于阐明成肌作用的调控机制非常有用，但它们往往因细胞来源和培养条件的不同而表现出很大程度的变异性。原代成肌细胞被认为是研究成肌作用最具生理相关性的模型。然而，由于它们在成年骨骼肌中的丰度较低，原代成肌细胞的分离在技术上具有挑战性。在本文中，我们描述了一种从成年小鼠骨骼肌中分离原代成肌细胞的改进方案。我们还描述了将它们培养并分化为肌管的方法。
