冬凌草甲素对急性单核细胞白血病细胞株THP-1的作用研究  

Anti-leukemia Effect of Oridonin on Acute Monoblastic Leukemia

作　　者：郭勇[1] 单卿卿[1] 龚玉萍[1] 林娟[1] 杨曦[1] 

机构地区：[1]四川大学华西医院血液科,成都610041

出　　处：《临床误诊误治》2015年第4期108-112,共5页Clinical Misdiagnosis & Mistherapy

基　　金：国家自然科学基金(30770912);四川省科技厅社会公益项目(2008SZ0017);教育部回国留学人员启动金(20071108-18-4)

摘　　要：目的探讨冬凌草甲素对急性单核细胞白血病细胞株THP-1的抗白血病效应及其作用机制。方法以急性单核细胞白血病细胞株THP-1为研究对象,应用改良四甲基偶氮唑盐法测定4、6、8μmol/L浓度冬凌草甲素处理72 h后THP-1细胞的增殖抑制率,以及计算2、4、6、8、10μmol/L冬凌草甲素处理24、72、120 h后的细胞生存率并绘制生长曲线;显微镜下观察4、6、8μmol/L冬凌草甲素处理24 h后THP-1细胞的形态学变化;采用流式细胞术检测0、4、6、8μmol/L冬凌草甲素处理24 h后THP-1细胞凋亡率;采用Western blot法检测6μmol/L冬凌草甲素处理THP-1细胞0、12、24 h后Akt/m TOR、Raf/MEK/ERK细胞信号转导通路的变化,以及凋亡调节蛋白Bcl-2和Bax的变化。结果 14、6、8μmol/L冬凌草甲素处理72 h后细胞增殖抑制率分别为(20.02±11.15)%、(45.99±12.76)%、(86.39±9.18)%,与4μmol/L冬凌草甲素比较,6、8μmol/L冬凌草甲素处理后细胞增殖抑制率均升高,差异均有统计学意义(P<0.05或P<0.01);冬凌草甲素处理72 h后细胞半抑制浓度为(6.12±1.48)μmol/L;细胞生长曲线显示,冬凌草甲素对THP-1细胞的生长抑制作用呈时间和浓度依赖性。2冬凌草甲素处理24 h后,THP-1细胞出现凋亡,形成凋亡小体,浓度越高细胞凋亡小体形成越多。30、4、6、8μmol/L冬凌草甲素(分别设为对照组及4、6、8μmol/L组)处理24 h后,THP-1细胞凋亡率分别为(3.7±1.1)%、(16.2±3.3)%、(30.1±4.3)%、(49.5±6.7)%,对照组与各浓度组凋亡率比较差异均有统计学意义(P<0.05或P<0.01);与4μmol/L组比较,6、8μmol/L组凋亡率均升高,差异均有统计学意义(P<0.05)。4经Western blot法检测6μmol/L冬凌草甲素处理THP-1细胞24 h后可抑制细胞内Akt/m TOR、Raf/MEK/ERK信号转导通路的活化,并下调Bcl-2、上调Bax的表达。结论冬凌草甲素通过抑制Akt/m TOR、Raf/MEK/ERK信号转导通路的活化,以及调节凋亡调节蛋白Bax和Bcl-2的表达而发挥抗Objective To investigate the anti-leukemia effect of oridonin on acute monoblastic leukemia cell line THP-1. Methods Acute monoblastic leukemia cell line THP-1 was cultured in vitro. Cell proliferation Inhibition rate after72 h of treatment of 4,6,8 μmol / L oridonin was examined using modified MTT assay,cell survival rate after 24,72,120 h of treatment of 2,4,6,8,10 μmol / L oridonin was calculated and the growth curve was drawn. The cellular morphologic changes after 24 h of treatment of 4,6,8 μmol / L oridonin were observed under a light microscope. The percentage of apoptosis of THP-1 cells after 24 h of treatment of 0,4,6,8 μmol / L oridonin was evaluated using flow cytometric analysis. The activation levels of Akt / m TOR,Raf / MEK / ERK signaling pathways and the expression levels of Bcl-2 and Bax after 0,12,24 h of treatment of 6 μmol / L oridonin were examined by Western blot. Results 1 Cell proliferation inhibition rate after 72 h of treatment of 4,6,8 μmol / L oridonin was（ 20. 02 ± 11. 15） %,（ 45. 99 ± 12. 76） %,（ 86. 39 ± 9. 18） % respectively,compared with 4 μmol / L oridonin treatment group,cell proliferation inhibition rate after 6,8 μmol / L oridonin treatment were higher,with statistically significant differences（ P〈0. 05 or P〈0. 01）. Cell growth curve showed that oridonin inhibited the growth of THP-1 cells in time- and dose- dependent manner and the IC50 of oridonin was（ 6. 12 ± 1. 48） μmol / L after 72 h of treatment. 2 After 24 h of treatment of oridonin,THP-1 cells apoptosis occurred and apoptotic bodies were formed,the higher the concentration of oridonin,the more apoptotic bodies were. 3 The percentage of apoptosis rate after 24 h of treatment of0,4,6,8 μmol / L oridonin were（ 3. 7 ± 1. 1） %,（ 16. 2 ± 3. 3） %,（ 30. 1 ± 4. 3） % and（ 49. 5 ± 6. 7） % respectively.The differences of apoptosis rate between various oridonin concentration groups and control group were statistically significant（ P〈0. 05 or P〈0. 01）. The differences

关 键 词：冬凌草素 白血病 单核细胞 急性 白血病抑制因子 凋亡调节蛋白质类 

分 类 号：R733.71[医药卫生—肿瘤]