Mechanical stimulation of cells for non-viral gene delivery

The main challenge of non-viral gene delivery is the design of effective and non-cytotoxic vectors and tools capable of targeted delivery of genetic material to intended sites to alter cellular function and/or structure (Pezzoli et al., 2012).Since their introduction, chemical vectors - cationic carriers able to self-assemble with anionic nucleic acids (NAs) into particles (complexes) in order to overcome cellular barriers - and physical methods - the application of membrane-disruptive forces to ease the NAs intracellular delivery - have made strides forward (Mehier-Humbert and Guy, 2005; Tros de Ilarduya et al., 2010). However, the poor efficiency of the former and the risk of potential cell damage of the latter are hampering their widespread application. In this context, we propose a novel in vitro gene delivery strategy relying on the delivery of polyethylenimine (PEI)-based polyplexes to mechanically stimulated cells, with the aim of easing the internalization of the genetic cargo, thus improving their transfection efficiency (TE). Specifically, high-frequencies vibrations (100 Hz) or cyclic deformation (10% cyclic strain) were used to induce a transient cell membrane destabilization in order to promote cell/complexes interactions thus increasing the uptake and the transgene expression. In our hands, when cells were properly stimulated, we observed a 10-to-100 fold-increase in TE of PEI/based polyplexes with respect to unstimulated transfected cells, with no effect on cell viability. Overall, coupling the use of a physical method with chemical vectors demonstrated to be an interesting technology to investigate the potential to drive effective gene transfer under well-defined mechanical environment.