ABSTRACT
Objective Bone marrow mesenchymal stem cells (BMSCs) were induced to differentiate to the special histological types of neurons in vitro.The morphological change of cells and positive expression of specific antigen on membrane were studied,and the function of connection between the induced BMSCs was also detected.The feasibility of BMSCs differentiate to the special histological types of neurons was investigated.Methods BMSCs were divided into group Ⅰ (induced with bFGF+GDNF),group Ⅱ (induced with bFGF+GDNF+WHI-P131 +Shh),and control group (no revulsive).The morphologic change of cells was observed,and the positive rate of neuron specific surface antigen and the content of dopamine were detected.Formation of mature synaptic structure was detected by immunohistochemical assay of postsynaptic density protein 95 (PSD-95) expression,and synaptic loop was shown by FM1-43 stain synaptic vesicles.Results By immunohistochemical staining,the positive rates of dopamine transporter (DAT) and tyrosine hydroxylase (TH) in group Ⅱ were significantly higher than those in group Ⅰ,and dopamine can been detected in cell culture supematant of group Ⅱ.After BMSCs was induced into dopamine neuron-like cells,number and length of cell protrusions,positive rate of PSD-95 and fluorescence intensity of FM1-43 in group Ⅱ were significantly higher than those of group Ⅰ.Conclusions There were no significant change in positive rate of neuron-specific surface markers,rate of cell survival and differentiation rate after BMSCs differentiated to dopaminergic neuron-like cells.The number and length of cell protrusions,content of dopamine in cell culture supematant,positive rate of dopaminergic neuron-specific surface antigen (DAT and TH),synaptic function index (positive rate of PSD-95 and fluorescence intensity of synaptic loop) of group Ⅱ were all significantly higher than that of group Ⅰ.
ABSTRACT
Objective To explore the differentiation of human bone marrow measenchymal stem cells (hMSCs) into neuron-like and dopaminergic neuron-like cells in vitro.Methods The hMSCs were isolated from adult human bone marrow and expanded on the flask undifferentiated state for 2 passages. After pretreatment with WHI-P131 for 48 h, the hMSCs were cultured at the medium containing 10 ng/ml basic fibroblast growth factor for 24 h, then incubated with all-trans-retinoic acid and glial-derived neurotrophic factor in serum-free media for 5 h. The surface markers of differentiated neuron were detected by immunocytochemical method and the transdifferentiation process was observed under light microscope.Results Under induction conditions, hMSCs progressively resumed typical neuronal morphological characteristics. After hMSCs were incubated in induction medium for 5 h, the percentage of NSE, nestin, GFAP, TH and DAT positive cells were (77.0?5.7)%, (54.2?3.7)%,(8.8?2.4)%, (36.5?15.8)% and (26.0?14.2)%, respectively. There were no positive expressions in the control group.Conclusion The hMSCs are able to differentiate into neuron-like and dopaminergic neuron-like cells in vitro.