April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Genomic Stability in Established RPE Transfectants
Author Affiliations & Notes
  • A. Vehr
    Ophthalmology, RWTH Aachen, Aachen, Germany
  • S. Johnen
    Department of Ophthalmology,
    RWTH Aachen University, Aachen, Germany
  • L. Wickert
    RBZ Rheinisches Bildungszentrum Köln gGmbH, Köln, Germany
  • G. Thumann
    Dept of Ophthalmology,
    RWTH Aachen University, Aachen, Germany
  • Footnotes
    Commercial Relationships  A. Vehr, None; S. Johnen, None; L. Wickert, None; G. Thumann, None.
  • Footnotes
    Support  None.
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 4098. doi:
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      A. Vehr, S. Johnen, L. Wickert, G. Thumann; Genomic Stability in Established RPE Transfectants. Invest. Ophthalmol. Vis. Sci. 2010;51(13):4098.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: : During the last few years intravitreal injection of neovascularization inhibitors have gained wide use in the treatment of neovascular AMD. However, in addition to the treatment burden of monthly intravitreal injections, long-term systemic safety and logistic concerns exist with the use of VEGF inhibitors, such as ranibizumab or bevacizumab. One solution to these concerns would be implantation of cells producing an anti-angiogenic factor. We have been able to stably transfect hRPE cells with PEDF, a neurotrophic and anti-angiogenic factor that could be used to deliver PEDF and also serve to replace degenerated RPE cells. However, before transplantation it is essential to insure that the cell’s genome has not been altered except for their expression of the inserted gene.

Methods: : During the last few years intravitreal injection of neovascularization inhibitors have gained wide use in the treatment of neovascular AMD. However, in addition to the treatment burden of monthly intravitreal injections, long-term systemic safety and logistic concerns exist with the use of VEGF inhibitors, such as ranibizumab or bevacizumab. One solution to these concerns would be implantation of cells producing an anti-angiogenic factor. We have been able to stably transfect hRPE cells with PEDF, a neurotrophic and anti-angiogenic factor that could be used to deliver PEDF and also serve to replace degenerated RPE cells. However, before transplantation it is essential to insure that the cell’s genome has not been altered except for their expression of the inserted gene.

Results: : A comparison to non-transfected human RPE cells showed that in our established transfectant clones the relative expression of PEDF was 280 to 760 times higher than in non-transfected cells. The gene expression of KRT8, KRT18, ZO-1, VEGFA, cathepsin D, NF-kB, c-ABL, and JNK1 was unchanged, and the expression of CRALBP, Bcl-2, and p53 was minimally, but not significantly down-regulated.

Conclusions: : We have shown that using electroporation protocols human RPE cells can be stably transfected without altering significantly the expression of genes that are specific and critical to the function of RPE cells in vivo.

Keywords: age-related macular degeneration • gene transfer/gene therapy • retinal pigment epithelium 
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