Hall Emily H, Schoenbach Karl H, Beebe Stephen J
Center for Pediatric Research, Eastern Virginia Medical School, Children's Hospital of the King's Daughters, Norfolk, Virginia 23510, USA.
DNA Cell Biol. 2005 May;24(5):283-91. doi: 10.1089/dna.2005.24.283.
Nanosecond pulsed electric fields (nsPEFs) are ultrashort pulses with high electric field intensity (kV/cm) and high power (megawatts), but low energy density (mJ/cc). To determine roles for p53 in response to nsPEFs, HCT116 cells (p53+/+ and p53-/-) were exposed to nsPEF and analyzed for membrane integrity, phosphatidylserine externalization, caspase activation, and cell survival. Decreasing plasma membrane effects were observed in both HCT116p53+/+ and p53-/- cells with decreasing pulse durations and/or decreasing electric fields. However, addition of ethidium homodimer-1 and Annexin-V-FITC post-pulse demonstrated greater fluorescence in p53-/- versus p53+/+ cells, suggesting a postpulse p53-dependent biological effect at the plasma membrane. Caspase activity was significantly higher than nonpulsed cells only in the p53-/- cells. HCT116 cells exhibited greater survival in response to nsPEFs than HL-60 and Jurkat cells, but survival was more evident for HCT116p53+/+ cells than for HCT116p53-/- cells. These results indicate that nsPEF effects on HCT116 cells include (1) apparent direct electric field effects, (2) biological effects that are p53-dependent and p53-independent, (3) actions on mechanisms that originate at the plasma membranes and at intracellular structures, and (4) an apparent p53 protective effect. NsPEF applications provide a means to explore intracellular structures and functions that can reveal mechanisms in health and disease.
纳秒级脉冲电场(nsPEFs)是具有高电场强度(千伏/厘米)和高功率(兆瓦)但能量密度低(毫焦/立方厘米)的超短脉冲。为了确定p53在响应nsPEFs中的作用,将HCT116细胞(p53+/+和p53-/-)暴露于nsPEF,并分析其膜完整性、磷脂酰丝氨酸外化、半胱天冬酶激活和细胞存活情况。随着脉冲持续时间的缩短和/或电场强度的降低,在HCT116p53+/+和p53-/-细胞中均观察到质膜效应减弱。然而,脉冲后添加碘化丙啶同型二聚体-1和膜联蛋白-V-异硫氰酸荧光素显示,p53-/-细胞中的荧光比p53+/+细胞更强,这表明在质膜上存在脉冲后p53依赖性生物学效应。仅在p53-/-细胞中,半胱天冬酶活性显著高于未脉冲处理的细胞。与HL-60和Jurkat细胞相比,HCT116细胞对nsPEFs的反应表现出更高的存活率,但HCT116p53+/+细胞的存活率比HCT116p53-/-细胞更明显。这些结果表明,nsPEF对HCT116细胞的影响包括:(1)明显的直接电场效应;(2)p53依赖性和非依赖性生物学效应;(3)对起源于质膜和细胞内结构的机制的作用;(4)明显的p53保护作用。nsPEF的应用提供了一种探索细胞内结构和功能的方法,这些结构和功能可以揭示健康和疾病中的机制。