摘要
目的:研究光辉霉素A(mithramy-cin A,MIT)对人肝癌细胞Huh-7的作用。方法:CCK-8法观察MIT对Huh-7细胞的生长抑制作用;Realtime-PCR法和蛋白质印迹法检测MIT作用后Huh-7细胞中Sp1、VEGF和c-Met表达情况。结果:MIT作用Huh-7细胞48h时细胞生长抑制作用与浓度成正相关。5×10-4μmol/L时细胞增殖的抑制率为2.9%,5μmol/L时达92.4%。随时间延长抑制率逐渐增高,作用24h时抑制率为6.15%,60h达72.65%。半数抑制浓度(IC50)为60.12nmol/L。与对照组Sp1mRNA表达量(1.4599±0.2696)相比,其表达量与浓度有关,500nmol/L时其表达量(0.2743±0.1043)下降约为81.21%,P=0.0027。且其表达量与时间有关,与表达量(1.459856±0.269603)相比,加药72h时其表达量(0.430778±0.125143)明显下降,P=0.0034。VEGF和c-Met的mRNA表达量变化有一个相同的趋势,随浓度和时间增加表达逐渐下调。MIT对Sp1、VEGF和c-Met在蛋白水平的表达有抑制作用。结论:MIT对Huh-7细胞有明显的生长抑制作用。MIT能抑制人肝癌Huh-7细胞Sp1及其下游基因表达。
OBJECTIVE：To study the effect of Mithramycin A（MIT） on cell proliferation and gene variation of Huh-7. METHODS：Cell growth was analyzed by CCK-8 assay. Real time-PCR and Western-blot methods were used to detect expression of Sp1,VEGF and c-Met after Huh-7 cells were treated with MIT. RESULTS：When treated with 5×10-4 μmol/L MIT,for 48 h the inhibition rate of cell proliferation was 2.9%. Treated with 5 μmol/L MIT,and it was 92.4%. It suggested that MIT inhibited the cell growth in a dose-dependent manner. Huh-7 cells cells were treated with 5×10-1 μmol/L MIT after different time. After 24 hours＇ treatment it was 6.15%. After 60 hours＇ treatment it was 72.65%. It suggested that MIT inhibited the cell growth in a time-dependent manner. The IC50 value of MIT for Huh-7 cells was 60.12 μmol/L. Sp1 mRNA expression showed a marked downregulation in 500 nmol/L group （0.274 3±0.104 3） compared with controlled group （1.459 9±0.269 6,P=0.002 7）. Meanwhile,It showed a marked downregulation in 72 h group （0.430 778±0.125 143） compared with 0 h group （1.459 856±0.269 603,P=0.003 4）. It suggested that Sp1 mRNA expression was assoiated with MIT in a dose-dependent and time-dependent manner.The trend of mRNA expression of VEGF and c-Met was in the same way.Treatment with MIT suppress the expression of Sp1 protein and its downstream target genes VEGF and c-Met. CONCLUSIONS：Mithramycin A could inhibit the cell growth. Treatment with MIT suppressed the expression of Sp1 and its downstream target genes in human liver cancer cell line Huh-7.
出处
《中华肿瘤防治杂志》
CAS
2009年第24期1917-1921,共5页
Chinese Journal of Cancer Prevention and Treatment
基金
上海市浦江人才计划项目(06PJ14096)
上海市科学技术委员会资助项目(054119628)
上海市浦东新区卫生系统重点学科项目(PWZXK2007-06)
