May 2003
Volume 44, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2003
High Glucose Suppresses Bovine Retinal Pericyte Proliferation: Implications for Retinopathy
Author Affiliations & Notes
  • P.P. Connell
    Ophthalmology Research Registrar, Master Misercordiae Hosp, Dublin, Ireland
  • G. Wei
    Biotechnology, Dublin City University, Dublin, Ireland
  • C.J. O' Brien
    Ophthalmology, Master Misercordiae Hosp, Dublin, Ireland
  • P. Cahill
    Vascular Health Research Centre, Dublin City University, Dublin, Ireland
  • Footnotes
    Commercial Relationships  P.P. Connell, None; G. Wei, None; C.J. O' Brien, None; P. Cahill, None.
  • Footnotes
    Support  1) Pharmacia ICO Fellowship(Ireland) 2) Phil Vizzard Diabetes Federation Ireland
Investigative Ophthalmology & Visual Science May 2003, Vol.44, 3886. doi:
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      P.P. Connell, G. Wei, C.J. O' Brien, P. Cahill; High Glucose Suppresses Bovine Retinal Pericyte Proliferation: Implications for Retinopathy . Invest. Ophthalmol. Vis. Sci. 2003;44(13):3886.

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

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Abstract

Abstract: : Purpose: Local retinal haemodynamics are altered in diabetic retinopathy.High glucose is integral in initiating many of these alterations, which change blood flow patterns. We have examined the effect of high glucose on bovine retinal pericyte cells and endothelial cells with respect to these autoregulatory elements. Methods: Pericyte cells were exposed to 25mM glucose and examined for changes in proliferating cell nuclear antigen (pCNA), Bax and BCl-2 protein expression and levels of caspase-3 activity. Clonal proliferation experiments, ethidium bromide/acridine orange dual stain for apoptosis were performed. Bovine retinal endothelial cells were exposed to increasing concentrations of glucose (maximum 25mM) acutely (3 hrs) and chronically (24 hrs). Constitutive endothelial nitric oxide synthase (eEnos), phosphorylated endothelial nitric oxide synthase activity (ppEnos) and cycloxygenase 1 (Cox-1) protein expression were examined. Media samples were assayed for Nitric Oxide activity with the DAN assay, and prostaglandin F1-alpha (PGF1-alpha) as an indicator of prostacyclin synthesis (PGI2). Results: Pericytes exhibited increasing apoptosis via acridine orange/ethidium bromide dual stain. The effect became manifest at 48 hrs and increased up to 7 days (Fig 1). Parallel clonal proliferation experiments revealed decreased cell counts with 25 mM glucose manifesting at 36 hrs. (Fig 2). PCNA protein expression at 72 hrs was reduced in the glucose treated cells (Fig 3). Bax protein expression increased. Caspase activity increased. Enos protein expression in BRECs decreased in a time and dose dependent manner, with maximal eNos suppression at 20 hrs, peaking at 25mM glucose treatments (Fig 4). NO activity decreased concordantly (Fig 5). Pre-treatment of BRECs with known activators of eNOS stimulated ppEnos acutely, liberating NO (Fig 6).Co-incubating with glucose abolished this effect. PGI was reduced but not significantly s in glucose treated cells. Cox 1 protein expression remained unchanged. Conclusions: We have shown an increase in apoptosis with high glucose with decrease in eNOS expression. Co-cultures should further our understanding of their complex interactions to further define the pathogenesis of diabetic retinopathy.

Keywords: diabetes • diabetic retinopathy • retinal culture 
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