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Stephan Hoffmann, Rizwan Masood, Ya Zhang, Shikun He, Stephen J. Ryan, Parkash Gill, David R. Hinton; Selective Killing of RPE with a Vascular Endothelial Growth Factor Chimeric Toxin. Invest. Ophthalmol. Vis. Sci. 2000;41(8):2389-2393.
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purpose. To determine the sensitivity of retinal pigment epithelial (RPE) cells
to a vascular endothelial growth factor (VEGF) chimeric toxin.
methods. A targeted toxin was developed using recombinant methods to fuse
VEGF165 to the diphtheria toxin (DT) translocation and
enzymatic domain (DT390-VEGF165). Human RPE
cells, choroidal endothelial cells (CECs), and scleral fibroblasts were
isolated, and a dose–response for
DT390-VEGF165 was determined by measurement of
cell proliferation and cell number. In parallel experiments, cultures
were pretreated with transforming growth factor (TGF)-β2.
VEGF-receptor (VEGFR-1 and -2) expression was determined using reverse
transcription–polymerase chain reaction and fluorescence-activated
cell sorting, and affinity was measured using Scatchard analysis.
results. RPE cells and CECs were similarly prone to killing by the VEGF-toxin,
but scleral fibroblasts were unaffected. Pretreatment with
TGF-β2 selectively increased the sensitivity of RPE cells
to the VEGF-toxin. RPE cells expressed both VEGFR-1 and -2 in vitro;
however, the expression of VEGFR-1 was very low. Pretreatment with
TGF-β2 (10 ng/ml) was associated with increased
expression of the VEGFR-1 in RPE cells and increased receptor affinity
for VEGF detected by Scatchard analysis.
conclusions. Dose-dependent killing of RPE cells by the
DT390-VEGF165 conjugate is selectively enhanced
by pretreatment with TGF-β2. This study provides further
strong support for the presence of functional VEGFRs on human RPE
cells, and demonstrates for the first time the ability to target a
normal nonendothelial cell type through VEGFR
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