Regulation of Pore Evolution via Progressive Electroporation Enhanced Intracellular Molecule Transport

Nonviral intracellular delivery based on pulsed-electric-field-induced electroporation is one of the most effective and widely used platforms in basic biological and biomedical research. However, the conventional bulk electroporation technique has exhibited limited performance in improving delivery efficiency with a single type of pulse, especially for in vivo small interfering RNA (siRNA) delivery. Pulse modulation has been confirmed effective in facilitating intracellular delivery. Nonetheless, pore evolution and regulation during and after electric exposure plays an essential role in the effective intracellular delivery of molecules with variable sizes. Here, we propose a progressive electroporation (PEP) strategy on the basis of multiple-pulse combination, which decouples the perforation process and delivery process compared to conventional bulk electroporation, efficiently improving delivery efficiency with regulation of the perforated pores. We demonstrated an important correlation between delivery efficiency enhancement and delayed pore resealing by quantitative investigations. The performance of this disruption-and-field-enhancement method also showed delivery advantages over conventional chemical systems. Moreover, we validated the improvement for siRNA knockdown efficacy in vivo. Overall, PEP helps provide a unique insight into improving intracellular delivery, by regulating pore dynamics rather than just inducing perforation. This strategic advancement of PEP may pave the way for the development of advanced wearable delivery systems with reduced energy consumption.