Khwanraj Kawinthra, Phruksaniyom Chareerut, Madlah Suriyat, Dharmasaroja Permphan
Department of Anatomy, Faculty of Science, Mahidol University, Bangkok 10400, Thailand.
Department of Anatomy, Faculty of Science, Mahidol University, Bangkok 10400, Thailand ; Department of Pharmacology, Faculty of Dentistry, Mahidol University, Bangkok 10400, Thailand.
Neurol Res Int. 2015;2015:734703. doi: 10.1155/2015/734703. Epub 2015 Nov 8.
The human neuroblastoma SH-SY5Y cell line has been used as a dopaminergic cell model for Parkinson's disease research. Whether undifferentiated or differentiated SH-SY5Y cells are more suitable remains controversial. This study aims to evaluate the expression of apoptosis-related mRNAs activated by MPP(+) and evaluate the differential expression of tyrosine hydroxylase (TH) in undifferentiated and retinoic acid- (RA-) induced differentiated cells. The western blot results showed a gradual decrease in TH in undifferentiated cells and a gradual increase in TH in differentiated cells from days 4 to 10 after cell plating. Immunostaining revealed a gradual increase in TH along with neuritic outgrowth in differentiated cells on days 4 and 7 of RA treatment. For the study on cell susceptibility to MPP(+) and the expression of apoptosis-related genes, MTT assay showed a decrease in cell viability to approximately 50% requiring 500 and 1000 μM of MPP(+) for undifferentiated and RA-differentiated cells, respectively. Using real-time RT-PCR, treatment with 500 μM MPP(+) led to significant increases in the Bax/Bcl-2 ratio, p53, and caspase-3 in undifferentiated cells but was without significance in differentiated cells. In conclusion, differentiated cells may be more suitable, and the shorter duration of RA differentiation may make the SH-SY5Y cell model more accessible.
人神经母细胞瘤SH-SY5Y细胞系已被用作帕金森病研究的多巴胺能细胞模型。未分化或分化的SH-SY5Y细胞哪种更合适仍存在争议。本研究旨在评估由MPP(+)激活的凋亡相关mRNA的表达，并评估酪氨酸羟化酶(TH)在未分化细胞和视黄酸(RA)诱导分化细胞中的差异表达。蛋白质印迹结果显示，接种细胞后第4至10天，未分化细胞中TH逐渐减少，分化细胞中TH逐渐增加。免疫染色显示，在RA处理的第4天和第7天，分化细胞中TH随着神经突生长而逐渐增加。对于细胞对MPP(+)的敏感性和凋亡相关基因表达的研究，MTT试验显示，未分化细胞和RA分化细胞分别需要500和1000μM的MPP(+)才能使细胞活力降低至约50%。使用实时RT-PCR，500μM MPP(+)处理导致未分化细胞中Bax/Bcl-2比值、p53和caspase-3显著增加，但在分化细胞中无显著变化。总之，分化细胞可能更合适，而较短时间的RA分化可能使SH-SY5Y细胞模型更易于使用。
