Non-Viral Gene Delivery Vehicles
An initial screen of several cationic polymers (branched PEI, linear
PEI, jetPEI, & Turbofect) and lipids (Lipofectamine LTX) in Jurkat T
cells revealed that the vehicles could be separated into three
significantly different cohorts (Figure 1A). For example, branched PEI
and some of the other polymeric vehicle formulations provided no
detectable luciferase expression, while Turbofect and a specific
nitrogen:phosphate ratio (N:P = 10:1) of linear PEI yielded modest
luciferase expression. The highest luciferase expression levels were
obtained with Lipofectamine LTX (Lipo-Lo = 1.25 µL and 1μg DNA; Lipo-Hi
= 2.75 µL and 1 μg DNA) and jetPEI (N:P = 5:1).
Subsequent transfections of pEF-GFP (Figure 1B) using the vehicles that
provided the highest luciferase expression levels (Lipofectamine,
Turbofect, and jetPEI) showed that the higher dose of Lipofectamine LTX
(Lipo-Hi) provided the highest transfection efficiency for Jurkat T
cells (25% EGFP+), while the transfection
efficiencies obtained with Turbofect and jetPEI were significantly lower
(4.5-4.8% EGFP+ cells). As observed with the
luciferase assays, the higher amount of Lipofectamine (Lipo-Hi) appeared
to provide a higher number of EGFP+ cells than the
lower amount (Lipo-Lo), but the difference was not statistically
significant. The histograms in Figure 1D-F also illustrate a distinct
increase in the number of fluorescent cells obtained with Lipo-Hi
compared to Lipo-Lo and Turbofect.
The effects of Lipofectamine, Turbofect, and jetPEI (5:1) on cell
metabolic activity were also measured with an MTT assay (Figure 1C).
Each vehicle significantly decreased cell metabolic activity
approximately 30-40% relative to control samples of untransfected
cells, but no significant differences were observed between the vehicles
themselves.
Overall, Lipofectamine LTX
appeared to be more efficient than Turbofect and branched or linear PEI.
This observation is in agreement with previous studies that have
compared the transfection efficiencies of cationic lipids and polymers
in other cell lines.31,32 These studies attribute the
higher transfection efficiency of the lipid vehicles to their higher
affinity for cell membranes, which allows them to be endocytosed more
quickly than cationic polymers like PEI.31Consequently, Lipofectamine (Lipo-Hi) was selected as the vehicle for
our subsequent studies.