摘要
目的观察丹参酮ⅡA(Tanshinone,TanⅡA)联合三氧化二砷(ATO)对人急性早幼粒细胞白血病细胞株(NB4细胞)凋亡的作用;并通过观察两药联合诱导NB4细胞凋亡时p53、Bcl-2表达的变化,探讨其可能的机制。方法通过Annexin V FITC/PI荧光染色观察1.0μg·mL-1TanⅡA分别联合0.25、0.5、1.0μmol·L-1的ATO作用于NB4细胞的形态学变化;流式细胞术Annexin V-FITC/PI法测定各组细胞凋亡率;流式细胞仪检测各组细胞p53、Bcl-2的表达。结果 (1)1.0μg·mL-1TanⅡA分别与0.5、1.0μmol·L-1的ATO联合作用NB4细胞24、48和72h的凋亡率均较相应单独用药组凋亡率明显增高(P<0.05);染色荧光显微镜观察l.0μg·mL-1TanⅡA联合1.0μmol·L-1ATO实验组较1.0μmol·L-1ATO对照组72h时更易见到中晚期凋亡细胞和死亡细胞;(2)1.0μg·mL-1TanⅡA与0.5、1.0μmol·L-1的ATO联合作用NB4细胞48h、72h表达p53蛋白的细胞较1.0μmol·L-1ATO单药组明显增多(P<0.05),作用高峰时间为72h;1.0μg·mL-1TanⅡA分别与0.25、0.5、1.0μmol·L-1的ATO联合作用NB4细胞24h时,表达Bcl-2蛋白的细胞数较1.0μmol·L-1ATO单药组显著下降(P<0.05),且与ATO的浓度呈负相关(r=-0.858,P=0.000)。结论联合1.0μg·mL-1TanⅡA可增强0.5、1.0μmol·L-1ATO诱导NB4细胞凋亡;TanⅡA联合ATO诱导NB4细胞p53表达增强和Bcl-2表达减少可能是其促凋亡机制之一。
Objective To investigate the apoptosis effect induced by Tanshinone ⅡA （Tan ⅡA） combined with arsenic trioxide （ATO） in human acute promyelocytic leukemia （APL） cell line NB4 cells and to explore the potential mechanism by detecting the expression changes of P53 and Bcl-2.Methods The NB4 cells were treated with 1.0μg · mL^-1 Tan ⅡA combined with ATO 0.25 μmol · L^-1,0.5μmol · L^-1 and 1.0 μmol · L^-1 respectively.The morphological changes of the cells were demonstrated by Annexin V-FITC/PI immunocytochemistry and the expression of P53 and Bcl-2 was measured by flow cytometry （FCM）.Results （1） The proliferative rate of NB4 cells treated by 1.0μg · mL^-1 Tan Ⅱ A combined with 1.0μmol · L^-1 and 0.5μmol · L^-1 ATO respectively during 24,48 and 72h was significantly inhibited compared with those treated by corresponding ATO alone group （P ＜ 0.05）.1.0 μg · mL^-1 Tan Ⅱ A combined with 1.0 μmol · L^-1 ATO was more easily to see middle to late period apoptosis cells and dead cells compared with 1.0μmol · L^-1 ATO alone group at 72h by Annexin V-FITC/PI immunocytochemistry study under fluorescence microscopy.（2） The expression of P53 in these two groups 1.0μg · mL^-1 Tan Ⅱ A combined with 0.5μmol · L^-1 and 1.0μmol · L^-1 ATO was significantly higher than that in ATO alone group at 48h and 72h（P ＜0.05） and the effect reached peak at 72h.The expression of Bcl-2 was significantly decreased in 1.0μg · mL^-1 Tan Ⅱ A combined with all different dose of ATO at 24h compared with 1.0pmol · L^-1 ATO alone group（P ＜ 0.05）.ATO concentration was significantly negatively correlated to Bcl-2 （r =-0.858,P =0.000）.Conclusion Combination treatment of 1.0μg · mL^-1 Tan Ⅱ with 0.5,1.0μmol · L^-1 ATO has significant synergistic effects on the apoptosis of NB4 cells.The synergistic effects of Tan ⅡA and ATO induced NB4 cells increase the expression of p53 and decrease the expression of Bcl-2,which maybe one of the mechanism of APL cell apoptosis.
出处
《宁夏医科大学学报》
2014年第2期178-182,190,F0004,共7页
Journal of Ningxia Medical University
基金
宁夏医科大学重点孵育项目(XZ201010)
