TIMP-3 Exhibits High Affinity for Elastin

Abstract

Abstract: : Purpose: Sorsby's fundus dystrophy (SFD) and Age Related Macular Degeneration (ARMD) are retinal diseases characterised by the respective accumulation of mutant and wild-type TIMP-3 in Bruch's membrane. TIMP-3 is an MMP inhibitor and may therefore reduce the rate of extracellular matrix turnover and cause thickening of Bruch's membrane. Since this is an important factor in the pathological progression of these diseases the purpose of this study was to determine the binding characteristics of TIMP-3 to specific macromolecular components of Bruch's membrane. Methods: COS-7 cells were transduced with wild type and SFD mutant TIMP-3. The cellular matrices were subsequently harvested and Western blotting and ELISA were used to detect and quantify the secreted TIMP-3 proteins. Component proteins of Bruch's membrane were coated onto wells of 96 well plates and binding of wild type and mutant TIMP-3 to these proteins was measured by ELISA. Results: The TIMP-3 proteins showed little or no binding to types I and IV collagen, but moderate binding to Laminin, Fibronectin, Heparin Sulphate and Chondroitin Sulphate A, B & C. Affinity for Elastin was significantly higher. Observed differences in affinity of the wild type and mutant TIMP-3 proteins for each of these matrix components were small and not statistically significant. Conclusions: Wild type and SFD mutant TIMP-3 show high affinity for elastin. Binding of TIMP-3 to the elastic layer of Bruch's membrane could explain the late pathology of SFD and ARMD.

Keywords: Bruch's membrane • age-related macular degeneration • gene transfer/gene therapy