柴胡皂甙-d对HepG2细胞恶性表型的逆转作用

目的 探讨柴胡皂甙-d (SSd)对人肝癌细胞株HepG2细胞恶性表型的逆转作用及其机制. 方法 常规培养HepG2细胞至对数生长期,四甲基偶氮唑盐(MTT)比色法观察SSd不同浓度、作用不同时间对HepG2细胞增殖情况的影响.实验组选择10 mg/L的SSd作用HepG2细胞48h,Giemsa染色法观察细胞形态学改变;化学发光法测定细胞上清液中甲胎蛋白(AFP)浓度;放射免疫法检测细胞上清液中白蛋白浓度;transwell小室实验观察细胞迁移率;RT-PCR检测p27基因mRNA表达.并与空白对照组做比较.数据在多组间比较采用随机分组单因素方差分析,在两组间比较采用t检验. 结果 10 mg/L的SSd作用48 h,对HepG2细胞的生长抑制最明显(F=265.06,P＜0.01).实验组HepG2细胞形态倾向于正常分化,形态变小、变圆,核变小、变圆或卵圆形,核/质比例缩小.与对照组比较,实验组穿膜细胞数明显减少[(50.60+4.04)个与(41.00+4.64)个,t= -7.32,P＜0.01];细胞上清液中白蛋白含量明显增多[(24.7+2.8)×10-3g/L与(31.1 +4.9)×10-3g/L,t=7.83,P＜ 0.05];AFP含量明显减少[(118.2±15.6)×10μ g/L与(70.3+5.7)×103μg/L,t=-10.72,P＜0.0l].实验组p27基因mRNA的相对表达量明显多于对照组(t=22.00,P＜0.05). 结论 SSd对人肝癌细胞株HepG2细胞恶性表型具有明显的逆转作用,其机制可能与SSd上调HepG2细胞p27基因mRNA表达使其进入分化过程有关.

Effect of Saikosaponins-d on reversing malignant phenotype of HepG2 cells in vitro

Objective To observe the effect of SSd on reversing the malignant phenotype of HepG2cells and to investigate its mechanism in order to prove that SSd is a new choice to prevent and treat HCC.Methods HepG2 cells were cultured and treated by different concentrationgs (0 mg/L,2.5 mg/L,5.0 mg/L,10.0 mg/L and 20.0 mg/L) of SSd for 24 h,and treated by 10 mg/L of SSd for 0 h,6 h,12 h,24 h,48 h and 72h respectively.The cell inhibition rates were measured by MTT assay.Then cells were treated by 10 mg/L SSd for 48 hr in experimental group and treated by no SSd as a control,their morphological changes were observed by contrast phase microscope.The concentrations of ALB and AFP in clear supernatant liquid of cells were detected by radio-immunity and chemiluminescence.The cell migration rates were observed by transwell method,the relative expression levels of p27 mRNA were messured by RT-PCR.Results The inhibitive effect of 10 mg/L SSd was the most significant among different concentrations (F=265.06,P＜0.01).The shape of HepG2from experimental group turned into small and round,and their volume ratios of nucleus to plasma decreased.ALB in supernatant liquid of HepG2 was higher (t= 7.83,P ＜ 0.05,and its AFP was lower (t=-10.72,P ＜ 0.01 )as compared to control group.Cells migrated were fewer and p27 mRNA expression of HepG2 was higher in experimental group than that in control group (t=22.00,P＜0.05).Conclusion SSd could reverse the malignant phenotype of HepG2 cells.It was suggested that the up-regulation of p27 mRNA expression play an important role in the differentiation of HepG2 cells treated by SSd.