April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Increased Autophagy is Associated With Transient Recovery of High Glucose-Induced Mitochondrial Fragmentation in Retinal Endothelial Cells
Author Affiliations & Notes
  • K. Trudeau
    Medicine & Ophthalmology,
    Boston University School of Medicine, Boston, Massachusetts
  • A. Molina
    Medicine,
    Boston University School of Medicine, Boston, Massachusetts
  • G. Las
    Medicine,
    Boston University School of Medicine, Boston, Massachusetts
  • O. Shirihai
    Medicine,
    Boston University School of Medicine, Boston, Massachusetts
  • S. Roy
    Medicine & Ophthalmology,
    Boston University School of Medicine, Boston, Massachusetts
  • Footnotes
    Commercial Relationships  K. Trudeau, None; A. Molina, None; G. Las, None; O. Shirihai, None; S. Roy, None.
  • Footnotes
    Support  NEI, NIH EY018218, Massachusetts Lions Organization, Fight for Sight, Undergraduate Research Opportunities Program at Boston University
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 5635. doi:
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    • Get Citation

      K. Trudeau, A. Molina, G. Las, O. Shirihai, S. Roy; Increased Autophagy is Associated With Transient Recovery of High Glucose-Induced Mitochondrial Fragmentation in Retinal Endothelial Cells. Invest. Ophthalmol. Vis. Sci. 2010;51(13):5635.

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

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Abstract

Introduction: : To investigate whether recovery from high glucose (HG)-induced mitochondrial fragmentation is associated with altered autophagy in rat retinal endothelial cells (RRECs), we assessed mitochondria shape change and autophagy induction after acute (24hours) exposure to HG.

Methods: : To assess mitochondrial shape, RRECs were grown in normal (5mM) and HG (30mM) medium for 0, 15, 30, 60, 120 minutes and 6 days. Cells were stained with (8nM) tetramethylrhodamine-ethyl-ester-perchlorate and imaged live using confocal microscopy. Average mitochondria shape within a cell was determined using Form Factor (FF) and Aspect Ratio (AR) values of the mitochondria, to assess branching and length. To analyze HG-induced formation and turnover of autophagosomes in RRECs, expression of LC3-II, a marker for autophagosomes, and p62, a marker for autophagic protein degradation, were examined by Western Blot analysis at 1, 2, 6, 48 hours and 6 days of HG.

Results: : Acute exposure of RRECs to HG media resulted in transient fragmentation of mitochondria within 30 minutes (0 min: FF=2.38, AR=2.11; 30min: FF=1.519, p=0.001; AR=1.89, p=0.009), followed by partial morphology recovery after 120 minutes (FF=2.01, p<0.001, AR=2.06, p=0.004). LC3-II expression was increased after 2 hours of HG (180±58% of normal, p=0.004) and p62 expression was reduced after 6 hours of HG (64±31% of normal, p=0.025). After 24 hours of HG mitochondrial fragmentation was observed again and remained for the rest of the 6 day experiment (FF=1.377, p<0.001, AR=1.793, p=0.006). LC3-II and p62 expression showed no change after 6 days of HG exposure.

Conclusions: : Findings from this study show that transient recovery from HG-induced mitochondrial fragmentation occurs with changes in autophagy-related proteins LC3-II and p62. Permanent mitochondrial fragmentation persists after 24 hours of HG with no change in LC3-II and p62. Together these observations suggest that autophagy may play a role in the recovery from HG-induced mitochondrial fragmentation. This could shed light on how HG contributes to mitochondrial dysfunction associated with diabetic retinopathy.

Keywords: mitochondria • diabetic retinopathy • apoptosis/cell death 
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