May 2008
Volume 49, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2008
Tracking of Flt23-KDEL Intraceptor in Endothelial Cells
Author Affiliations & Notes
  • H. Uehara
    Ophthalmology, Med Coll of Georgia, Augusta, Georgia
  • J. Simonis
    Ophthalmology, Med Coll of Georgia, Augusta, Georgia
  • N. Singh
    Ophthalmology, Med Coll of Georgia, Augusta, Georgia
  • B. Ambati
    Ophthalmology, Med Coll of Georgia, Augusta, Georgia
  • Footnotes
    Commercial Relationships  H. Uehara, None; J. Simonis, None; N. Singh, None; B. Ambati, None.
  • Footnotes
    Support  NIH
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 4588. doi:
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      H. Uehara, J. Simonis, N. Singh, B. Ambati; Tracking of Flt23-KDEL Intraceptor in Endothelial Cells. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4588.

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

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Abstract

Purpose: : Vascular endothelial growth factor (VEGF) and its homologues are considered as a principal mediators angiogenesis. Although these are promising targets of medical reagent to inhibit angiogenesis, VEGF inhibition as a therapeutic strategy has not reached its full potential. Our laboratory has previously shown that Flt intraceptors, intracellularly expressed recombinant subunits of Flt (VEGF receptor 1) with KDEL (Flt23K), an endoplasmic reticulum retention peptide sequesters VEGF within ER and disrupts VEGF autocrine loops. Flt intraceptors inhibited hypoxia-induced VEGF and injury-induced corneal angiogenesis. In this study, we sought to track Flt23K expression in living cells to elucidate how to optimize its inhibitory ability.

Methods: : We synthesized pCMV.Flt23-YFP-KDEL, and observed fluorescence in transfected culture cells with fluorescence microscope and confocal microscope.

Results: : The most common fluorescence pattern is fluorescence deposits over cytoplasm. It is possible these are present in ER, but seem more consistent with deposition in other vesicles such as proteasome or lysosome. In some cells, fibers of fluorescence are observed, which resemble cytoskeleton. This may indicate cytoskeletal involvement in the inhibition of Flt23K.

Conclusions: : Flt23K seems to be trafficked in proteasome, lysosome, and cytoskeleton. This may relate to our previous findings of its intraceptor-induced unfolded protein response and apoptosis.

Keywords: vascular endothelial growth factor • inhibitory receptors 
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