小鼠脂肪干细胞分离培养方法

摘要

目的：建立一种简单高效的小鼠脂肪干细胞分离培养方法，为小鼠模型中间充质干细胞研究提供充足的细胞来源。方法体外分离小鼠腹股沟皮下脂肪组织，应用0．1％胶原酶NB4消化，含10％ FBS的低糖DMEM培养基贴壁培养小鼠脂肪干细胞，并对其细胞形态、体外增殖能力、多向分化潜能及表面标记进行分析研究。结果体外分离培养的小鼠脂肪干细胞具有良好的细胞形态、极强的增殖能力，体外经三向诱导，具有成脂、骨及软骨分化潜能，表面标记CD34、CD105阳性，Sca-1高表达，不表达CD45及 SSEA-1。结论利用该实验方法，能培养出高纯度、具有良好干性及极强增殖能力的小鼠脂肪干细胞。%Objective To establish a simple and effective method for isolation and culture of mouse adipose-derived stem cells (mASCs)in vitro,in order to provide the sufficient sources of seed cells for the research of mesenchymal stem cells.Methods The mouse inguinal fat tissues were isolated in vitro and performed a digestion with 0.1% collagenase type NB4,then adipose-derived stem cells(ASCs)were seeded and adhered to the culture dishes in low glucose DMEM containing 10% fetal calf serum.The cellu-lar morphology,in vitro proliferation capacity,multidifferentiation potential and immunophenotype were assessed.Results The mASCs showed good cell morphology,extremely strong proliferation capacity and potential of adipogenesis,osteogenesis and chon-drogenesis via in vitro three-dimensional induction.The cellular surface antigen phenotype was consistent with that reported by lit-erature,and the expression of CD34 and CD105 was positive,Sca-1 was highly expressed,CD45 and SSEA-1 were not expressed. Conclusion Using the experimental methods in this research can culture the high purity of mASCs with the excellent stem cell properties and extremely strong proliferative ability.