基于声表面波微流控芯片的单细胞可修复声致穿孔机理研究

Sonoporation employs the acoustic cavitation of microbubbles to induce transient disruption of cell membranes, thereby enabling transport of exogenous drug-loaded micro/nano particles and DNA molecules into the cytoplasm of living cells. Sonoporation has shown great promise in the drug delivery and gene therapy. However, there is still no method or tool suitable for controlling the initial the radius and cavitation position of the microbubbles, as well as the inertial cavitation dose, which limits the safety and effectiveness of the sonoporation. This project integrates the monodisperse microbubble fabrication, spatial manipulation and cavitation into a surface acoustic wave (SAW)-based microfluidic device, aiming to real-time, quantitative and precise investigate the interactions between the microbubble and cell. The critical shear stress generated by cavitation microjetting that induces the repairable sonoporation will be analyzed by a high-speed micro-PIV system. Furthermore, when the initial radius and the cavitation position of the microbubble are both controlled, the influence of the acoustic field (waveform, frequency and intensity), microbubble radius, and the distance between the microbubble and the cell on the membrane integrity and cell viability will be revealed by analysis of the single cell sonoporation. The results of proposed studies contribute to improve the safety and effectiveness of sonoporation and establish a foundation for the clinical applications of gene therapy. This project is of great scientific and practical importance.

声致穿孔效应是利用微泡空化作用，瞬间提高细胞膜的通透性，能促进微纳药物颗粒、DNA分子等进入细胞内，在药物输送、基因治疗等方面显示出巨大潜力。然而，由于目前尚无合适方法控制微泡初始粒径和空化位置、精确调控微泡瞬态空化剂量，声致穿孔效应的安全性和有效性受到制约。本项目将单分散微泡制备、微泡及细胞空间定点操控、微泡瞬态空化通过声表面波微流控芯片有机集成,旨在实时、定量、精确研究微泡与细胞之间的相互作用；利用高速Micro-PIV技术，分析微泡空化激发的微激流在细胞膜表面形成可修复声孔对应的临界剪切应力；研究微泡尺寸、微泡细胞相对位置均可精确调控条件下，声场（波形、频率、强度）、微泡粒径、微泡与细胞距离等参量与细胞穿孔程度及活性的关系。项目成果有助于提高声致穿孔的安全性和有效性，为声致穿孔在基因治疗的临床应用奠定基础，具有重要科学意义和应用价值。

声致穿孔效应是利用微泡空化作用，提高细胞膜的通透性，能促进微纳药物颗粒、DNA 分子等进入细胞内，在药物输送、基因治疗等方面显示出巨大潜力。然而，由于目前尚无合适方法精确控制微泡初始粒径和空化位置，声致穿孔效应的安全性和有效性受到制约。本项目将微泡及细胞空间定点操控、微泡瞬态空化通过声表面波微流控芯片有机集成，旨在实时、定量、精确研究微泡与细胞之间的相互作用。在超声精确操控方面，利用倾斜形叉指换能器实现了对不同空间位置微泡的选择形状操控，通过声表面波联合体波操控实现了对不同粒径颗粒的多尺度操控与分选；在超声的导致细胞穿孔的研究中，通过对声场中相位的调控，实现对微泡与细胞之间相对距离的精确操控，当微泡靠近细胞时，提高声压，使微泡破碎，可实现对单细胞的声致穿孔效应，穿孔效率达到82.4%±6.5%，细胞的存活率为90%±8.7%。进一步，利用超声热效应实现对热敏脂质体的释放，通过对聚焦型叉指换能器施加不同的射频信号，可使得超声加热的温度稳定控制在37℃，42℃，50℃，结果表明当温度为42℃可导致热敏脂质体释放，并且超声声流效应可促进药物释放，提高细胞摄取效率。本项目的研究提高了声致穿孔的效率，保证了细胞的活性，为声致穿孔在基因治疗的临床应用奠定基础，具有重要科学意义和应用价值。