May 2007
Volume 48, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2007
The Angiopoietin/Tie-2 System Regulates the Actin Cytoskeleton in Retinal Pericytes
Author Affiliations & Notes
  • J. Cai
    Ophthalmology & Visual Sciences, The University of Texas Medical Branch, Galveston, Texas
  • M. E. Boulton
    Ophthalmology & Visual Sciences, The University of Texas Medical Branch, Galveston, Texas
  • P. Hykin
    Ophthalmology & Visual Sciences, Moorfields Eye Hospital, London, United Kingdom
  • Footnotes
    Commercial Relationships J. Cai, None; M.E. Boulton, None; P. Hykin, None.
  • Footnotes
    Support Diabetes UK
Investigative Ophthalmology & Visual Science May 2007, Vol.48, 1737. doi:https://doi.org/
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      J. Cai, M. E. Boulton, P. Hykin; The Angiopoietin/Tie-2 System Regulates the Actin Cytoskeleton in Retinal Pericytes. Invest. Ophthalmol. Vis. Sci. 2007;48(13):1737. doi: https://doi.org/.

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

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Abstract

Purpose:: We have previously reported that the angiopoietin/Tie-2 system plays an important role in recruitment and maturation of pericytes in diabetic retinopathy (Hykin et al, ARVO 2004). The aim of this study was to investigate the effect of angiopietin-1 (Ang-1) and -2 (Ang-2) on Tie-2 expression and phosphorylation and cytoskeletal organization in retinal pericytes.

Methods:: Pericytes were treated with angiopoietin-1 or -2 (100 ng/ml) for up to 48 hours. Tie-2 receptor density was determined by flow cytometric analysis. To assess Tie-2 phosphorylation retinal pericytes were treated with angiopoietins and phosphorylated Tie-2 was immunoprecipitated from the cell lysate by anti-tyrosine phosphorylation monoclonal antibody, followed by ELISA for Tie-2. To observe angiopoietin-induced changes in the cytoskeleton, retinal pericytes were grown on slides precoated with Matrigel and the actin cytoskeleton labeled with phalloidin conjugated with TRITC.

Results:: Flow cytometry demonstrated that, compared to normal controls, angiopoietin-treated pericytes exhibited an increase in levels of Tie-2 of 79.3 and 38.6% for Ang-1 and Ang-2 respectively. Ang-1 induced tyrosine phosphorylation of Tie-2 in pericytes within 5 minutes, reaching a maximum level at 1 hour, followed by a slight down-regulation. In contrast, the kinetics of Tie2 phosphorylation induced by Ang-2 was a gradual up-regulation with maximal phosphorylation occurring at 48 hours. Control pericytes were flat with thin, uniform, parallel actin filaments (stress fibres) departing from single foci and extending throughout the length of the cell, with the presence of lamellipodia. There were few dominant, elongated pseudopodia and few actin containing fine cell extensions. Ang-1 treatment caused dramatic reorganization of the actin cytoskeleton that resulted in a reduction in number of stress fibres and lamellipodia, with an uneven thickening of the remaining stress fibres and an increase in both dominant leading-edge pseudopodia observed at the ends of the cells and in invadopodia. By contrast, Ang-2-treated retinal pericytes remained flat with the partial disassociation of actin into aggregates. Tie-2 antisense treatment abolished both the effects of Ang-1 and Ang-2 on changes in actin cytoskeleton in the retinal pericytes.

Conclusions:: Tie-2 expression in retinal pericytes is functional and angiopoietin-1 may exert migratory effects on the pericytes via alterations in the F-actin cytoskeletal organization of the pericytes. This may be important in the recruitment of pericytes to new vessels in diabetic retinopathy.

Keywords: growth factors/growth factor receptors • diabetic retinopathy • cytoskeleton 
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