J4-4324 — Annual report 2012
1.
Does cell membrane fluidity affects gene electrotransfer and/or electrofusion of CHO-K1 and B16-F1 cells?

When a cell is exposed to high intensity external electric field hydrophilic pores are fomrd in the cell membrane. The increased permeability enables transfer of drugs and molecules into the cells and in case of mutual cell contats also electrofusion of cells. Thus electroporation i simportnat for different applications from electrotrasnfection to electrofusion of cells. However the mechanism of pore formation on the cellular level are not completly understood. Only few studies analysed role of membrane fluidity. We therefore set a protocol for measuring fluidity by means of spectrofluorimetric measuremnts of anisotropy on two cell lines. We obtained that fluidity does not influence electrofusion while for gene electrotrasnfer further experiments are needed.

B.03 Paper at an international scientific conference

COBISS.SI-ID: 9393236
2.
Analysis of different mechanisms involved in gene electrotransfer

We peformed a broad analysis of electrotransfection in vitro from visualization on the membrane level to systematic comparison of protocols and theoretical analysis. We demonstrated that the electrophoresis is key for the efficient electrogene transfection in cases where plasmid DNA concentrations are relatively low (typically in vivo). In contrast, for in vitro the efficient electropermeabilization suffices for transfection. We also demostrate and explain observed differences on cells in suspension and plated cells, and also comment which methodology is more suitable for each applicationa. Presented results are important for understanding of the mechanisms of gene electrotransfection and for improvement of the protocols. We also determined that use of low DNA concentrations better represents in vivo conditions and allows better transfer of conclusions from in vitro to in vivo conditions.

B.03 Paper at an international scientific conference

COBISS.SI-ID: 9393492