硒甲基硒代半胱氨酸对神经母细胞瘤细胞增殖及凋亡的影响

Effects of Se-methyl Selenocysteine on Proliferation and Apoptosis of Neuroblastoma

目的观察硒甲基硒代半胱氨酸(MSC)对神经母细胞瘤(NB)细胞增殖及凋亡的影响,并初探其作用机制。方法不同浓度MSC处理NB细胞株SH-SY5Y和SK-N-DZ。用CCK-8法检测各组细胞增殖情况;用流式细胞仪检测各组细胞周期和凋亡情况;采用蛋白免疫印迹(Western Blot)法检测各组凋亡相关蛋白。结果MSC作用于NB细胞(SH-SY5Y细胞和SK-N-DZ细胞)24 h后,细胞存活率明显降低;随MSC浓度升高,SH-SY5Y细胞和SK-N-DZ细胞存活率均明显下降,SH-SY5Y细胞和SK-N-DZ细胞的半抑菌浓度(IC50)分别为(47.89±1.74)μmol/L和(49.42±0.38)μmol/L,后期仅选用浓度分别为5μmol/L、25μmol/L和50μmol/L MSC处理NB细胞。与空白对照组比较,25μmol/L和50μmol/L MSC干预后SK-N-DZ细胞早期凋亡率明显增加,差异均有统计学意义(P<0.01或P<0.001);不同浓度MSC处理后,SH-SY5Y和SK-N-DZ细胞周期G_(0)/G_(1)、S、G_(2)/M比较,差异均无统计学意义(P>0.05);与空白对照组比较,5μmol/L、25μmol/L和50μmol/L MSC干预后SK-N-DZ细胞caspase-3和SH-SY5Y细胞caspase-9蛋白表达水平明显升高,差异有统计学意义(P<0.01或P<0.001)但caspase-8蛋白表达水平比较差异无统计学意义(P>0.05)。结论MSC主要通过激活内源性凋亡通路诱导细胞凋亡抑制NB细胞生长,与细胞周期无相关性。

Objective To analyze the effect of Se-methyl selenocysteine(MSC)on the proliferation and apoptosis of neuroblastoma cells.Methods The neuroblastoma cell lines SH-SY5Y and SK-N-DZ were treated with different concentrations of MSC.Cell proliferation was detected by CCK-8.Cell cycle and apoptosis was detected by flow cytometry,and apoptosis related proteins was detected by Western Blot.Results The survival rate of NB cells(SH-SY5Y cells and SK-N-DZ cells)was significantly inhibited by MSC for 24 hours.With the increase of MSC concentration,the survival rate of SH-SY5Y cells and SK-N-DZ cells significantly decreased.After analysis,the semi inhibitory concentration(IC50)of SH-SY5Y cells and SK-N-DZ cells was(47.89±1.74)μmol/L and(49.42±0.38)μmol/L.respectively.In the later stage of this study,only 5μmol/L,25μmol/L,and 50μmol/L MSC were used to treat NB cells.Compared with blank control group,the early apoptosis rate of the cells increased significantly after treatment with 25μmol/L and 50μmol/L MSC(P<0.01 or P<0.001).After treatment with different concentrations of MSC,there was no significant difference in G_(0)/G_(1),S,G_(2)/M ratio between SH-SY5Y and SK-N-DZ(P>0.05).Compared with blank control group,the protein expression levels of caspase-3 of SK-N-DZ cells and caspase-9 of SH-SY5Y cells were significantly higher after treatment with 5μmol/L,25μmol/L,and 50μmol/L MSC(P<0.01 or P<0.001),which was dose-dependent,but there was no significant difference in caspase-8 protein expression level(P>0.05).Conclusion MSC can induce apoptosis and inhibit the growth of neuroblastoma cells mainly through endogenous apoptosis pathway,there is no significant correlation with cell cycle.