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A. Kumar, A. El-Osta, A. A. Hussain, J. Marshall; Sequestration of MMPs 2&9 by Ageing Human Bruch’s Membrane: Relevance to Functional Deterioration. Invest. Ophthalmol. Vis. Sci. 2009;50(13):3403.
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© ARVO (1962-2015); The Authors (2016-present)
To determine the level of binding and sequestration of matrix metalloproteinase (MMP) enzymes by ageing human Bruch’s membrane.
Freshly isolated Bruch’s-choroid preparations (14 eyes, age range 21-88 years) were mounted in Ussing chambers and eluted with phosphate buffered saline (PBS) at a hydrostatic pressure of 200mm H2O. The ensuing fluid transported across the preparation was collected at timed intervals over a 6-8 hour period and the amount of released MMP species quantified by gelatin zymography and densitometry. At the end of the experiment, the remaining bound/trapped MMP pool was extracted with SDS-sample buffer and quantified.
The releasable pool of MMPs 2&9 was routinely removed from the preparation within 6 hours of elution. The bound pool was observed to increase exponentially with ageing of donor, the increase in MMP-9 being considerably marked in comparison to MMP-2. Furthermore, increased amount of polymeric forms of MMPs 2&9 (indicative of oxidative damage and cross-linking) were also observed as a function of ageing of Bruch’s.
Greater sequestration of MMPs with age is expected to diminish the free pool available for activation, resulting in reduced matrix turnover and deterioration in the structural and functional aspects of the membrane.
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