Kinoshita Manabu, Hynynen Kullervo
Department of Radiology, Brigham and Women's Hospital, and Harvard Medical School, Boston, Massachusetts, USA.
Pharm Res. 2005 May;22(5):716-20. doi: 10.1007/s11095-005-2586-7. Epub 2005 May 17.
To investigate the possibility of intracellular delivery of Bak BH3 peptide using sonoporation effect by microbubble-enhanced ultrasound.
HeLa and BJAB cells were exposed to 1.696-Mhz focused ultrasound with 2% microbubble contrast agents (OPTISON). Cell-impermeable calcein was used as an indicator for successful sonoporation, and propidium iodide staining was used for cell viability assessment. Peptides were also exposed to ultrasound with OPTISON and analyzed with mass spectrometry for evaluation of stability under ultrasound exposure. The effect of transduced Bak BH3 peptide was evaluated by the cell viability of successfully sonoporated cells.
Bak BH3 peptides did not undergo mechanical degradation with microbubble-enhanced ultrasound exposure. With the increase of acoustic energy exposure, the sonoporation efficiency saturated both in BJAB and HeLa cells, while direct cell death rate by ultrasound exposure tended to increase. When BJAB cells were treated with 100 microM Bak BH3 peptides, and ultrasound exposure with ultrasound contrast agents (OPTISON), an increased 35% cell death was confirmed. On the other hand, although HeLa cells had a similar trend, they failed to exhibit statistical significance.
Our results suggest that microbubble-enhanced focused ultrasound peptide transduction is possible. Further optimization of ultrasound exposure conditions may be necessary.
研究利用微泡增强超声的声孔效应实现Bak BH3肽细胞内递送的可能性。
将HeLa细胞和BJAB细胞暴露于1.696兆赫兹聚焦超声及2%的微泡造影剂（OPTISON）中。使用细胞不可渗透的钙黄绿素作为声孔形成成功的指标，并用碘化丙啶染色评估细胞活力。肽也与OPTISON一起暴露于超声下，并用质谱分析以评估超声暴露下的稳定性。通过成功进行声孔形成的细胞的活力来评估转导的Bak BH3肽的效果。
Bak BH3肽在微泡增强超声暴露下未发生机械降解。随着声能暴露的增加，BJAB细胞和HeLa细胞的声孔形成效率均达到饱和，而超声暴露导致的直接细胞死亡率趋于增加。当用100微摩尔的Bak BH3肽处理BJAB细胞，并与超声造影剂（OPTISON）一起进行超声暴露时，确认细胞死亡率增加了35%。另一方面，虽然HeLa细胞有类似趋势，但未显示出统计学意义。
我们的结果表明，微泡增强聚焦超声介导的肽转导是可能的。可能需要进一步优化超声暴露条件。
