Blau H M, Webster C, Chiu C P, Guttman S, Chandler F
Exp Cell Res. 1983 Apr 1;144(2):495-503. doi: 10.1016/0014-4827(83)90431-7.
The interpretation of the majority of studies of Duchenne muscular dystrophy (DMD) has been complicated by the heterogeneous composition of the cultures used. In addition to muscle cells, muscle tissue contains adipocytes and fibroblasts and the proportion of these cell types varies, especially in disease states. To overcome this problem we developed culture conditions which permitted isolation and characterization of pure populations of clonally derived human muscle cells [1, 2]. Here we report the successful application of these methods to muscle cells from biopsies of individuals with diagnosed DMD. The normal and mutant human muscle cells were used in experiments of muscle differentiation in the same manner as cell lines. Frozen-stored cells were thawed, plated in a series of replicate plates, and allowed to differentiate under similar culture conditions. Yet, in contrast with cell lines, the cells were karyotypically normal, not altered by adaptation to long-term culture, and had a finite lifespan. We have systematically analysed specific properties of the normal and DMD muscle cells which differentiated in culture. The kinetics and extent of myoblast fusion, myotube morphology, and the accumulation and distribution of membrane acetylcholine receptors were monitored. In addition, the isozyme composition of creatine kinase and its intracellular and extracellular distribution were determined. Our results indicate that DMD muscle cells are fully capable of initiating myogenesis in culture and do not differ from normal muscle in several important parameters of differentiation.
大多数关于杜氏肌营养不良症（DMD）的研究解读因所用培养物的异质组成而变得复杂。除了肌肉细胞外，肌肉组织还含有脂肪细胞和成纤维细胞，并且这些细胞类型的比例各不相同，尤其是在疾病状态下。为了克服这个问题，我们开发了培养条件，允许分离和鉴定克隆衍生的人类肌肉细胞的纯群体[1,2]。在这里，我们报告了这些方法在诊断为DMD的个体活检肌肉细胞中的成功应用。正常和突变的人类肌肉细胞以与细胞系相同的方式用于肌肉分化实验。将冷冻保存的细胞解冻，接种到一系列重复的培养皿中，并在相似的培养条件下使其分化。然而，与细胞系不同的是，这些细胞的核型正常，不会因适应长期培养而改变，并且具有有限的寿命。我们系统地分析了在培养中分化的正常和DMD肌肉细胞的特定特性。监测了成肌细胞融合的动力学和程度、肌管形态以及膜乙酰胆碱受体的积累和分布。此外，还测定了肌酸激酶的同工酶组成及其细胞内和细胞外分布。我们的结果表明，DMD肌肉细胞在培养中完全能够启动肌生成，并且在几个重要的分化参数上与正常肌肉没有差异。
