The perforation of plasma membrane by ultrasound-driven microbubbles (i.e., sonoporation) provides a temporary window for transporting macromolecules into the cytoplasm that is promising with respect to drug delivery and gene therapy. To improve the efficacy of delivery while ensuring biosafety, membrane resealing and cell recovery are required to help sonoporated cells defy membrane injury and regain their normal function. Blebs are found to accompany the recovery of sonoporated cells. However, the spatiotemporal characteristics of blebs and the underlying mechanisms remain unclear. With a customized platform for ultrasound exposure and 2-D/3-D live single-cell imaging, localized membrane perforation was induced with ultrasound-driven microbubbles, and the cellular responses were monitored using multiple fluorescent probes. The results indicated that localized blebs undergoing four phases (nucleation, expansion, pausing and retraction) on a time scale of tens of seconds to minutes were specifically involved in the reversibly sonoporated cells. The blebs spatially correlated with the membrane perforation site and temporally lagged (about tens of seconds to minutes) the resealing of perforated membrane. Their diameter (about several microns) and lifetime (about tens of seconds to minutes) positively correlated with the degree of sonoporation. Further studies revealed that intracellular calcium transients might be an upstream signal for triggering blebbing nucleation; exocytotic lysosomes not only contributed to resealing of the perforated membrane, but also to the increasing bleb volume during expansion; and actin components accumulation facilitated bleb retraction. These results provide new insight into the short-term strategies that the sonoporated cell employs to recover on membrane perforation and to remodel membrane structure and a biophysical foundation for sonoporation-based therapy.

超声驱动的微泡对质膜的穿孔（即，sonoporation) 为将大分子转运到细胞质中提供了一个临时窗口，这在药物递送和基因治疗方面是有希望的。为了在确保生物安全的同时提高递送效率，需要膜重新密封和细胞恢复，以帮助经声穿孔的细胞抵抗膜损伤并恢复其正常功能。发现气泡伴随着声穿孔细胞的恢复。然而，气泡的时空特征和潜在机制仍不清楚。借助用于超声曝光和 2-D/3-D 活单细胞成像的定制平台，通过超声驱动的微泡诱导局部膜穿孔，并使用多个荧光探针监测细胞反应。结果表明，局部气泡经历了四个阶段（成核、膨胀、暂停和回缩）在几十秒到几分钟的时间尺度上专门参与可逆声穿孔细胞。气泡在空间上与膜穿孔部位相关，并且在时间上滞后（大约几十秒到几分钟）穿孔膜的重新密封。它们的直径（大约几微米）和寿命（大约几十秒到几分钟）与声孔化程度呈正相关。进一步的研究表明，细胞内钙瞬变可能是触发起泡成核的上游信号。胞吐溶酶体不仅有助于穿孔膜的重新密封，而且还有助于膨胀过程中气泡体积的增加；和肌动蛋白成分的积累促进了气泡的收缩。