Cationic lipid-mediated gene transfer: effect of serum on cellular uptake and intracellular fate of lipopolyamine/DNA complexes

• Published in: Biochimica et biophysica acta Vol. 1368; no. 2; pp. 276 - 288
• Main Authors: Escriou, Virginie, Ciolina, Carole, Lacroix, Florence, Byk, Gerardo, Scherman, Daniel, Wils, Pierre
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 19.01.1998
• Subjects: 3T3 Cells; Animals; Blood; Cationic lipid; Cations; Cell Line; Cellular uptake; Cytoplasm - chemistry; DNA; Fibroblasts; Fluorescent Dyes; Gene transfer; Lipids; Mice; Plasmids - analysis; Plasmids - genetics; Plasmids - metabolism; Rhodamines; Serum; Spermine - chemistry; Transfection - methods; DOPE, dioleoyl phosphatidylethanolamine; lipofection, cationic lipid-mediated transfection; MEM, minimum essential medium; Cellular uptake; lipoplexes, cationic lipid/DNA complexes; Gene transfer; DMEM, Dulbecco's modified Eagle's medium; Cationic lipid; DNA; FBS, fetal bovine serum; Serum; PBS, phosphate-buffered saline, pH 7.4
• ISSN: 0005-2736; 0006-3002; 1879-2642
• DOI: 10.1016/S0005-2736(97)00194-6

Most of the cationic lipids used for gene transfer experiments drastically lose their efficiency in the presence of serum. We used a cationic lipid with a spermine head group and its fluorescent analog to study the cellular uptake and the intracellular fate of lipoplexes in the presence and absence of serum. We found that the amount of DNA and lipid taken up by the cells was not related to the efficacy of the gene transfer. When the lipofection was performed in the presence of serum, lipoplexes were contained within small intracellular vesicles. In the absence of serum, the vesicles were larger and heterogeneous in size and shape. By analysis of their size distribution, we showed that lipoplexes preformed in the absence of serum tended to aggregate. This aggregation was inhibited in the presence of serum. We used a carbonate formulation that led to the preformation of large particles: those large particles gave a high lipofection efficiency in the presence of serum and their intracellular distribution was identical to that observed in the absence of serum.