Slow and fast myosin heavy chain content defines three types of myotubes in early muscle cell cultures

We prepared monoclonal antibodies specific for fast or slow classes of myosin heavy chain isoforms in the chicken and used them to probe myosin expression in cultures of myotubes derived from embryonic chicken myoblasts. Myosin heavy chain expression was assayed by gel electrophoresis and immunoblotting of extracted myosin and by immunostaining of cultures of myotubes. Myotubes that formed from embryonic day 5-6 pectoral myoblasts synthesized both a fast and a slow class of myosin heavy chain, which were electrophoretically and immunologically distinct, but only the fast class of myosin heavy chain was synthesized by myotubes that formed in cultures of embryonic day 8 or older myoblasts. Furthermore, three types of myotubes formed in cultures of embryonic day 5-6 myoblasts: one that contained only a fast myosin heavy chain, a second that contained only a slow myosin heavy chain, and a third that contained both a fast and a slow heavy chain. Myotubes that formed in cultures of embryonic day 8 or older myoblasts, however, were of a single type that synthesized only a fast class of myosin heavy chain. Regardless of whether myoblasts from embryonic day 6 pectoral muscle were cultured alone or mixed with an equal number of myoblasts from embryonic day 12 muscle, the number of myotubes that formed and contained a slow class of myosin was the same. These results demonstrate that the slow class of myosin heavy chain can be synthesized by myotubes formed in cell culture, and that three types of myotubes form in culture from pectoral muscle myoblasts that are isolated early in development, but only one type of myotube forms from older myoblasts; and they suggest that muscle fiber formation probably depends upon different populations of myoblasts that co-exist and remain distinct during myogenesis.

我们制备了针对鸡肌球蛋白重链异构体快、慢类型的单克隆抗体，并使用它们来检测源自鸡胚成肌细胞的肌管培养物中的肌球蛋白表达。通过对提取的肌球蛋白进行凝胶电泳和免疫印迹以及对肌管培养物进行免疫染色来测定肌球蛋白重链的表达。由胚胎第5 - 6天的胸肌成肌细胞形成的肌管合成了快、慢两种类型的肌球蛋白重链，它们在电泳和免疫学上是不同的，但由胚胎第8天或更老的成肌细胞培养物中形成的肌管仅合成快类型的肌球蛋白重链。此外，在胚胎第5 - 6天成肌细胞的培养物中形成了三种类型的肌管：一种仅含有快肌球蛋白重链，第二种仅含有慢肌球蛋白重链，第三种同时含有快和慢重链。然而，在胚胎第8天或更老的成肌细胞培养物中形成的肌管是单一类型的，仅合成快类型的肌球蛋白重链。无论胚胎第6天的胸肌成肌细胞是单独培养还是与等量的胚胎第12天的肌肉成肌细胞混合培养，形成且含有慢类型肌球蛋白的肌管数量是相同的。这些结果表明，慢类型的肌球蛋白重链可由细胞培养中形成的肌管合成，在发育早期分离的胸肌成肌细胞在培养中形成三种类型的肌管，但较老的成肌细胞仅形成一种类型的肌管；并且它们表明肌肉纤维的形成可能取决于在肌发生过程中共存且保持不同的不同成肌细胞群体。