MOE Key Laboratory of Molecular Genetics and Breeding, College of Marine Life Sciences, Ocean University of China, Qingdao 266003, China.
MOE Key Laboratory of Molecular Genetics and Breeding, College of Marine Life Sciences, Ocean University of China, Qingdao 266003, China; Laboratory of Tropical Marine Germplasm Resources and Breeding Engineering, Sanya Oceanographic Institution, Ocean University of China, Sanya, China.
Gene. 2021 Nov 15;802:145869. doi: 10.1016/j.gene.2021.145869. Epub 2021 Aug 2.
Skeletal myoblasts are activated satellite cells capable of proliferation and differentiation. Studies on mammalian myoblast differentiation and myogenesis could be carried out in vitro thanks to the availability of mouse myoblast cell line C2C12. Lacking of muscle cell line hinders the studies of teleost fish myogenesis. Here, we established a continuous skeletal muscle cell line from juvenile rockfish (Sebastes schlegelii) muscle using explant method and subcultured more than 50 passages for over 150 days. Stable expression of myoblast-specific marker, MyoD (myoblast determination protein) and the potential of differentiation into multi-nucleated skeletal myotubes upon induction suggested the cell line were predominately composed of myoblasts. Transcriptome analysis revealed a total of 4375 genes differentially expressed at four time points after the switch to differentiation medium, which were mainly involved in proliferation and differentiation of myoblasts. KIF22 (kinesin family member 22) and POLA1 (DNA polymerase alpha 1) were identified as the key genes involved in fish myoblast proliferation whereas MYL3 (myosin light chain 3) and TNNT2 (troponin T2) were determined as the crucial genes responsible for differentiation. In all, the continuous myoblasts cultured in this study provided a cell platform for future studies on marine fish myoblast differentiation and myogenesis. The molecular process of myoblast differentiation revealed in this study will open a window into the understanding of indeterminate muscle growth of large teleost.
成肌细胞是能够增殖和分化的激活卫星细胞。由于鼠成肌细胞系 C2C12 的可用性，哺乳动物成肌细胞分化和肌发生的研究可以在体外进行。缺乏肌肉细胞系阻碍了硬骨鱼肌发生的研究。在这里，我们使用外植体方法从小鲈（Sebastes schlegelii）肌肉中建立了连续的骨骼肌细胞系，并进行了超过 50 代的传代培养，超过 150 天。成肌细胞特异性标志物 MyoD（成肌决定蛋白）的稳定表达以及在诱导后分化为多核骨骼肌肌管的潜力表明，该细胞系主要由成肌细胞组成。转录组分析显示，在切换到分化培养基后的四个时间点，共有 4375 个基因差异表达，这些基因主要参与成肌细胞的增殖和分化。KIF22（驱动蛋白家族成员 22）和 POLA1（DNA 聚合酶 alpha 1）被鉴定为参与鱼类成肌细胞增殖的关键基因，而 MYL3（肌球蛋白轻链 3）和 TNNT2（肌钙蛋白 T2）被确定为负责分化的关键基因。总之，本研究中培养的连续成肌细胞为未来海洋鱼类成肌细胞分化和肌发生的研究提供了细胞平台。本研究中揭示的成肌细胞分化的分子过程将为理解大型硬骨鱼不定型肌肉生长打开一扇窗。
