Annexin V/PI双标记检测高电压脉冲电场诱导K562细胞凋亡的研究

Study on apoptosis of K562 cells staining with Annexin V/PI induced by high-voltage pulse electric field

目的:探讨高电压脉冲电场诱导红白血病细胞K562和淋巴细胞凋亡的最佳参数。方法:调节电场强度和脉冲频率分别作用K562细胞和淋巴细胞后,用Annexin V/PIKit标记凋亡细胞,流式细胞术检测凋亡细胞的百分比;设计正交试验确定电场诱导K562细胞凋亡的最优参数组合;电场作用K562细胞与淋巴细胞(1∶1)混合悬液进行选择性诱导凋亡;MTT法检测电场作用后的K562细胞与淋巴细胞的增殖。结果:K562细胞的凋亡率随着脉冲频率的增加而升高,当脉冲频率为30Hz时,凋亡率高达79.13%;K562细胞和淋巴细胞的凋亡率均随着电场强度的增加而升高,在电场强度达到30kV/cm时,二者的凋亡率分别达82.40%、57.08%。诱导K562细胞凋亡的最优参数组合为脉冲宽度1μs、电场场强25kV/cm、脉冲个数为12个。K562细胞与淋巴细胞混合液在电场作用下发生凋亡的细胞百分比具有显著性差异(P<0.05);K562细胞与淋巴细胞经相同条件的电场作用后,二者生存率比较,差异显著(P<0.05)。结论:利用高电压脉冲电场使癌细胞选择性失活是可行的。

Objective：To investigate the optimization parameters for apoptosis of erythroleukemia K562 cells and lymphocytes induced by high-voltage pulse electric field.Methods：The apoptotic cells of K562 cells and lymphocytes were labeled by Annexin V/PI under different electric field intensity and pulse frequency,and the percentage of apoptotic/dying cells were detected by FCM.Designing an orthogona experiment was to define the best parameter array.The mixed cell suspension comprising of K562 cells and lymphocytes（1∶1）treated with electric field was selectively induced to apoptosis.The proliferation of K562 cells and lymphocytes were assayed by MTT.Results：The percentage of apoptotic K562 cells was increased following by pulse frequency,and the maximum apoptotic ratio reached 79.13% in 30 Hz.The percentage of apoptotic K562 cells and lymphocytes was increased following by electric field intensity,and the apoptotic ratio reached 82.40%,57.08% in 30 kV/cm.The optimization parameter array was 1 μs pulse extent,25 kV/cm electric field intensity and 12 pulse counts.There was a significant difference between the apoptotic ratio of mixing K562 cells and lymphocytes（P〈0.05）.The difference between survival rate was significant（P〈0.05）.Conclusion：It is feasible to make selective inactivation of cancer cells by high intensity pulsed electric fields.