Induction of Bystander and Abscopal Effects after Electroporation-Based Treatments.

Electroporation-based antitumor therapies, including bleomycin electrotransfer, calcium electroporation, and irreversible electroporation, are very effective on directly treated tumors, but have no or low effect on distal nodules. In this study, we aimed to investigate the abscopal effect following calcium electroporation and bleomycin electrotransfer and to find out the effect of the increase of IL-2 serum concentration by muscle transfection. The bystander effect was analyzed in in vitro studies on 4T1tumor cells, while abscopal effect was investigated in an in vivo setting using Balb/c mice bearing 4T1 tumors. ELISA was used to monitor IL-2 serum concentration. We showed that, similarly to cell treatment with bleomycin electrotransfer, the bystander effect occurs also following calcium electroporation and that these effects can be combined. Combination of these treatments also resulted in the enhancement of the abscopal effect in vivo. Since these treatments resulted in an increase of IL-2 serum concentration only in mice bearing one but not two tumors, we increased IL-2 serum concentration by muscle transfection. Although this did not enhance the abscopal effect of combined tumor treatment using calcium electroporation and bleomycin electrotransfer, boosting of IL-2 serum concentration had a significant inhibitory effect on directly treated tumors.

Correlation between the loss of intracellular molecules and cell viability after cell electroporation.

Control of membrane permeability to exogenous compounds by membrane electroporation can lead to cell death, which is related to permanent membrane damage, oxidation stress, leakage of intracellular molecules. In this study, we show that the predominant cell death modality after the application of high voltage electric pulses is related with inability to reseal of initial pores (first stage irreversible electroporation, FirEP). After moderately strong electric pulses, initial pores reseal, however, some cell still die later on due to electric field induced cell stress which leads to delayed cell death (late-stage irreversible electroporation, LirEP). According to our data, the period in which the majority of cells commit to either pore resealing or complete loss of barrier function depends on the intensity of electric field treatment but did not exceed 35 min. Additionally, we show that after electroporation using electric pulse parameters that induce LirEP, some cells can be rescued by supplementing medium with compounds obtained from irreversibly electroporated cells. We determined that the intracellular molecules that contribute to the increase of cell viability are larger than 30 kDa. This serves to prove that the loss of intracellular compounds plays a significant role in the decrease of cell viability after electroporation.