GFP转基因鼠咀嚼肌卫星细胞的培养及影响因素

探索骨骼肌卫星细胞作为组织工程种子细胞的可能性。取新生绿色荧光蛋白(GFP)转基因鼠面部咀嚼肌,分离出肌卫星细胞,体外进行原代和传代培养,建立GFP转基因小鼠肌卫星细胞系。通过倒置显微镜观察肌卫星细胞的形态、生长曲线测定等,观察肌卫星细胞的增殖与分化能力,荧光显微镜观察GFP荧光的保持及表达,并观察不同浓度的FBS,IGF-1以及不同种植密度对肌卫星细胞生长的影响。结果表明:体外培养的咀嚼肌卫星细胞增殖速度快,15%的FBS,接种密度在5×104 cells/ml,IGF-1在100ng/ml有显著的促进增殖作用。荧光显微镜观察发现,肌卫星细胞经传代后仍然保持其特有的GFP荧光表达。

The Culture and Influence Factors on Proliferation of Skeletal Muscles Satellite Cells from GFP Transgenic Mouse in vitro

To investigate the possibility of skeletal muscles satellite cells as a seed of tissue engineering,The newborn transgenic mice were acquired to separate skeletal muscles satellite cells with enzyme digestion method.Cells were cultured and subcultured in vitro.Morphological observation,growth curve were investigated to evaluate the proliferation and differentiation characteristics of skeletal muscles satellite cells,fluorescence microscope was used to observe the GFP expression.We also observed the effect of different concentration of FBS,different density of implant cells and IGF-1 on the proliferation of skeletal muscle cells.The result showed that the cells expressed strong proliferative ability in vitro culture and they were positive with immunocytochemical stain of anti-sarcometric actin antibody,immunocytofluorescence suggested the good retain of GFP fluorescence in skeletal muscles satellite cells.15?S,5x104 cells/ ml implant density and 100ng/ml IGF-1 can obviously promote the prolification of cells.