鸦胆子油乳注射液联合热疗对人膀胱癌细胞株BIU-87细胞增殖及p-Akt表达的影响

Effects of brucea javanica oil emulsion combined with thermotherapy on proliferation and p-Akt expression in human bladder cancer BIU-87 cells

目的探讨鸦胆子油乳注射液(简称鸦油乳)联合热疗对人膀胱癌细胞株BIU-87膀胱癌细胞形态、增殖、细胞周期及磷酸化丝氨酸苏氨酸蛋白激酶(p-Akt)蛋白表达的影响。方法采用倒置光学显微镜观察鸦油乳(10 ml/L)与热疗单独或联合对人BIU-87膀胱癌细胞形态学的影响;用四甲基偶氮唑蓝(MTT)法检测鸦油乳(5、10、20、30 ml/L)与热疗单独或联合对人BIU-87膀胱癌细胞增殖的抑制作用;采用流式细胞仪检测鸦油乳(10 ml/L)与热疗单独或联合对人BIU-87膀胱癌细胞周期的影响;用蛋白质印迹法(Western blotting)检测鸦油乳(10 ml/L)与热疗单独或联合作用于人BIU-87膀胱癌细胞24 h后p-Akt蛋白的表达。结果倒置光学显微镜结果显示,热疗组、鸦油乳组和鸦油乳联合热疗组的细胞数均明显减少,部分细胞变圆、体积变小、核固缩,但鸦油乳联合热疗组变化最明显,活细胞数最少。与热疗组和鸦油乳组比较,鸦油乳联合热疗对BIU-87膀胱癌细胞增殖的抑制作用显著增强(P<0.05),呈时间和剂量依赖性;鸦油乳联合热疗作用于人BIU-87膀胱癌细胞24、48、72 h的半数抑制浓度(IC50)分别为84、62、35 mg/L,低于单药鸦油乳的101、79、57 mg/L。鸦油乳联合热疗组作用24 h后人BIU-87膀胱癌细胞S期比率为(16.79±2.78)%,显著低于热疗组的(29.52±3.31)%和鸦油乳组的(26.34±0.86)%(P<0.05)。鸦油乳联合热疗组p-Akt蛋白的相对表达量为(0.51±0.02),显著低于热疗组的(0.98±0.05)和鸦油乳组的(0.70±0.04)(P<0.05)。结论鸦油乳联合热疗能增强对人BIU-87膀胱癌细胞的增殖抑制作用,下调p-Akt蛋白表达可能是其作用机制之一。

Objective To study the effects of brucea javanica oil emulsion combined with thermotherapy on the morphology, prolifera-tion, cell cycle and p-Akt expression of human bladder cancer BIU-87 cells. Methods Cytomorphology of human bladder cancer BIU-87 cells treated by brucea javanica oil emulsion（ 10 ml/L） or hyperthermia alone or in combination was observed by inverted optical microscope. The proliferation of human bladder cancer BIU-87 cells treated by brucea javanica oil emulsion （5, 10, 20 and 30 ml/L） or hyperthermia alone or in combination was assessed by MTT. Cell cycles of human bladder cancer BIU-87 cells treated by brucea javanica oil emulsion or hyperthermia alone or in combination were detected by flow cytometer. The expression of p-Akt treated by brucea javanica oil emulsion or hyperthermia alone or in combination was determined by Western blotting. Results Under optical microscope, the number of tumor cells in brucea javanica oil emulsion group, hyperthermia group and combined groups decreased obviously, and some cells became round and smaller with nucleus pyknosis, especially in the combined group. Compared with the other groups, the combined group showed stronger inhibition to the proliferation of human bladder cancer BIU-87 cells in a time-dose dependent manner （ P 〈 0. 05 ）. The IC50 of the combined group was 84, 62 and 35 mg/L at 24-, 48- and 72-hour, lower than that （ 101 , 79 and 57 rag/L） of the brueea javanica oil emulsion group. The rate of the tumor cells arrested at S-phase was （ 16.79 ± 2.78 ） % in combination group, lower than that [ （29.52 ± 3.31 ） % ] in the hyperthermia group and that [ （26.34 ± 0.86） % ] in the brucea javanica oil emulsion group （ P 〈 0.05 ）. The p-Akt expression was （0.51 ± 0.02）, lower than that （0.98 ± 0.05 ） in the hyperthermia group and that （0.70 ± 0.04） in the brucea javanica oil emulsion group （ P 〈 0.05 ）. Conclusion Brucea javanica oil emulsion combined with hyperthermia enhances the inhibition to the proliferation of human bladder cancer BIU-87 cells, and the mechanism may be related to the down-regulation of p-Akt.