纳秒级陡脉冲电场诱导人黑色素瘤细胞裸鼠皮下-第三军医大学学报.doc

纳秒级陡脉冲电场诱导人黑色素瘤细胞裸鼠皮下移植瘤凋亡作用及机制的研究* *第一作者: 杨方黎 ,女 ,26岁 ,硕士,研究方向:妇科肿瘤的临床治疗 联系电话电子邮箱:daisy-117@163.com 通讯作者: 唐均英,女,44岁,博士,教授,研究方向:妇科肿瘤的临床治疗 杨方黎 唐均英 米彦 姚陈果 王剑飞 重庆医科大学附属第一医院,400016 【摘要】 目的：观察纳秒级陡脉冲对人黑色素瘤细胞裸鼠皮下移植瘤的诱导凋亡作用、Caspase-3表达的影响。方法：取雌性4～6周龄BALB/C裸鼠20只,建立人黑色素瘤皮下移植瘤模型，设对照组和陡脉冲处理组,采用参数为300ns、20KV/cm、f=1hz的脉冲电场处理5分钟，分别观察处理组和对照组各时相瘤体体积大小变化，绘制抑瘤曲线；电镜观察肿瘤超微结构改变；免疫荧光检测caspase -3蛋白表达；RT-PCR方法检测caspase -3 mRNA表达；荧光分光光度计测定Ca2＋浓度。结果：陡脉冲治疗组裸鼠肿瘤体积明显小于对照组(P0.01)；电镜下出现典型的细胞凋亡改变；免疫荧光结果为caspase -3蛋白表达处理组比对照组明显增多；RT-PCR结果为caspase -3mRNA表达处理组比对照组明显增多；荧光分光光度计测量处理组Ca2＋浓度显著增加。结论：纳秒级陡脉冲电场能诱导在体人黑色素异体移植瘤细胞产生凋亡。 【关键词】纳秒级陡脉冲；人黑色素瘤细胞A375；凋亡；caspase -3； Ca2+ 浓度 Experimental studies on apoptosis of A375 cells of Xenografted tumor induced by nanosecond pulsed electric fild and It’s mechanism. 【Abstract】 Objective：To investigate the effect of Nanosecond Steep Pulse on growth and apoptosis in A375 cells of Xenografted tumor. Methods： After establishment of 4 to 6 week old female nude mouse model carried with human melanoma by subcutaneous inoculationg,20 mice were randomly divided into 2 groups, treatment group and control group,them were exposed to nanosecond steep pulse (20kV/cm,300ns,1 Hz in 5 minutes),to observe the changes of the tumor body volume and calculate it’s inhibition rate. The caspase-3 was determinated by immunofluorescence and RT-PCR.The apoptosis of A375 cells was detected by transmission electron microscope. The change of [Ca2+]i in A375 cells were observed by fluorescence spectrophotometer using Fluo-3/AM as the calcium fluorenscent indicator. Results：The tumor body volume of treatment group were smaller than control group(P0.01), The typical morphological features of apoptotic cells were observed by transmission electron microscope, the treatment group’s caspase-3 incresed compared with control group through immunofluorescence and RT-PCR. The [Ca2+]i was markedly increased by treatment with nanosecond steep pulse(P0.01). Conclusion：Nanosecond steep