Ultrasound enhances retrovirus-mediated gene transfer.
Viral vector systems are efficient for transfection of foreign genes into many tissues. Especially, retrovirus based vectors integrate the transgene into the genome of the target cells, which can sustain long term expression. However, it has been demonstrated that the transduction efficiency using retrovirus is relatively lower than those of other viruses. Ultrasound was recently reported to increase gene expression using plasmid DNA, with or without, a delivery vehicle. However, there are no reports, which show an ultrasound effect to retrovirus-mediated gene transfer efficiency. Retrovirus-mediated gene transfer systems were used for transfection of 293T cells, bovine aortic endothelial cells (BAECs), rat aortic smooth muscle cells (RASMCs), and rat skeletal muscle myoblasts (L6 cells) with beta-galactosidase (beta-Gal) genes. Transduction efficiency and cell viability assay were performed on 293T cells that were exposed to varying durations (5 to 30 seconds) and power levels (1.0 watts/cm(2) to 4.0 watts/cm(2)) of ultrasound after being transduced by a retrovirus. Effects of ultrasound to the retrovirus itself was evaluated by transduction efficiency of 293T cells. After exposure to varying power levels of ultrasound to a retrovirus for 5 seconds, 293T cells were transduced by a retrovirus, and transduction efficiency was evaluated. Below 1.0 watts/cm(2) and 5 seconds exposure, ultrasound showed increased transduction efficiency and no cytotoxicity to 293T cells transduced by a retrovirus. Also, ultrasound showed no toxicity to the virus itself at the same condition. Exposure of 5 seconds at the power of 1.0 watts/cm(2) of an ultrasound resulted in significant increases in retrovirus-mediated gene expression in all four cell types tested in this experiment. Transduction efficiencies by ultrasound were enhanced 6.6-fold, 4.8-fold, 2.3-fold, and 3.2-fold in 293T cells, BAECs, RASMCs, and L6 cells, respectively. Furthermore, beta-Gal activities were also increased by the retrovirus with ultrasound exposure in these cells. Adjunctive ultrasound exposure was associated with enhanced retrovirus-mediated transgene expression in vitro. Ultrasound associated local gene therapy has potential for not only plasmid-DNA-, but also retrovirus-mediated gene transfer.
Naka
Toshio
T
Department of Internal Medicine, Division of Coronary Heart Disease, Hyogo College of Medicine, Nishinomiya, Hyogo, Japan.
Sakoda
Tsuyoshi
T
Doi
Takashi
T
Tsujino
Takeshi
T
Masuyama
Tohru
T
Kawashima
Seinosuke
S
Iwasaki
Tadaaki
T
Ohyanagi
Mitsumasa
M
D000818:Animals; D001011:Aorta / Q000166:cytology / Q000473:pathology / Q000528:radiation effects*; D002417:Cattle; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology / Q000473:pathology / Q000528:radiation effects*; D005786:Gene Expression Regulation / Q000502:physiology / Q000528:radiation effects*; D018014:Gene Transfer Techniques; D051379:Mice; D032446:Myoblasts / Q000166:cytology / Q000473:pathology / Q000528:radiation effects*; D051381:Rats; D012190:Retroviridae / Q000235:genetics / Q000502:physiology / Q000528:radiation effects*; D013997:Time Factors; D014161:Transduction, Genetic; D014162:Transfection; D014465:Ultrasonics; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D001616:beta-Galactosidase