March 2012
Volume 53, Issue 14
Free
ARVO Annual Meeting Abstract  |   March 2012
Effect Of Divalent Metal Chelation On The MMP System And Transport Characteristics Of Human Bruch’s Membrane
Author Affiliations & Notes
  • Yunhee Lee
    Ophthalmology, King's College London, Lambeth Palace Rd, London, United Kingdom
  • Ali Hussain
    Division of Molecular Therapy, UCL Institute of Ophthalmology, London, United Kingdom
  • John Marshall
    Division of Molecular Therapy, UCL Institute of Ophthalmology, London, United Kingdom
  • Footnotes
    Commercial Relationships  Yunhee Lee, None; Ali Hussain, None; John Marshall, None
  • Footnotes
    Support  Fight For Sight, UK
Investigative Ophthalmology & Visual Science March 2012, Vol.53, 6500. doi:
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      Yunhee Lee, Ali Hussain, John Marshall; Effect Of Divalent Metal Chelation On The MMP System And Transport Characteristics Of Human Bruch’s Membrane. Invest. Ophthalmol. Vis. Sci. 2012;53(14):6500.

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Abstract

Purpose: : Divalent metal deposition in Bruch’s of elderly donors and those with age-related macular degeneration (AMD) may serve to trap and stabilise both lipid and proteinaceous debris and by enhancing free radical production, increase the degree of collagen cross-links within the membrane. These changes decrease the porosity of the membrane resulting in a decrease in its transportation capability. This study has examined the effects of metal chelation on the components of the matrix metalloproteinase (MMP) system and on the hydraulic conductivity of Bruch’s membrane.

Methods: : Intact Bruch’s-choroid preparations were obtained from two donors aged 79 and 83 years and incubated with (a) phosphate buffered saline (PBS, control) and (b)10mM EGTA for 1 hour. Tissue samples wereseparated from the incubation medium and both fractions examined for MMP activity by gelatin zymography. Another 20 donor Bruch’s-choroid preparations (age range 58-93 years) were mounted in open-type Ussing chambers and perfused with PBS at a pressure of 2000Pa to determine basal hydraulic conductivity over a period of 8 hours. In seven preparations, at the 8-hour mark, the perfusion medium was switched to 10mM EGTA and effects on hydraulic conductivity monitored.

Results: : Metal chelation with EGTA showed enhanced release of active MMP-2, a constitutive enzyme involved in the maintenance of the membrane. Perfusion with PBS removed nearly all free and mobile components of Bruch’s within 8 hours. Samples perfused with PBS showed stable hydraulic conductivities of 1.14 ±0.4m/s/Pa over a perfusion period of 12 hours. In 7 preparations, switching to 10mM EGTA at the 8-hour mark, resulted in improved hydraulic conductivities over the next three 1-hour sampling times. Thus after 1, 2 and 3 hours, conductivities were increased to 2.67 ±1.3 (p<0.05), 3.62 ±1.9 (p<0.05), and 8.19 ± 2.86 (p<0.005) Mean ± SD respectively.

Conclusions: : Divalent metal chelation releases bound active MMP2 and considerably improves the hydraulic conductivity of Bruch’s membrane. Thus metal chelation appears to provide a new therapeutic avenue to address the degenerative changes associated with AMD.

Keywords: Bruch's membrane • extracellular matrix • age-related macular degeneration 
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