Hongyi Yuan1, Hairong Du1, Zi’ao Yuan1, Xiaoliang Zhi1, Minghai Zhang1*, Weijun Guan2
1College of Wildlife and Protected Area, Northeast Forestry University, Harbin, 150040, China - 2Institute of Animal Sciences of Chinese Academy of Agricultural Sciences, Beijing, 100193, China
Objective: To establish a method for isolation, culture and identification of skeletal muscle satellite cells (MuSCs) in vitro, that play the main role in skeletal muscle generation after birth, and to investigate its biological characteristics.
Methods: In this study, chest muscles of 18-day-old Peking duck embryos were selected as the re-search material, the muscle were cut into minced meat under sterile conditions, by enzyme digestion using 0.1% collagenase I and 0.25% trypsin. After digestion,filtration and resuspended, the isolated cells were cultured in 37℃ with 5% CO2. Skeletal MuSCs were confirmed by immunofluorescence and the RT-PCR of specific markers Pax7, Myf5 and MyoD,
Result: High purity satellite cells were obtained by two-step enzyme digestion,the growth kinetics of various MuSC passages were assessed and growth curves were typically sigmoidal. Pax7,Myf5 and MyoD were positive by immunofluorescence and RT-PCR. Differentiation into adipocytes was assessed by oil red O staining, and differentiation into osteoblasts was evaluted by alizarin red staining. Moreover, long multicore muscle fibers and beating cardiomyogenic cells were detected after induction of myogenic differentiation.
Conclusion: This study confirmed the multi potent differentiation potential of skeletal muscle satellite cells, which can provide basic data for stem cell research.
Peking duck, Biological characteristic, Directed differentiation in vitro, Isolation, Skeletal muscle satellite cell.