Bian Yanchao, He Xiaohong, Mu Ren, Li Xiangchen, Ma Yuehui, Li Yunzhang, Guan Weijun
College of Animal Veterinary, Inner Mongolia Agricultural University, Hohhot 010018, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2013 Nov;29(11):1196-9.
To isolate and culture skeletal muscle satellite cells of Luxi cattle embryo and study its biological characteristics.
The skeletal muscles were taken from the limbs of Luxi cattle embryo of 30-50 d old to isolate and culture the skeletal muscle satellite cells by differential adherence method and co-digestion of type I collagenase and trypsin. Satellite cell surface markers, desmin, MyoD, c-Met, Myf5 and pax7 were detected by immunocytochemistry and RT-PCR, and its biological characteristics were researched.
The satellite cells were cultured to over passage 15. Cell viability was (97.90 ± 0.96)% after cryopreservation. The immunocytochemical staining showed that the specific surface antigen markers desmin and MyoD were positive, and RT-PCR also indicated the positive expressions of desmin, Myf5, c-Met and pax7. Colony formation was (56.39 ± 1.41)%. The karyotype analysis demonstrated that the isolated satellite cells were all derived from normal cattle embryos. Bovine satellite cells were successfully induced into osteoblasts and neuron-like cells by various inductors, and were positively expressed after alizarin red and toluidine blue staining. The osteoblast specific genes osteopontin and type I collagen and the nerve cell specific genes MAP-2 and nestin were positive by RT-PCR.
The skeletal muscle satellite cells of the Luxi cattle embryo are successfully isolated and cultured in vitro , They have the ability of being differentiated into osteoblasts and neuron-like cells.
分离培养鲁西黄牛胚胎骨骼肌卫星细胞并研究其生物学特性。
取30 - 50日龄鲁西黄牛胚胎四肢骨骼肌，采用差速贴壁法及Ⅰ型胶原酶和胰蛋白酶联合消化法分离培养骨骼肌卫星细胞。通过免疫细胞化学和RT-PCR检测卫星细胞表面标志物结蛋白、肌分化抗原（MyoD）、肝细胞生长因子受体（c-Met）、肌原性决定因子5（Myf5）和配对盒基因7（pax7），并研究其生物学特性。
卫星细胞培养至15代以上。冻存后细胞活力为（97.90±0.96）%。免疫细胞化学染色显示特异性表面抗原标志物结蛋白和MyoD呈阳性，RT-PCR也表明结蛋白、Myf5、c-Met和pax7呈阳性表达。集落形成率为（56.39±1.41）%。核型分析表明分离的卫星细胞均来源于正常牛胚胎。通过各种诱导剂成功将牛卫星细胞诱导分化为成骨细胞和神经元样细胞，茜素红和甲苯胺蓝染色后呈阳性表达。RT-PCR检测成骨细胞特异性基因骨桥蛋白和Ⅰ型胶原以及神经细胞特异性基因微管相关蛋白2（MAP-2）和巢蛋白呈阳性。
成功体外分离培养鲁西黄牛胚胎骨骼肌卫星细胞，其具有向成骨细胞和神经元样细胞分化的能力。
