Efficiency of High and Low Voltage Pulse Combinations for Gene Electrotransfer in Muscle, Liver, Tumor and Skin

Abstract

Gene electrotransfer is gaining momentum as an efficient methodology for nonviral gene transfer. In skeletal muscle, data suggest that electric pulses play two roles: structurally permeabilizing the muscle fibers and electrophoretically supporting the migration of DNA toward or across the permeabilized membrane. To investigate this further, combinations of permeabilizing short high-voltage pulses (HV; hundreds of V/cm) and mainly electrophoretic long low-voltage pulses (LV; tens of V/cm) were investigated in muscle, liver, tumor, and skin in rodent models. The following observations were made: (1) Striking differences between the various tissues were found, likely related to cell size and tissue organization; (2) gene expression is increased, if there was a time interval between the HV pulse and the LV pulse; (3) the HV pulse was required for high electrotransfer to muscle, tumor, and skin, but not to liver; and (4) efficient gene electrotransfer was achieved with HV field strengths below the detectability thresholds for permeabilization; and (5) the lag time interval between the HV and LV pulses decreased sensitivity to the HV pulses, enabling a wider HV amplitude range. In conclusion, HV plus LV pulses represent an efficient and safe option for future clinical trials and we suggest recommendations for gene transfer to various types of tissues. 1261

Control by pulse parameters of DNA electrotransfer into solid tumors in mice

“…This high field is lower than that used for skin (200 V cm À1 ), but much higher than that used in the combination protocol for muscle (80-100 V cm À1 ). 18 The differences in transfection efficiency between different tissues, as well as different tumor histological types, can be explained by the unique characteristics of each tissue, 5,26 namely, the delivery and distribution of macromolecules, including plasmid DNA, is affected by the amount of DNases, necrosis and macrophage infiltration as well as increased interstitial fluid pressure and the amount of extracellular matrix components, such as collagen and hyaluronan. In addition, in vitro and in vivo studies have elucidated the effect of the shape, size and orientation of cells in tissue and cell density on electric field distribution, which are all crucial factors for effective electrotransfection.…”

“…[10][11][12][13][14][15][16] A new protocol with high transfection efficacy in muscle and skin has recently been described, consisting of a combination of one high-field (HV, B1000 V cm À1 ) and one (or several) low-field (LV, B100 V cm À1 ) pulse. [17][18][19][20][21][22][23] For simplicity, we called all along the paper 'field' what indeed is the voltage to electrode width ratio, that is, an experimentally controlled parameter. In these studies, the authors suggested two functions of these electric pulses.…”

A microfluidic biochip for the nanoporation of living cells

“…This temporary permeabilization of the plasma membrane allows genes or drugs entering into the cell cytosol. This technique is very attractive in various applications such as electrochemotherapy Mir et al 2006), cutaneous and subcutaneous tumour nodule treatments (Mir et al 1998), evaluation of the cytotoxicity of nonpermeant or poorly permeant anticancer drugs (Labanauskiene et al 2007), gene electrotransfer to various animal tissues (Andre and Mir 2004;Andre et al 2008) and latterly with in vivo tests (Gothelf and Gehl 2010;Heller and Heller 2006).…”

Therapeutic levels of erythropoietin (EPO) achieved after gene electrotransfer to skin in mice

“…Not only combinations of high voltage pulse (HV) and low voltage pulse (LV) pulses are efficient in skin as well as different kinds of tissues, 29 but also LV pulse combinations are effective in transfecting reporter genes into skin with plate electrodes. 22 …”