Vitamin C effect on cells exposed to external electric field

Abstract

Electroporation is based on the formation of pores in the cell membrane when cells are exposed to external electric fields (Chabot et al. 2013). Electroporation allows transmembrane delivery of various exogenous hydrophilic molecules, including a vitamin C (Markelc et al. 2012). Vitamin C is well known for its antioxidative properties. However, at high intracellular concentration vitamin C can induce significant cell toxicity leading to cell death (Maramag et al. 1997). Here we performed the in vitro analysis of vitamin C electrotransfer effect on cell viability during or after electroporation. CHO (Chinese Hamster Ovary) cell culture was used for experiments. Electroporation was performed by using electric field at 1400 V/cm voltage for the duration of 100 μs. Cells were electroporated in laboratory made EP medium (pH 7.1, conductivity 0.1 S/m, osmolarity 270 mOsm). Generation of ROS during electroporation was evaluated by incubating cells with DCFDA dye for 30 min prior to electroporation. 70 % of ethanol was added 10 min after electroporation and additionally incubated for 10 min. Afterwards fluorescence was evaluated with fluorimeter (Tecan GeniosPro). Cell viability change because of vitamin C addition before or after electroporation at concentrations from 0.01 to 5 mM was evaluated using clonogenic assay. For this, 400 cells were plated into 40 mm petri dishes with growth medium after 10 min of incubation post electroporation. Colonies were measured 6 days after treatment.[...]