In vitro differentiation of human umbilical cord blood-derived mesenchymal stem cells into dopaminergic neurons

doi:10.3969/j.issn.2095-4344.2012.49.012

Abstract

Abstract:

BACKGROUND: Parkinson's disease treated by cell replacement therapy has become a hot research at present, and how to obtain adequate and effective dopaminergic neurons in vitro is a strategy in the treatment of the disease.
OBJECTIVE: To investigate the feasibility of human umbilical cord blood-derived mesenchymal stem cells differentiating into dopaminergic neurons in vitro.
METHODS: The fifth passage human umbilical cord blood-derived mesenchymal stem cells were inoculated with 5×103/mL. The cells were pre-inducted with 20 ng/mL epidermal growth factor+20 ng/mL basic fibroblast growth factor for 24 hours. Then the cells were divided into four groups: the control group was not given induction medium but only cultured in DMEM/F12 culture medium containing 10% fetal bovine serum; the other three induced groups were induced with 100 μmol/L ascorbic acid, 1 μmol/L all-trans retinoic acid 50 μg/mL glial cell line-derived neurotrophic factor alone or in combination respectively.
RESULTS AND CONCLUSION: The expression of tyrosine hydroxylase, dopamine transporter and dopamine receptor D2 mRNA could be seen in the induced groups. After induction and differentiation, the cells could express the neurons and glial cell-specific antigens. Compared with the control group, the positive rates of nestin, neuron-specific enolase, glial fibrillary acidic protein, tyrosine hydroxylase, dopamine transporter and dopamine receptor D2 in the induced groups were significantly increased (P < 0.05); the positive rates in the all-trans retinoic acid+glial cell line-derived neurotrophic factor group and the combination induced group were significantly higher than those in the ascorbic acid group (P < 0.05), but the increased degree of the combination induced group was significantly higher than that of the all-trans retinoic acid+glial cell line-derived neurotrophic factor group (P < 0.05). Ascorbic acid, all-trans retinoic acid+glial cell line-derived neurotrophic factor, all-trans retinoic acid+glial cell line-derived neurotrophic factor+ascorbic acid can promote the human umbilical cord blood-derived mesenchymal stem cells to differentiate into dopaminergic neurons, and the combined induction can get the best effect.

CLC Number:

R394.2

Wu Xiao-hua, Chen Nai-yao. In vitro differentiation of human umbilical cord blood-derived mesenchymal stem cells into dopaminergic neurons[J]. Chinese Journal of Tissue Engineering Research, 2012, 16(49): 9186-9191.

Figures/Tables 6

酪氨酸羟化酶阳性细胞率：抗坏血酸，全反式维甲酸 +胶质细胞源性神经营养因子，抗坏血酸+全反式维甲酸+胶质细胞源性神经营养因子各诱导组酪氨酸羟化酶阳性细胞率分别为(7.98±1.59)%，(13.50±1.45)%，(19.17±2.27)%。多巴胺转运体阳性细胞率：抗坏血酸，全反式维甲酸 +胶质细胞源性神经营养因子，抗坏血酸+全反式维甲酸+胶质细胞源性神经营养因子各诱导组多巴胺转运体阳性细胞率分别为(6.22±1.62)%，(12.07±1.42)%，(17.61±1.45)%。多巴胺受体D2阳性细胞率：抗坏血酸，全反式维甲酸+胶质细胞源性神经营养因子，抗坏血酸+全反式维甲酸+胶质细胞源性神经营养因子各诱导组多巴胺受体D2阳性细胞率分别为(0.22±0.08)%，(0.39±0.15)%，(0.48± 0.13)%。与对照组比较，各诱导组Nestin、神经元特异性烯醇化酶、胶质纤维酸性蛋白、酪氨酸羟化酶、多巴胺转运体、多巴胺受体D2阳性细胞率均明显升高(P < 0.05)。与抗坏血酸组比较，全反式维甲酸+胶质细胞源性神经营养因子组和抗坏血酸+全反式维甲酸+胶质细胞源性神经营养因子组均明显升高(P < 0.05)，且抗坏血酸+全反式维甲酸+胶质细胞源性神经营养因子组升高幅度高于全反式维甲酸+胶质细胞源性神经营养因子组(P < 0.05)。 2.4 Real Time RT-PCR检测结果与对照组相比，各诱导组中酪氨酸羟化酶mRNA、多巴胺转运体 mRNA、多巴胺受体D2 mRNA的含量均有所增加，其中以抗坏血酸+全反式维甲酸+胶质细胞源性神经营养因子组酪氨酸羟化酶mRNA、多巴胺转运体mRNA、多巴胺受体D2mRNA的含量增加最为明显，全反式维甲酸+胶质细胞源性神经营养因子组次之，抗坏血酸组单独诱导较弱，差异有显著性意义(P < 0.05)，见表1。"

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