Abstract：Objective The aims of this study were to establish a reliable method for primary culture, purification and identification of skeletal muscle satellite cells from fetal rat in vitro and to provide experiment means for investigating fetal and preterm skeletal muscle developmental programming. Methods Fetal rats were obtained by caesarean section on day 18 of gestation. The limb skeletal muscles were isolated from fetal rats under the surgical microscope, disassociated with type I collagenase and trypsin by two-steps digestion method, and purified via differential adhesion. Growth medium contained 70% DMEM/F12ham, 20% FBS, 10% HS and 5ng/ml bFGF. Differentiation medium contained 98% DMEM/F12ham and 2% HS. CCK-8 assay was used to record fetal rat skeletal muscle satellite cells growth curves. The expression of desmin was identified by immunocytochemistry staining. The differentiation of satellite cells was observed by inverted microscopy.Results Fetal rat skeletal muscle satellite cells entered the logarithmic growth period after 3 d of culture in vitro, and the platform phase was on day 5 to day 6. The positive rate of desmin was beyond 90%. Cells could fuse with each other and form myotubes with rhythmic contraction phenomenon when cultured in the differentiation medium.Conclusion High purity and viability primary fetal rat skeletal muscle satellite cells with potential of myogenic differentiation could be isolated and purified by digestion and preplate technique.
余慕雪, 戴杰民, 郭楚怡, 卢珍通, 杨佩军, 庄思齐. 胎鼠骨骼肌卫星细胞的原代培养、纯化与鉴定[J]. 中国生育健康杂志, 2016, 27(4): 335-339.
YU Muxue, DAI Jiemin, GUO Chuyi, LU Zhentong, YANG Peijun, ZHUANG Siqi. Primary culture, purification and identification of skeletal muscle satellite cells from fetal rat. Chinese Journal of Reproductive Health, 2016, 27(4): 335-339.
