April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Inhibitory Effect of Cx43 on Oxidative Stress Induced VEGF in Human Retinal Pigment Epithelial Cells
Author Affiliations & Notes
  • C. M. Hutnik
    Ophthalmology, Ivey Eye Institute, London, Ontario, Canada
  • S. Khimdas
    Ophthalmology, Ivey Eye Institute, London, Ontario, Canada
  • H. Liu
    Ophthalmology, Ivey Eye Institute, London, Ontario, Canada
  • C. Pocrnich
    Ophthalmology, Ivey Eye Institute, London, Ontario, Canada
  • D. Laird
    Anatomy and Cell Biology, University of Western Ontario, London, Ontario, Canada
  • C. Shao
    Anatomy and Cell Biology, University of Western Ontario, London, Ontario, Canada
  • Footnotes
    Commercial Relationships  C.M. Hutnik, None; S. Khimdas, None; H. Liu, None; C. Pocrnich, None; D. Laird, None; C. Shao, None.
  • Footnotes
    Support  None.
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 1433. doi:
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      C. M. Hutnik, S. Khimdas, H. Liu, C. Pocrnich, D. Laird, C. Shao; Inhibitory Effect of Cx43 on Oxidative Stress Induced VEGF in Human Retinal Pigment Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1433.

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

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Abstract

Purpose: : To determine the effect of connexin43 (Cx43) expression and gap junctional intercellular communication (GJIC) on the expression and secretion of vascular endothelial growth factor (VEGF) from the retinal pigment epithelium under normal cell culture and oxidative stress conditions.

Methods: : Stable cell lines of ARPE-19 were produced in which Cx43 was either over-expressed, down-regulated by targeted shRNA, or functionally inhibited by co-expression of a disease-linked dominant-negative mutant (G21R). Pharmacologic blockade of GJIC was accomplished with flufenamic acid. Oxidant challenge was performed with tert-butyl hydroperoxide (tBH). VEGF gene expression and secretion were assessed by real-time PCR and ELISA, respectively. Serum was collected from wild-type mice, mice expressing a dominant-negative mutant of Cx43 and Cx43 null mice.

Results: : Over-expression of Cx43 in ARPE-19 cells reduced both gene expression and secretion of VEGF. Down-regulation of Cx43 increased gene expression and secretion of VEGF. Increased secretion of VEGF was also observed in ARPE-19 cells expressing a dominant negative mutant of Cx43 or when GJIC was blocked. Over-expression of Cx43 reduced tBH-induced secretion of VEGF from ARPE-19 cells. Finally, Cx43 mutant mice and Cx43 knock-out mice displayed relatively higher levels of serum VEGF than wild-type mice.

Conclusions: : Expression of Cx43 in ARPE-19 cells reduces VEGF secretion under normal cell culture conditions and protects against tBH-induced VEGF secretion.

Keywords: retinal pigment epithelium • oxidation/oxidative or free radical damage • gap junctions/coupling 
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