May 2008
Volume 49, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2008
Role of Intralysosomal Redox-Active Iron in Oxidative Stress-Induced Damage in Trabecular Meshwork Cells
Author Affiliations & Notes
  • Y. Lin
    Duke University, Durham, North Carolina
    Ophthalmology,
  • C. Luna
    Duke University, Durham, North Carolina
    Ophthalmology,
  • P. Gonzalez
    Duke University, Durham, North Carolina
    Ophthalmology,
  • K. Franz
    Duke University, Durham, North Carolina
    Chemistry,
  • D. L. Epstein
    Duke University, Durham, North Carolina
    Ophthalmology,
  • P. B. Liton
    Duke University, Durham, North Carolina
    Ophthalmology,
  • Footnotes
    Commercial Relationships  Y. Lin, None; C. Luna, None; P. Gonzalez, None; K. Franz, None; D.L. Epstein, None; P.B. Liton, None.
  • Footnotes
    Support  NIH Grants EY01894, EY016228, and EY05722; Research to Prevent Blindness
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 1622. doi:https://doi.org/
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      Y. Lin, C. Luna, P. Gonzalez, K. Franz, D. L. Epstein, P. B. Liton; Role of Intralysosomal Redox-Active Iron in Oxidative Stress-Induced Damage in Trabecular Meshwork Cells. Invest. Ophthalmol. Vis. Sci. 2008;49(13):1622. doi: https://doi.org/.

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

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Abstract

Purpose: : In vitro studies have demonstrated the accumulation of iron during aging in different tissues. This accumulation of iron is believed to contribute to the age-related oxidative damage and has been associated with several age-related diseases, including Alzheimer’s disease, Parkinson’s disease, type 2 diabetes, cardiovascular diseases, and macular degeneration. Here we investigated the potential role of intralysosomal redox-active iron in oxidative stress-induced damage in trabecular meshwork (TM) cells.

Methods: : Confluent cultures of porcine TM cells were used for these studies. Chronic oxidative stress was applied using the hyperoxic model (40% O2 versus physiological 5% O2 for two weeks); acute oxidative stress was applied by incubating the cells for three hours with H2O2 (250 µM, 500 µM, 750 µM, 1000 µM). The levels of mRNA for several iron-regulated genes were quantified by real-time PCR using specific primers. Protein levels of ferritin light chain (FTL) were analyzed by western-blot. Cytotoxicity was assayed with the MultiTox-Fluor Multiplex from Promega. Lysosomal labilization was evaluated by fluorometry using lysotracker red.

Results: : Exposure of porcine TM primary cultures to chronic oxidative stress increased the mRNA content of several genes involved in iron homeostasis, including transferrin receptor (TFRC, 1.85±0.25 fold), ferritin H (FTH, 2.01±0.16 fold), FTL (2.44±1.01), and metallothionein (MT1A, 6.09±2.00 fold). Similar to what has been reported in the TM from glaucoma donors, the mRNA of ceruloplasmin (CP, 0.47 ±0.1 fold) was downregulated with chronic oxidative stress. Western-blot analysis also demonstrated elevated levels of FTL suggesting an increase in the labile iron pool size in the stressed cultures. Pre-treatment of porcine TM cells with the intralysosomal iron chelator desferrioxamine (dfo) completely protected in a dosage-dependent manner against H2O2-induced cell death and H2O2-induced lysosomal permeabilization.

Conclusions: : Our results indicate that chronic exposure of TM cells to oxidative stress induces changes in the expression of iron-regulated genes, which suggests the intracellular accumulation of redox-active iron in the stressed cultures. Due to its potential of generating reactive oxygen species through Fenton reactions, the accumulation of iron in TM cells with age may play a role in the oxidative stress damage observed in the outflow pathway of POAG donors. Iron chelation therefore may represent a novel therapeutic approach to POAG.

Keywords: trabecular meshwork • oxidation/oxidative or free radical damage • aging 
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