Correlation of the physicochemical and structural properties of pDNA/cationic liposome complexes with their in vitro transfection.
In this study, we characterized the conventional physicochemical properties of the complexes formed by plasmid DNA (pDNA) and cationic liposomes (CL) composed of egg phosphatidylcholine (EPC), 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) (50/25/25% molar ratio). We found that these properties are nearly unaffected at the studied ranges when the molar charge ratio (R(±)) between the positive charge from the CL and negative charge from pDNA is not close to the isoneutrality region (R(±) = 1). However, the results from in vitro transfection of HeLa cells showed important differences when R(±) is varied, indicating that the relationships between the physicochemical and biological characteristics were not completely elucidated. To obtain information regarding possible liposome structural modifications, small-angle X-ray scattering (SAXS) experiments were performed as a function of R(±) to obtain correlations between structural, physicochemical, and transfection properties. The SAXS results revealed that pDNA/CL complexes can be described as being composed of single bilayers, double bilayers, and multiple bilayers, depending on the R(±) value. Interestingly, for R(±) = 9, 6, and 3, the system is composed of single and double bilayers, and the fraction of the latter increases with the amount of DNA (or a decreasing R(±)) in the system. This information is used to explain the transfection differences observed at an R(±) = 9 as compared to R(±) = 3 and 6. Close to the isoneutrality region (R(±) = 1.8), there was an excess of pDNA, which induced the formation of a fraction of aggregates with multiple bilayers. These aggregates likely provide additional resistance against the release of pDNA during the transfection phenomenon, reflected as a decrease in the transfection level. The obtained results permitted proper correlation of the physicochemical and structural properties of pDNA/CL complexes with the in vitro transfection of HeLa cells by these complexes, contributing to a better understanding of the gene delivery process.
School of Chemical Engineering, University of Campinas, UNICAMP, Campinas, SP, Brazil.
Tiago A
TA
Balbino
Antônio A M
AA
Gasperini
Cristiano L P
CL
Oliveira
Adriano R
AR
Azzoni
Leide P
LP
Cavalcanti
Lucimara G
LG
de La Torre
D002412:Cations; D004247:DNA / Q000737:chemistry*; D005229:Fatty Acids, Monounsaturated / Q000737:chemistry; D006367:HeLa Cells; D006801:Humans; D008081:Liposomes / Q000737:chemistry*; D010713:Phosphatidylcholines / Q000737:chemistry; D010714:Phosphatidylethanolamines / Q000737:chemistry; D010957:Plasmids / Q000737:chemistry*; D000644:Quaternary Ammonium Compounds / Q000737:chemistry; D053838:Scattering, Small Angle; D055672:Static Electricity; D013329:Structure-Activity Relationship; D014162:Transfection / Q000379:methods*; D014961:X-Ray Diffraction
D002412:Cations; D005229:Fatty Acids, Monounsaturated; D008081:Liposomes; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; C020888:1,2-dielaidoylphosphatidylethanolamine; D004247:DNA; C070046:1,2-dioleoyloxy-3-(trimethylammonium)propane