April 2010
Volume 51, Issue 13
ARVO Annual Meeting Abstract  |   April 2010
Distinct Biological Properties of Biochemically Distinct PEDF Protein Versions on Tumor Cells
Author Affiliations & Notes
  • M. M. Deshpande
    National Eye Institute, Bethesda, Maryland
  • S. Locatelli-hoops
    National Eye Institute, Bethesda, Maryland
  • P. Subramanian
    National Eye Institute, Bethesda, Maryland
  • S. Becerra
    National Eye Institute, Bethesda, Maryland
  • Footnotes
    Commercial Relationships  M.M. Deshpande, None; S. Locatelli-hoops, None; P. Subramanian, None; S. Becerra, None.
  • Footnotes
    Support  NIH
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 4506. doi:
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      M. M. Deshpande, S. Locatelli-hoops, P. Subramanian, S. Becerra; Distinct Biological Properties of Biochemically Distinct PEDF Protein Versions on Tumor Cells. Invest. Ophthalmol. Vis. Sci. 2010;51(13):4506.

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

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Purpose: : Pigment epithelium-derived factor (PEDF) is a multi-functional serpin protein. It is negatively regulated in tumor cells, inhibits tumor formation and metastasis, blocks angiogenesis, and it induces apoptosis in tumor and endothelial cells. In contrast, it promotes retinal cell survival and neuronal differentiation, protects RPE cells against oxidative stress, and plays a role in expansion of neural stem cells. The mechanisms that confer this multimodal functionality to PEDF are not clear.The purpose of this study is to investigate the biological activities of PEDF protein versions.

Methods: : Recombinant human PEDF was purified from the culture media of BHK cells harboring an expression plasmid with full length human PEDF cDNA by ammonium sulfate precipitation, followed sequentially by cation- and anion-exchange column chromatography.We used the human prostrate adenocarcinoma cell lines LNCaP and PC-3, the breast cancer cell lines MDA-231 and 4T1, and retinal R28 and ARPE-19 cells. Cell viability assays were performed using CellTiter-Glo (Promega) and Cell Counting Kit-8 (Dojindo). Real-time impedance of cells was followed with ACEA microelectronic system (Roche).

Results: : Two PEDF protein peaks were identified after anion-exchange column chromatography. The PEDF version eluting with lower ionic strength and having lesser negative charge was termed PEDF1. The version eluting with higher ionic strength and greater negative charge was labeled PEDF2.A marked difference in the biological action of PEDF1 and PEDF2 on cell viability was observed. PEDF2 emerged as being highly potent in lowering cell viability in all tumor cell lines tested. In contrast, PEDF1 minimally affected tumor cell viability but protected R28 cells against death due to serum starvation, and ARPE-19 cells against H2O2/TNF alpha-mediated damage.

Conclusions: : We have identified two distinct biochemical versions of PEDF. The two versions had clear distinct biological effects on tumor cell viability. The multifunctional modality of PEDF may be explained by the presence of differential post-translational modifications that may regulate its biological activity.

Keywords: apoptosis/cell death • protein purification and characterization • tumors 

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