April 2009
Volume 50, Issue 13
ARVO Annual Meeting Abstract  |   April 2009
Mechanistic Insight into the Interaction of Pigment-epithelium Derived Factor and Its Receptor PEDF-R for Retinal Cell Survival
Author Affiliations & Notes
  • P. Subramanian
    NEI/NIH, Bethesda, Maryland
  • S. Locatelli-Hoops
    NEI/NIH, Bethesda, Maryland
  • S. Choxi
    NEI/NIH, Bethesda, Maryland
  • G. M. Seigel
    SUNY, Buffalo, New York
  • S. P. Becerra
    NEI/NIH, Bethesda, Maryland
  • Footnotes
    Commercial Relationships  P. Subramanian, None; S. Locatelli-Hoops, None; S. Choxi, None; G.M. Seigel, None; S.P. Becerra, None.
  • Footnotes
    Support  None.
Investigative Ophthalmology & Visual Science April 2009, Vol.50, 2336. doi:
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      P. Subramanian, S. Locatelli-Hoops, S. Choxi, G. M. Seigel, S. P. Becerra; Mechanistic Insight into the Interaction of Pigment-epithelium Derived Factor and Its Receptor PEDF-R for Retinal Cell Survival. Invest. Ophthalmol. Vis. Sci. 2009;50(13):2336.

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

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Purpose: : Pigment epithelium-derived factor (PEDF), a neurotrophic factor, has high affinity receptors on the surface of target cells. PEDF-R is a transmembrane protein present in the retina. It exhibits phospholipase A2 (PLA2) activity and high affinity for PEDF. Recently we have identified a PEDF-binding region in PEDF-R. In this study, we investigate the biological significance of PEDF: PEDF-R interactions for survival of R28 rat retinal cells.

Methods: : Plasma membrane proteins were isolated from rat retina R28 cells with 0.1% NP-40. PEDF-R protein was detected by immunoblotting. PLA2 activity was determined by spectrophotometric assays. Cell survival and death was monitored using real-time microelectronic cell sensor system (ACEA) and CellTiter-Glo luminescent assay. Peptides were chemically synthesized and recombinant PEDF protein was purified from conditioned media of BHK cells harboring expression vectors for full-length human PEDF.

Results: : PEDF-R was immunodetected as an 80-kDa protein in the plasma membrane fractions of R28 cells. The immunodetected protein band was blocked by incubating the antibody with a specific peptide antigen. R28 plasma membrane fraction exhibited PLA2 activity, which was further stimulated by the addition of PEDF in a concentration-dependent manner. Serum starvation of R28 cells induced cell death. Exogenous addition of nanomolar concentrations of PEDF promoted R28 cell survival in a dose-dependent fashion. Peptides from the longest extracellular region of PEDF-R were generated and tested for PEDF binding. Preincubation of PEDF with PEDF-R-derived peptides containing PEDF affinity blocked the survival effect of PEDF on R28 cells. No effect on cell viability was observed with peptide alone or with peptides lacking PEDF affinity.

Keywords: cell membrane/membrane specializations • cell survival • neuroprotection 

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