OPTIMIZING SHOCK WAVE-MEDIATED TRANSFECTION: A PARAMETRIC STUDY

Not all medications can effectively reach their targets by conventional administration methods.Alternative drug delivery methods, such as shock wave-mediated drug delivery (SWDD), have been proposed to overcome this challenge [1].SWDD is a physical method that has been under study for three decades [2].Despite progress, its efficiency is still low and has yet to be used clinically.Mechanical parameters, such as cavitation and shock wave characteristics, like amplitude and impulse, are presumed to play roles in SWDD [2].In order to improve the current understanding of these phenomena, we ultimately aim to study the interaction of shock waves with individual cells.Here, we present a novel and smaller-scale experimental setup that yielded a significant improvement (at least 40%) [4] in shock wave-mediated transfection efficiency with a specific cell type and Green Fluorescence Protein (GFP).This approach can pave the way for future microfluidic development and, in the long run, clinical application.Shock waves are high-amplitude, short-duration mechanical pulses that can noninvasively reach deep inside the body with negligible heating effects.These features give them superiority over other methods to be used as a means of drug delivery.Despite significant research, SWDD struggles to achieve its full potential due to a limited understanding of the key factors influencing its effectiveness [2].With a more detailed parametric study of the involved variables on a microfluidic scale and real-time monitoring of the phenomenon, better optimization will be achievable.In our apparatus, shown in Fig. 2 and3, shock waves were produced using a high-voltage electric spark gap in water.The experiments were conducted at 5 and 10 kV with varying shock wave exposures, while other parameters remained constant.We used human embryonic kidney (HEK293) cells and pEGFP-N1, a GFPencoding plasmid, for transfection.Transfection efficiency and cell viability were assessed using flow cytometry and trypan blue assay, respectively.The optimal DNA concentration for transfection was determined through titration experiments.Two flow cytometry readout times were also examined: 24-and 48-h.Preliminary transfection results, Fig. 4, identified 50 g/ml and 24-hour incubation as optimal.Transfection results, presented in Fig. 1 (a), show a different trend than previously reported in the literature [2, 4].In those works, it was shown that increasing the number of shock wave exposures increased dye internalization/ transfection efficiency and decreased cell viability.However, our results show that increasing the number of shock wave exposures initially raises the transfection efficiency but soon reaches a plateau.Increasing the number of shock waves decreases cell viability, as expected based on the literature.The trend of transfection efficiency matches our dye internalization results, which are partially presented in [3].Transfection efficiency is significantly influenced by the voltage (released energy); shock waves generated at 5 kV appear largely ineffective for inducing transfection.So, we suggest weaker shock waves shouldn't be used in future studies.By refining the shock wave parameters and potentially transitioning to a microfluidic platform, this approach has the potential to enhance SWDD efficiency significantly.