Lipid-mediated gene transfer of acidic fibroblast growth factor into human corneal endothelial cells

被引:22
作者
Dannowski, H
Bednarz, J
Reszka, R
Engelmann, K
Pleyer, U
机构
[1] Charite Univ Med Berlin, Dept Ophthalmol, D-13353 Berlin, Germany
[2] Univ Klinikum Hamburg Eppendorf, Dept Ophthalmol, D-20246 Hamburg, Germany
[3] AG Drug Targeting, Max Delbruck Ctr Mol Med, D-13092 Berlin, Germany
[4] Tech Univ Dresden, Univ Klinikum Carl Gustav Carus, Dept Ophthalmol, D-01307 Dresden, Germany
关键词
gene transfer; liposomes; fibroblast growth factor; corneal endothelial cells; cell; proliferation;
D O I
10.1016/j.exer.2004.08.024
中图分类号
R77 [眼科学];
学科分类号
100212 ;
摘要
The aim of this study was to optimize non-viral gene transfer conditions and investigate the effect of fibroblast growth factor-1 (FGF-1) gene transfer on human corneal endothelial cell (HCEC) proliferation. Five non-viral vectors (Lipofectin(TM), DMRIE-C(TM), DAC-30, Effectene(TM), FuGene(TM)6) were used to transfect HCEC with plasmids coding for enhanced green fluorescent protein (EGFP) and FGF-1. Transfection efficiency and toxicity (n = 6) were quantified and optimized using the EGFP construct by FRCS-analysis. Using optimal conditions HCEC were transfected with the FGF-1 plasmid and cell proliferation as well as expression of FGF-1 were determined at days 4 and 7 by counting and western blotting, respectively. Lipofectin (17 +/- 2.02%) transfected HCEC more successfully than DMRIE-C (11 +/- 1.46%), Effectene (9 +/- 0.62%), FuGene (9 +/- 0.93%) and DAC-30 (7 +/- 0.59%). Toxicity of the lipids ranged from 2 to 4%. Optimal HCEC proliferation was achieved with DAC-30/FGF-1 (P<0.05), whereas all other vectors did not result in significantly increased cell proliferation. However, all of the transfected cells produced FGF-1 in different amounts as indicated by western blotting. Efficient and almost non-toxic transfer of the FGF-1 gene into HCEC can be successfully achieved by lipid-based techniques. Using optimal conditions significantly increased cell proliferation was independent on gene transfer efficiency. This may indicate that even a low transfection rate is sufficient to produce a concentration of FGF-1 that will have a stimulatory effect on HCECs. (C) 2004 Elsevier Ltd. All rights reserved.
引用
收藏
页码:93 / 101
页数:9
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