bFGF促增殖后骨髓间充质干细胞向多巴胺能神经元样细胞的分化

Differentiation of Bone Marrow Mesenchymal Stem Cells into Dopaminergic Neuronlike Cells after Their Proliferation was Improved by bFGF

目的探讨碱性成纤维细胞生长因子(Basic fibroblast growth factor,bFGF)体外作用于骨髓间充质干细胞(Mesen-chymal stem cells,MSCs)后,诱导其向神经元样细胞和多巴胺能神经元样细胞定向分化的情况。方法从鼠骨髓中获得MSCs,培养传代,用MTT法检测bFGF对骨髓MSCs生长的影响。10 ng/ml bFGF作用2 d后,通过IBMX、细胞因子bFGF、GDNFI、L-1β、中脑神经胶质细胞条件培养基和中脑神经细胞膜碎片等分组联合诱导骨髓MSCs向神经元样细胞、多巴胺能神经元样细胞分化,免疫细胞化学方法鉴定特异标志NSE、MAP-2a,b和TH的表达。结果在一定范围内,bFGF对骨髓MSCs具有明显的促增殖作用。分化的神经元样细胞表达NSE、MAP-2a,b和TH,联合应用GDNFI、L-1β、中脑条件培养基和中脑神经细胞膜碎片诱导7 d后,NSE阳性率为(27.774±2.747)%,MAP-2a,b为(28.006±3.080)%,TH为(3.098±0.352)%。结论体外骨髓MSCs被诱导分化成神经元样细胞和多巴胺能神经元样细胞,为帕金森等中枢神经系统疾病的细胞移植治疗奠定了基础。

Objective To explore the differentiation of bone marrow measenchymal stem cells（ MSCs） intoneuron-likeanddopaminergic neuron-like ceils after their proliferation was improved by basic fibroblast growth factor（bFGF） in vitro,Methods The MSCs derived from rat bone marrow were cultured and pass, aged, and the effect of bFGF on their growth was detected by MTT method. After pretreatment with bFGF for 2 d, the MSCs were cultured at the medium containing 0.5 mmol/L IBMX for 2 d, then the medium was replaced with induction media conraining GDNF, IL-1β, mesencephalic glial-cell-conditioned medium and flash-frozen rnesencephalic membrane fragments. After incubation for 7 d, the surface markers of differentiated neuron, such as NSE, MAP-2a, b and TH were detected by immunocytochemical method. Results The bFGF at a certain concentration range improved the proliferation of MSCs significantly. After MSCs were incubated in induction medium for 7 d, the percentages of NSE, MAP-2a, b and TH-positive ceils were （27.774 ± 2. 747） %, （28.006 ± 3.080 ） % and （3.098± 0.352 ） % respectively. Conclusion The differentiations of MSCs into neuron-like and dopaminergic neuron-like cells were induced in vitro. The study laid a foundation of treatment of central nervous system disease,such as Parkinson＇ s disease,by neurotransplantation.