三尖杉酯碱及氮烯苯酸诱导肿瘤细胞凋亡的时-效 ,量-效关系以及凋亡与坏死的消长变化

The relationships between dose effect and time effect of apoptosis induced by harringtonine and benzoazene in tumor cells and the change of growth and decline between apoptosis and necrosis

目的 :研究三尖杉酯碱及氮烯苯酸诱导肿瘤细胞凋亡的量 效 ,时 效关系以及凋亡与坏死的消长变化。方法 :以S1 80 腹水癌荷瘤小鼠为体内模型 ,HL60 细胞为体外模型 ,以Hoechst3 3 3 42染色细胞 ,荧光显微镜下观察并计算凋亡指数。透射电镜判定凋亡 ;流式细胞仪检测DNA分布加以验证。用重复测量方差分析对凋亡指数进行量化处理 ;以台盼蓝染色阳性作为坏死的参数 ,观察凋亡与坏死的关系。结果 :三尖杉酯碱和氮烯苯酸均诱导了肿瘤细胞凋亡 ,在荧光显微镜及透射电镜下见典型的凋亡改变。DNA直方图出现亚G1峰。对HL6 0细胞 ,在 2～ 2 4小时时间范围内 ,三尖杉酯碱在 0 .0 2～ 1.0 μg/ml ,氮烯苯酸在 0 .2～ 5 μg/ml浓度范围内 ,凋亡指数随药物浓度增加及时间延长而上升 ,2 4小时后 ,凋亡指数下降 ,台盼蓝染色阳性坏死细胞明显增加。在本实验的最高浓度组(三尖杉酯碱 5 .0 μg/ml,氮烯苯酸 2 5 μg/ml )凋亡指数最低 ,台盼蓝阳性率一直处于最高水平。在荷S1 80 腹水癌小鼠 ,氮烯苯酸在 2 5～ 2 5 0mg/kg和 2～ 48小时范围内 ,凋亡指数随剂量增加及时间延长而上升 ,48小时后 ,凋亡指数下降 ,台盼蓝染色阳性细胞明显增加。结论 :三尖杉酯碱及氮烯苯酸诱导肿瘤细胞凋亡 ,在一定剂量和时间范围内呈现明显的量 ?

Objective: To study the relationship on the dose effect and time effect of apoptosis induced by harringtonine (HT) and benzoazene (BAZ) in tumor cells and the change of growth and decline between apoptosis and necrosis. Methods:Ascitic cells of sarcoma 180 (S 180 ) of mice in vivo and human promyelocyte leukemia HL60 cells in vitro were used as models. The cells were stained with Hoechst 33342 and were observed under fluorescence microscope,the apoptosis index was counted. The ultra structure of the cells was examined with transmission electron microscope and DNA distribution was analyzed with FCM to define apoptosis. The statistical method of repeated measure analysis of variance was used to analyze the data. The trypan blue positive rate was used as a parameter of necrosis to investigate the possible relationship between apoptosis and necrosis. Results:Both of HT and BAZ could induce apoptosis of tumor cells. The typical apoptosis was confirmed by the examination of fluorescence microscope and transmission electron microscope. The sub G1 peak was found in DNA histogram by FCM. In HL60 cells ,during the 2～24 hrs,in the dose range of 0.02～1.0 μg/ml of HT,and of 0.2～5 μg/ml of BAZ,the apoptosis index was rising with the time passing and the concentration increasing. After 24 hrs,the apoptosis index reduced and the trypan blue positive rate rised. The highest concentration of drugs in this experiment (HT in 5.0 μg/ml,BAZ in 25 μg/ml) induced the lowest apoptosis index and the highest trypan blue positive rate (necrosis). In the mice bearing S 180 tumor,during 2～48hrs,and in the dose range of 25～250 mg/kg of BAZ,the apoptosis index was rising with the time passing and the dose increasing. Forty eight hours later,the apoptosis index decreased and the trypan blue positive rate increased. Conclusions:Antitumor drugs induce apoptosis of tumor cells,the relationship between dose effect and time effect was found in certain range of dose and time. High dose of drugs kills tumor cells by necrosis mainly. It implicated that the threshold value of intensity and time of damage may be important in the switching on and the regulating of apoptosis and necrosis.