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Tissue inhibitor of metalloproteinase-3 differentially binds to components of Bruch’s membrane
  1. M A Majid1,
  2. V A Smith1,
  3. F J Matthews1,
  4. A C Newby2,
  5. A D Dick1
  1. 1Academic Unit of Ophthalmology, University of Bristol, Bristol Eye Hospital, Bristol, UK
  2. 2Bristol Heart Institute, University of Bristol, Bristol Royal Infirmary, Bristol
  1. Correspondence to: V A Smith Bristol Eye Hospital, Lower Maudlin Street, Bristol BS1 2LX, UK;Val.Smith{at}bristol.ac.uk

Abstract

Background: Sorsby’s fundus dystrophy (SFD) is caused by mutations in tissue inhibitor of metalloproteinase (TIMP)-3 and, with the exception of early onset, is similar to age-related macular degeneration. The pathological features of this condition relate to the accumulation of TIMP-3 in Bruch’s membrane.

Aims: To compare the extracellular membrane-binding characteristics of wild-type and four SFD-mutant TIMP-3s.

Methods: COS-7 cells were transfected with wild-type, Ser-181, Gly-167, Ser-156 and Tyr-168 SFD-mutant TIMP-3 cDNA. The TIMP-3 proteins subsequently synthesised were harvested, analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, semiquantified by ELISA and used in binding assays on the basis of the retention of the wild-type and SFD-mutant TIMP-3 proteins by components of Bruch’s membrane.

Results: SFD-mutant TIMP-3s could not be distinguished from wild-type TIMP-3 by the extents to which they aggregated or adhered to type-I collagen, type-IV collagen, fibronectin, laminin, elastin, chondroitin sulphates A, B and C, and heparin sulphate. Of these macromolecules, the wild-type and SFD-mutant TIMP-3s exhibited greatest affinity for elastin and laminin.

Conclusion: The similarity in the physical and extracellular membrane-binding characteristics of wild-type and SFD-mutant TIMP-3s indicates that these properties are not responsible for the difference in timing of onset of SFD and age-related macular degeneration.

  • ARMD, age-related macular degeneration
  • DMEM, Dulbecco’s modified Eagle medium
  • ECM, extracellular membrane
  • FCS, fetal calf serum
  • GAG, glycosaminoglycan
  • HRP, horseradish peroxidase
  • MMP, matrix metalloproteinase
  • RPE, retinal pigment epithelial
  • SDS, sodium dodecyl sulphate
  • SFD, Sorsby’s fundus dystrophy
  • TIMP-3, tissue inhibitor of metalloproteinase-3
  • TMB, 3,3′,5,5′-tetramethyl benzidine

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Footnotes

  • Competing interests: None declared.

  • Published Online First 12 July 2006

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