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The aim of the present study was to construct ultrasound targeted microbubble destruction (UTMD) combined with a polyethylenimine/pEGFP/nuclear localization sequence (PEI/DNA/NLS) complex gene delivery system and to evaluate the transfection efficiency of enhanced green fluorescent protein (EGFP) gene delivery to 293T cells by this system.
The formation of PEI/DNA/NLS complexes and the protective effects of PEI/NLS were verified by gel electrophoresis. Solutions of plasmid DNA, PEI/DNA complexes, PEI/DNA/NLS complexes, SonoVue/DNA, SonoVue/PEI/DNA complexes, and SonoVue/PEI/DNA/NLS complexes were transduced into 293T cells via ultrasound irradiation. The expression of green fluorescent protein was observed using an inverted microscope and the transfection efficiency was detected by flow cytometry after a 24 h in vitro incubation. Cell activity was detected using a CCK-8 assay.
Gel electrophoresis confirmed the formation of PEI/DNA/NLS complexes and showed that PEI/NLS had protective effects on DNA but only for a limited time. Inverted microscope observations showed more green fluorescent protein could be expressed under the joint action of the PEI/DNA/NLS complexes with UTMD as well as demonstrated a higher transfection efficiency by flow cytometry analysis. Further, the activity of the cells detected by CCK-8 was greater than 80%.
This study successfully constructed UTMD combined with a PEI/DNA/NLS complex gene delivery system, demonstrating that the transfection efficiency of the system was higher than that of UTMD+PEI or PEI/DNA/NLS complexes alone, without subsequently increasing the rate of cell injury. It can potentially be used for the clinical application of gene transfection to treat ischemic cardiomyopathy and inherited diseases.