10.1021/mp200374e.s002
Francesco Cardarelli
Francesco
Cardarelli
Daniela Pozzi
Daniela
Pozzi
Angelo Bifone
Angelo
Bifone
Cristina Marchini
Cristina
Marchini
Giulio Caracciolo
Giulio
Caracciolo
Cholesterol-Dependent
Macropinocytosis and Endosomal
Escape Control the Transfection Efficiency of Lipoplexes in CHO Living
Cells
American Chemical Society
2012
lipid composition
cationic liposome formulations
imaging approaches
DOTAP
intracellular delivery
zwitterionic lipid dioleoylphosphatidylethanolamine
zwitterionic helper dioleoylphosphocholine
DNA
intracellular fate
uptake mechanism
Transfection Efficiency
physicochemical properties
Chinese hamster ovary
cell uptake
endosomal compartments
DOPE
DOPC
lysosome marker
lipoplexe
Endosomal Escape Control
CHO Living CellsHere
lipoplex macropinocytosis
2012-02-06 00:00:00
Journal contribution
https://acs.figshare.com/articles/journal_contribution/Cholesterol_Dependent_Macropinocytosis_and_Endosomal_Escape_Control_the_Transfection_Efficiency_of_Lipoplexes_in_CHO_Living_Cells/2553337
Here we investigate the cellular uptake mechanism and
final intracellular
fate of two cationic liposome formulations characterized by similar
physicochemical properties but very different lipid composition and
efficiency for intracellular delivery of DNA. The first formulation
is made of cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane
(DOTAP) and the zwitterionic helper dioleoylphosphocholine (DOPC),
while the second one is made of the cationic 3β-[<i>N</i>-(<i>N</i>,<i>N</i>-dimethylaminoethane)-carbamoyl]
cholesterol (DC-Chol) and the zwitterionic lipid dioleoylphosphatidylethanolamine
(DOPE). Combining pharmacological and imaging approaches we show that
both DOTAP-DOPC/DNA and DC-Chol-DOPE/DNA lipoplexes are taken up in
Chinese hamster ovary (CHO) living cells mainly through fluid-phase
macropinocytosis. Our results also indicate that lipoplex macropinocytosis
is a cholesterol-sensitive uptake mechanism. On the other side, both
clathrin-mediated and caveolae-mediated endocytosis play a minor role,
if any, in the cell uptake. Colocalization of fluorescently tagged
lipoplexes and Lysosensor, a primary lysosome marker, reveals that
poorly efficient DOTAP-DOPC/DNA lipoplexes are largely degraded in
the lysosomes, while efficient DC-Chol-DOPE/DNA systems can efficiently
escape from endosomal compartments.