Increased sequestration of matrix metalloproteinases in ageing human Bruch's membrane: implications for ECM turnover.
OBJECTIVE
The ageing of Bruch's membrane is associated with progressive reduction in the degradation of the capacity for ECM turnover mediated by the matrix metalloproteinase (MMP) system. In this study, the free and bound pools of all gelatinase species were quantified to aid in assessing the likelihood of reduced availability of pro-MMPs for activation in ageing Bruch's membrane.


METHODS
Bruch's membrane from macular locations (10 eyes; donor age range, 21-84 years) was mounted in Ussing chambers and eluted with phosphate-buffered saline to release the free pool of MMPs. Free and bound pools of MMPs were subjected to gelatin zymography, and individual gelatinase species were quantified by densitometric scans.


RESULTS
The zymograms displayed six gelatinase species: four corresponding to the pro- and active forms of MMP-2 and -9 and two high-molecular-weight polymeric forms designated HMW1 and -2, corresponding to approximate molecular masses of 195 and 391 kDa, respectively. The ageing of Bruch's membrane was associated with an exponential increase in the percentage of pro-MMPs bound to the membrane (pro-MMP-2: %age bound = 0.54 exp(0.04 x age), r = 0.87, P < 0.01; and pro-MMP-9: %age bound = 5.0 exp(0.03 x age), r = 0.8, P < 0.01). A similar exponential increase was seen in the percentage of bound HMW1 species (%bound = 11.7 exp(0.018 x age; P < 0.05). The HMW2 species was virtually all bound to the membrane, but some release was observed in the very elderly.


CONCLUSIONS
The ageing of Bruch's membrane was associated with progressive sequestration of MMPs reducing the free concentration and potential for activation. These changes may underlie the reduction in degradation that leads to the age-related increase in the thickness of the membrane.
Kumar
Anupma
A
Department of Ophthalmology, The Rayne Institute, St. Thomas' Hospital, London, United Kingdom. anupmakumar@hotmail.com
El-Osta
Austen
A
Hussain
Ali A
AA
Marshall
John
J
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000375:Aging / Q000502:physiology*; D016570:Bruch Membrane / Q000201:enzymology*; D003720:Densitometry; D005109:Extracellular Matrix / Q000378:metabolism*; D005260:Female; D006801:Humans; D008297:Male; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism*; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D008875:Middle Aged; D055815:Young Adult
D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9