Medical Engineering Program, University of Hong Kong, Pokfulam, Hong Kong SAR, China.
Ultrasound Med Biol. 2013 Jun;39(6):1075-86. doi: 10.1016/j.ultrasmedbio.2013.01.008. Epub 2013 Mar 15.
For sonoporation to be established as a drug/gene delivery paradigm, it is essential to account for the biological impact of this membrane permeation strategy on living cells. Here we provide new insight into the cellular impact of sonoporation by demonstrating in vitro that this way of permeating the plasma membrane may inadvertently induce repressive cellular features even while enhancing exogenous molecule uptake. Both suspension-type (HL-60) and monolayer (ZR-75-30) cells were considered in this investigation, and they were routinely exposed to 1-MHz pulsed ultrasound (pulse length, 100 cycles; pulse repetition frequency, 1 kHz; exposure period, 60 s) with calibrated field profile (spatial-averaged peak negative pressure, 0.45 MPa) and in the presence of microbubbles (cell:bubble ratio, 10:1). The post-exposure morphology of sonoporated cells (identified as those with calcein internalization) was examined using confocal microscopy, and their cell cycle progression kinetics were analyzed using flow cytometry. Results show that for both cell types investigated, sonoporated cells exhibited membrane shrinkage and intra-cellular lipid accumulation over a 2-h period. Also, as compared with normal cells, the deoxyribonucleic acid synthesis duration of sonoporated cells was significantly lengthened, indicative of a delay in cell cycle progression. These features are known to be characteristics of a cellular stress response, suggesting that sonoporation indeed constitutes as a stress to living cells. This issue may need to be addressed in optimizing sonoporation for drug/gene delivery purposes. On the other hand, it raises opportunities for developing other therapeutic applications via sonoporation.
为了使声孔作用成为一种药物/基因传递范例，必须考虑这种膜渗透策略对活细胞的生物学影响。在这里，我们通过体外实验提供了对声孔作用的细胞影响的新见解，证明这种渗透质膜的方法即使在增强外源性分子摄取的同时，也可能无意中诱导抑制性细胞特征。在这项研究中考虑了悬浮型（HL-60）和单层型（ZR-75-30）细胞，并且它们通常在存在微泡的情况下暴露于 1MHz 脉冲超声（脉冲长度为 100 个周期；脉冲重复频率为 1kHz；暴露时间为 60s），并具有校准的场分布（空间平均负压力峰值为 0.45MPa）。使用共聚焦显微镜检查声孔化细胞（鉴定为具有钙黄绿素内化的细胞）的暴露后形态，并用流式细胞术分析其细胞周期进展动力学。结果表明，对于研究的两种细胞类型，声孔化细胞在 2 小时内表现出细胞膜收缩和细胞内脂质积累。此外，与正常细胞相比，声孔化细胞的脱氧核糖核酸合成持续时间明显延长，表明细胞周期进展延迟。这些特征是细胞应激反应的特征，表明声孔作用确实对活细胞构成应激。在优化声孔作用以用于药物/基因传递目的时，可能需要解决这个问题。另一方面，它为通过声孔作用开发其他治疗应用提供了机会。
