纳秒电脉冲跨膜损伤铂耐药肿瘤细胞核质效应研究

Our previous investigations have manifested that nanosecond electric pulses (nsEP)induced DNA breaks in cisplatin-resistant human ovarian cancer cells COC1/DDP, and the circult analysis demonstrated the highest potential in the nucleoplasm. The findings suggested that nsEP can trans-membranous manupulate the nucleus.nsEP may be a technique to treat refractory cancers, considering the role of nucleus in platium resistance. The objective of the present study was to determine the posibility of using nsEP against chemoresistant cancers, and to explore physical and biological mechanisms. Platium-sensitive and -resistant human cancer cell pairs (COC1 and COC1/DDP, SKOV3 and SKOV3/DDP, A2780 and C30, and SW480 and SW480/Oxa) were subjected to nsEP. Potentials in subcellular units were calculated with an equvalent circult, the kinetics of DNA damages and cell death followed, the mode of cell-death (apoptosis and necrosis) explored, and the therapeutic efficiency in vivo tested. The expected findings can be used to interpret the response difference between chemosensitive and chemoresistant cells (partically whether the biological property of cells be a determinant for response to nsEP, and the response to anticaner drugs after nsEP exposure). The expected findings can also serve as a reference for developing nsEP therapeutic modalities and devices.

申请者在执行NSFC项目(11174376)中发现纳秒电脉冲(nsEP)可引起顺铂耐药人卵巢癌细胞DNA断裂和致细胞死亡，电路分析显示核质高电压；提示nsEP可跨膜实现对胞核的直接影响，而核为肿瘤铂耐药的重要靶点。本项目将进一步研究其生物物理机制。拟选择顺铂和/或奥沙利铂敏感和耐药肿瘤细胞对，(1)计算nsEP在细胞内亚细胞单元诱导的电压，及胞质内电压的分配；(2)探讨nsEP诱导DNA损伤和细胞死亡的特点和动态变化，及DNA损伤修复在其中的效应；探讨胞质电压对线粒体的损伤效应；(3)探讨nsEP致肿瘤细胞死亡的方式；阐明药物敏感和耐受肿瘤细胞对nsEP反应性的差异和特点，及nsEP对细胞化疗反应性的影响；(4)探讨nsEP治疗耐药肿瘤的体内疗效。预期阐明nsEP对耐药肿瘤细胞的作用特点。研究结果可用于探索新的耐药肿瘤非药物治疗手段，并可为相关仪器的开发提供理论参考数据。

对纳秒电脉冲(nsEP)对顺铂(DDP)耐药肿瘤细胞的影响进行探讨。(1) 多层电介模型计算表明不同的频域和不同的持续时间对细胞内膜和外膜有选择性的影响，时域、频域分析显示细胞上有一个窗口，当持续时间在10(-8)~10(-6) s，细胞内膜电压将高于细胞外膜电压。(2) nsEP致细胞DNA断裂、死亡在处理后2 h、8 h出现峰值；2 h断裂为SSB，8 h断裂为DSB；8 h凋亡率升高；提示nsEP致细胞死亡存在两种模式，早期细胞死亡与电损伤引起的细胞膜损伤相关，晚期细胞死亡主要由凋亡介导，二种死亡方式对耐药肿瘤治疗的意义不同。(3) 耐药细胞与非耐药细胞的反应性存在差异，耐药细胞损伤较轻、损伤修复更快，提示细胞的生物学特性是其对nsEP反应性的决定因子之一。(4) 成功建立裸鼠原位卵巢癌模型，该模型成功率高，发生腹腔、远处转移，可用于物理治疗实验，评价疗效时应以生存时间为第一疗效终点。上述结果提示nsEP可发展用于耐药肿瘤治疗，电场参数(强度、脉宽)需要个性化设定。