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目的研究酪氨酸激酶受体B(tyrosine kinase receptor,TrkB)及其配体脑源性神经营养因子(brain-derived neurotrophic factor,BDNF)的表达水平对神经母细胞瘤(neuroblastoma,NB)细胞化疗敏感性的影响。方法Western-blot技术检测不同纳摩尔浓度全反式维甲酸(all trans-retinoic acid, ATRA)诱导后TrkB蛋白水平变化;四甲基偶氮蓝比色(MTT)法检测细胞存活率;流式细胞仪(flow cytometer,FCM)检测细胞凋亡率;透射电镜检测凋亡细胞的形态。结果(1)不同浓度ATRA(1、10、100 nmol／L)处理神经母细胞瘤细胞SY5Y 5 d,TrkB蛋白水平随ATRA浓度增加而增加;(2)BDNF 10 ng／ml+ATRA 10 nmol／L+顺铂(Cisplatin,CP)5μg／ml作用组细胞存活率、凋亡率同单用CP组比较均无显著差异,BDNF(50、100 ng/ml)加相同浓度ATRA及CP组细胞存活率均明显高于单用CP组,凋亡率均明显低于单用CP组,BDNF 100 ng／ml组存活率高于BDNF 50 ng／ml组,凋亡率低于BDNF 50 ng／ml组。ATRA 1 nmol／L+BDNF 50 ng／,ml+CP5μg/ml组细胞存活率、凋亡率同单用CP组比较无显著差异,ATRA 10、100 nmol／L加相同浓度BDNF及CP组细胞存活率均明显高于单用CP组,凋亡率均明显低于单用CP组,ATRA 100 nmol／L组细胞存活率高于ATRA 10 nmol／L组,凋亡率低于ATRA 10 nmol／L组。(3)透射电镜技术观察到CP作用组可见到较多细胞呈凋亡改变,而ATRA 10 nmol／L+BDNF 50 ng／ml+CP 5μg/ml组细胞形态多数正常。结论SY5Y对化疗药顺铂的敏感性受TrkB及BDNF水平的影响,二者水平越高越容易产生化疗耐药。
Objective To investigate the expression of tyrosine kinase receptor (TrkB) and its brain-derived neurotrophic factor (BDNF) in neuroblastoma (NB) The impact of sensitivity. Methods Western blotting was used to detect the level of TrkB protein after induced by all-trans retinoic acid (ATRA) in different nanomolar concentrations. Cell viability was measured by MTT assay. Flow cytometry The apoptosis rate was detected by flow cytometer (FCM). The morphology of apoptotic cells was detected by transmission electron microscopy. Results (1) After treatment with different concentrations of ATRA (1, 10, 100 nmol / L) for 5 days on SY5Y cells, TrkB protein level increased with increasing ATRA concentration. (2) The survival rate and apoptosis rate of L + cisplatin (5μg / ml) group were not significantly different from that of CP group alone. The survival rates of BDNF group (50,100 ng / ml) plus ATRA and CP group Were significantly higher than that of CP group. The apoptosis rate was significantly lower than that of CP group. The survival rate of BDNF 100 ng / ml group was higher than that of BDNF 50 ng / ml group, and the apoptosis rate was lower than that of BDNF 50 ng / ml group. ATRA 1 nmol / L + BDNF 50 ng /, ml + CP 5μg / ml group of cell survival rate, apoptosis rate compared with the single CP group no significant difference, ATRA 10,100 nmol / L plus the same concentration of BDNF and CP group cell survival The rates of apoptosis were significantly lower in CP group than those in CP group. The survival rate of ATRA 100 nmol / L group was higher than that of ATRA 10 nmol / L group, and the apoptosis rate was lower than that of ATRA 10 nmol / L group. (3) Transmission electron microscopy (TEM) showed that more cells were found to be apoptotic when treated with CP, while most of the cells in ATRA 10 nmol / L + BDNF 50 ng / ml + CP 5μg / ml group were normal. Conclusion The sensitivity of SY5Y to Cisplatin is affected by TrkB and BDNF levels. The higher the level of SY5Y, the more likely it is to develop chemoresistance.