他米巴罗汀对神经母细胞瘤SH-SY5Y细胞增殖及酪氨酸激酶受体A、N-myc表达的影响

Effect of Tamibarotene on the proliferation and Tyrosine kinase receptor A, N-myc expressions of neuroblastoma SH-SY5Y cells

目的研究他米巴罗汀(Tamibarotene,Am80)对神经母细胞瘤SH-SY5Y细胞增殖抑制能力及酪氨酸激酶受体A(TrkA)、N-myc (即MYCN) mRNA和蛋白表达的影响,为治疗神经母细胞瘤提供实验依据。方法采用不同浓度(0、10、20、40、80、160 μmol/L)Am80作用SH-SY5Y细胞48 h。采用细胞计数试剂盒(Cell Counting Kit-8,CCK-8)检测细胞增殖情况。采用反转录-聚合酶链反应(RT-PCR)和Western-blot法检测48 h时SH-SY5Y细胞中TrkA、MYCN mRNA和蛋白的表达情况。结果当Am80浓度为10 μmol/L时,Am80对SH-SY5Y细胞无显著抑制作用[(3.51±1.68)%,抑制率<5%];当Am80浓度为20 μmol/L时,显示弱抑制作用[(9.60±1.97)%,抑制率<10%];Am80浓度为80 μmol/L时,对SH-SY5Y细胞增殖的抑制率显著增强[(57.43±4.95)%];TrkA随着Am80浓度增加,表达量增加;Am80能显著降低SH-SY5Y细胞MYCN的表达(10 μmol/L:0.65±0.05比20 μmol/L:0.36±0.06),差异有统计学意义(P<0.05)。结论Am80可抑制SH-SY5Y细胞的增殖,随着Am80浓度的增加,对细胞的抑制作用逐渐增强,Am80浓度越高对肿瘤细胞的抑制作用越显著。其机制可能与通过下调MYCN来提升TrkA的表达有关。

Objective To study the effect of Tamibarotene on the SH-SY5Y cell proliferation inhibition ability and the mRNA and protein expressions of tyrosine kinase receptor a (TrkA) and N-myc (MYCN) in order to provide some experimental bases for the treatment of neuroblastoma. Methods The SH-SY5Y cells were treated with different concentrations of Am80 (0, 10, 20, 40, 80, 160 μmol/L) for 48 h, then Cell Counting Kit-8(CCK-8)was used to test the cell proliferation.Reverse transcription PCR(RT-PCR)and Western blot were used to test the mRNA and protein expressions of TrkA and MYCN at 48 hours. Results When the concentration was 10 μmol/L, Am80 had no significant inhibitory effect on SH-SY5Y cells[(3.51±1.68)%, inhibition ratio<5%];but when the concentration was 20 μmol/L, it showed weak inhibition[(9.60±1.97)%, inhibition ratio<10%]. The inhibition rate of SH-SY5Y cell proliferation[(57.43±4.95)%] was significantly enhanced at Am80 with a concentration of 80 μmol/L.The concentrations of Am80 could effectively inhibit SH-SY5Y cell proliferation in a dose-dependent manner(P<0.05). The expression of TrkA increased with the increase of Am80 concentration.Am80 significantly decreased the expression of MYCN in SH-SY5Y cells(10 μmol/L: 0.65±0.05 vs.20 μmol/L: 0.36±0.06), and the difference was statistically significant(P<0.05). Conclusions It is suggested that Am80 can effectively inhibit SH-SY5Y cell proliferation in a concentration-dependent manner.The underlying mechanism involves increasing the expression of TrkA by down-regulation of MYCN.