苦参碱抑制神经母细胞瘤LA-N-5细胞增殖及MYCN基因mRNA的表达

Matrine inhibits the proliferation of neuroblastoma LAN5 cell and MYCN mRNA expression

摘要:

目的：神经母细胞瘤是4岁以下儿童最常见的恶性实体肿瘤,MYCN基因的高表达导致预后更加恶劣。苦参碱作为中药苦参的重要成分对多种肿瘤有治疗作用，该实验拟应用苦参碱作用神经母细胞瘤细胞(LA-N-5)，对肿瘤细胞增殖及MYCN基因mRNA表达受抑制情况进行初步研究，希望可以为神经母细胞瘤的治疗开拓新的思路。方法：以终浓度为0.25 mg/mL，0.50 mg/mL，0.75 mg/mL，1.00 mg/mL苦参碱作用神经母细胞瘤LA-N-5细胞, MTT法检测LA-N-5细胞增殖情况变化，采用SYBR 绿色荧光染料Ⅰ的实时荧光定量RT-PCR检测MYCN基因mRNA表达受抑制情况。结果：苦参碱对LA-N-5细胞增殖的抑制呈明显的剂量、时间依赖性，随剂量的增加，作用时间的延长，抑制效果逐渐加强。1.00 mg/mL作用72 h后肿瘤细胞增殖抑制效率达36.3%，单细胞MYCN基因mRNA表达9.7±0.35拷贝，抑制效率达44.6%。结论：应用苦参碱在体外环境作用LA-N-5细胞，可以有效抑制该细胞的增殖、原癌基因MYCN mRNA的表达，为临床上神经母细胞瘤的治疗开拓了新的思路。

Abstract:

OBJECTIVE: Neuroblastoma is the most common malignant solid tumor in children under 4 years. Amplification of MYCN oncogene is associated with advanced-stage disease, rapid tumor progression, resistance to treatment, and poor outcome. Matirne has the anti-tumor activity. This study was designed to investigate the effects of matrine on LA-N-5 cell line proliferation and MYCN gene mRNA expression. METHODS: Neuroblastoma LA-N-5 cells were treated by 0.25,0.50,0.75 or 1.00 mg/mL matrine. MTT was used to measure the levels of the proliferation of LA-N-5 cells cultured with different concentrations of matrine. MYCN gene mRNA expression in LA-N-5 cells was measured using real time RT-PCR with SYBR GREEN I fluorescence. RESULTS: The proliferation of LA-N-5 cells was obviously inhibited by matrine in a dose-and time-dependent manner. Matrine of 1.00 mg/mL treatment for 72 hrs produced a best effect, with an inhibitory rate of LA-N-5 cell proliferation of 36.3% and an inhibitory rate of MYCN gene mRNA expression of 44.6%. CONCLUSIONS: Matrine may inhibit the growth of neuroblastoma cells and down-regulate MYCN mRNA expression. It may be promising as a new drug for treatment of neuroblastoma.