Vol. 284, Issue 5, L817-L825, May 2003
Transfection of lung cells in vitro and in vivo: effect of
antioxidants and intraliposomal bFGF
Xiaoping
Luo1,
Rosetta
Belcastro2,3,
Judy
Cabacungan2,3,
Vicky
Hannam3,
Anna
Negus3,
Yanxia
Wen3,
Jonathan
Plumb3,
Jim
Hu2,3,
Brent
Steer3,
David R.
Koehler2,
Gregory P.
Downey3,4,5, and
A. Keith
Tanswell2,3,5,6
1 Department of Pediatrics, Tongji Hospital, Tongji
Medical College, Huazhong University of Science and Technology,
Wuhan 430074, People's Republic of China; 2 The
Canadian Institutes for Health Research Group in Lung Development
and 3 Lung Biology Programme, Hospital for Sick
Children Research Institute, and the Departments of
4 Medicine, 5 Physiology, and
6 Paediatrics, University of Toronto, Toronto, Ontario
M5G 1X8, Canada
We hypothesized that constitutive
formation of reactive oxygen species by respiratory cells is a barrier
to gene transfer when liposome-DNA complexes are used, by contributing
to rapid degradation of plasmid DNA. When plasmid DNA is complexed to
liposomes it is protected against H2O2-mediated
degradation but not that mediated by the hydroxyl radical. Treatment of
distal rat fetal lung epithelial cells (RFL19Ep) with the
vitamin E analog Trolox (50 µM) reduced intracellular plasmid
degradation. Both Trolox (50 µM) and an iron chelator,
phenanthroline (0.1 µM), significantly increased transgene
expression in RFL19Ep approximately twofold, consistent
with a hydroxyl radical-mediated inhibition of transgene expression.
When basic fibroblast growth factor (bFGF; 20 ng/ml), a growth factor
with antioxidant properties, was included within liposomes, we observed
a significantly greater enhancement of RFL19Ep transgene
expression (~2-fold) over that seen with Trolox or phenanthroline.
Inclusion of bFGF within liposomes also significantly enhanced
(~4-fold) transgene expression in mice following intratracheal instillation.
gene transfer; reactive oxygen species; growth factors; basic
fibroblast growth factor
