Ishii Atsuhiro, Hiruta Yuki, Heinemann Dag, Heisterkamp Alexander, Kanazawa Hideko, Terakawa Mitsuhiro
School of Integrated Design Engineering, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, 223-8522, Japan.
Faculty of Pharmacy, Keio University, 1-5-30 Shibakoen, Minato, Tokyo, 105-8512, Japan.
J Biophotonics. 2017 Dec;10(12):1723-1731. doi: 10.1002/jbio.201600323. Epub 2017 May 2.
Micro-/nanosphere-mediated femtosecond laser cell perforation is one of the high throughput technologies used for macro-molecule-delivery into multiple cells. We have demonstrated the delivery of plasmid-DNA/liposome complexes into cells using biodegradable polymer microspheres and a femtosecond laser and investigated the intracellular localization of the complexes by delivering fluorescence-labeled plasmid-DNA/liposome complexes into cells. The utilization of liposomes increases the number of complexes delivered into the cytoplasm by laser illumination, which contributed to the increased transfection rate. In the experiment involving polystyrene (PS) microspheres of different diameters, the fluorescence of the complexes was detected in the nucleus as well as cytoplasm after laser illumination for PS microspheres of 3.0 μm diameter. The direct delivery of complexes into the nucleus is probably attributed to the enhancement of the nuclear membrane permeability by the enhanced optical field obtained close to the nucleus. These revelations on the intracellular localization of foreign DNA would provide effective laser-based transfection. Picture: Intranuclear delivery of plasmid-DNA/liposome complexes by utilizing dielectric microspheres and a femtosecond laser.
微/纳米球介导的飞秒激光细胞穿孔是用于将大分子递送至多个细胞的高通量技术之一。我们已经证明了使用可生物降解的聚合物微球和飞秒激光将质粒-DNA/脂质体复合物递送至细胞中，并通过将荧光标记的质粒-DNA/脂质体复合物递送至细胞中来研究复合物在细胞内的定位。脂质体的使用增加了通过激光照射递送至细胞质中的复合物数量，这有助于提高转染率。在涉及不同直径聚苯乙烯（PS）微球的实验中，对于直径为3.0 μm的PS微球，激光照射后在细胞核以及细胞质中检测到了复合物的荧光。复合物直接递送至细胞核可能归因于靠近细胞核处获得的增强光场增强了核膜通透性。这些关于外源DNA在细胞内定位的发现将提供有效的基于激光的转染。图：利用介电微球和飞秒激光进行质粒-DNA/脂质体复合物的核内递送。
