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HIV-1-mediated delivery of a short hairpin RNA targeting vascular endothelial growth factor in human retinal pigment epithelium cells
  1. G Lombardi1,
  2. A Calistri1,
  3. M Curtarello1,
  4. G L Giudice2,
  5. S Piermarocchi3,
  6. G Prosdocimo4,
  7. G Palù1,
  8. C Parolin5
  1. 1
    Department of Histology, Microbiology and Medical Biotechnologies, University of Padua, Padua, Italy
  2. 2
    Department of Ophthalmology, S. Antonio Hospital, Padua, Italy
  3. 3
    Department of Neurosciences, Section of Ophthalmology, University of Padua, Padua, Italy
  4. 4
    Department of Ophthalmology, Conegliano Hospital, Conegliano (Treviso), Italy
  5. 5
    Department of Biology, University of Padua, Padua, Italy
  1. Professor C Parolin, Department of Biology, University of Padua, Viale G. Colombo, 3, 35131 Padua, Italy; cristina.parolin{at}unipd.it

Abstract

Background: Vascular endothelial growth factor (VEGF) has been shown to play a major role in the pathological neovascularisation that occurs in degenerative retinal diseases like age-related macular degeneration (AMD). Although several approaches to attenuate VEGF show significant promise, repeated treatments are required to achieve therapeutic benefits. As lentiviruses efficiently and stably infect resting cells, a human immunodeficiency virus type 1 (HIV-1)-based vector was used for the delivery and long-term endogenous expression of a short hairpin RNA (shRNA) specific for VEGF in postmitotic human retinal pigment epithelium (RPE) cells.

Methods: An HIV-1 vector expressing a shRNA targeting VEGF was developed and adopted to transduce RPE cell cultures, in both normoxic and hypoxic conditions in vitro. Intracellular VEGF expression was analysed by western blotting, and the release of VEGF in culture supernatants was determined by ELISA.

Results: At least 90% of RPE cells were successfully transduced by HIV-1 virions. Inhibition of VEGF expression and reduction by 95% of VEGF release in transduced cells were achieved. Moreover, shRNA-VEGF effectively and specifically prevented hypoxia-induced VEGF upregulation.

Conclusion: HIV-1-mediated delivery of a shRNA-VEGF leading to gene expression knockdown could represent a novel therapeutic strategy against neovascularisation-related eye diseases.

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Footnotes

  • Competing interests: None.

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