基于阵列微泡的超声调控细胞膜通透性机制及方法研究

Ultrasound-triggered drug delivery with microbubbles is capable of releasing drug, improving the localized drug concentration, which has shown a promising applications in the cancer treatment. Improving the cell membrane permeability based on the stable cavitation is crucial for the ultrasound drug delivery as the drug can be delivered into cells. The mechanism of the sonoporation based on the stable cavitation, however, still unclear, and a method for parallel single-cell sonoporation is needed to be developed. To address this issue, this project will investigate the sonoporation mechanism at the single bubble-single cell level. Moreover, this project will fabricate the monodisperse microbubble array and excite all microbubbles cavitation simultaneously to realize the parallel sonoporation. The detailed research contents include: fabrication and generation of monodisperse microbubble array, obtaining the resonant frequency of the microbubble and excitation the bubble array resonant; trapping the cell at the microbubble surface by the second acoustic radiation force and quantitative analysis the influence of mechanical force and the shear stress induced by the stable cavitation on the sonoporation; dynamic quantitative analysis the change of transmembrane current during the sonoporation and resealing process by path clamp system. This project not only has the significance in understanding the interaction between the ultrasound and cells, but also has the potential applications in biomedical area for parallel single-cell sonoporation.

超声联合微泡介导药治疗可有效释放药物，提高局部药物浓度，在肿瘤治疗方面具有良好的应用前景。如何利用空化效应提高细胞膜通透性，促进大分子药物进入细胞内部是超声给药治疗的关键。然而，由于缺少并行化大规模单细胞声致穿孔方法，基于稳态空化效应的细胞穿孔机制并不清楚。针对以上问题，本项目研发阵列微泡芯片，同时激发微泡稳态空化，实现并行化细胞穿孔，在单微泡-单细胞水平上研究声致穿孔的物理机制。具体研究内容为：制备阵列微泡模板，利用表面张力产生阵列微泡，每个微泡粒径相同、相互独立，获取微泡共振频率，激发微泡稳态空化效应；通过微泡振动产生的二阶辐射力捕获细胞，量化分析微泡振动产生的机械应力及流体剪切力对细胞穿孔的影响；借助膜片钳技术测量跨膜电流，动态量化分析细胞开孔及恢复过程。本项目不仅在理解超声与细胞之间的相互作用方面具有重要科学意义，同时提出超声并行化单细胞穿孔方法在生物医学领域具有潜在应用价值。

超声联合微泡介导药治疗可有效释放药物，提高局部药物浓度，在肿瘤治疗方面具有良好的应用前景。如何利用空化效应提高细胞膜通透性，促进大分子药物进入细胞内部是超声给药治疗的关键。然而，由于缺少并行化大规模单细胞声致穿孔方法，基于稳态空化效应的细胞穿孔机制并不清楚。本项目基于标准MEMS工艺设计加工了尺寸均一可控的微气泡阵列，通过超声激发微气泡阵列的共振吸引、捕获细胞，利用气泡阵列的稳态空化，实现了单细胞水平的并行、高效的声致穿孔效应调控。针对微气泡阵列实现的高通量声致穿孔效应，通过单个微气泡对单细胞声致穿孔进行了详细的物理机制探究，分析了并量化了超声激励引起气泡振动产生的声辐射力大小、剪切力大小、流体曳力，研究表明超声辐射力导致细胞最终被捕获在振荡气泡表面，剪切力促进细胞表面微孔的生成。基于气泡阵列可调控细胞膜通透性的技术，揭示了该技术调控细胞膜通透性的具体物理机制，通过调控超声参数，实现了快速同源或非同源细胞的融合效应。进一步，基于气泡阵列调控细胞膜通透性的技术，通过调控超声参数，实现了快速、高效的细胞裂解效应。本项目不仅在理解超声与细胞之间的相互作用方面具有重要科学意义，同时提出超声并行化单细胞穿孔方法在生物医学领域具有潜在应用价值。