The focus of the proposed experiments centers on the use of monoclonal antibodies which react with unique carbohydrate moieties and provide a high-resolution analysis system of extracellular matrix moieties in human skin. The central hypothesis is that proteoglycans provide unique water- structuring and visco-elastic properties of the extracellular matrix and that subtle changes in these molecules can have profound effects on the properties of human skin, especially those associated with aging. The availability of unique monoclonal antibodies to specific carbohydrate epitopes on proteoglycans allows the identification of a distinct fingerprint for various layers of skin as a function of human age. Preliminary results establish certain monoclonal antibodies to be unique to specific skin layers and to subtly change as a function of age. We propose to build on these preliminary observations to establish a normative data base for skin samples as a function of age. In addition, by using these monoclonal antibodies, we propose to isolate and characterize the epitope- containing macromolecules from human skin specimens, in this case, proteoglycans. Proteoglycans will be isolated and epitope-reactive molecules will be obtained, and these macromolecules characterized. This project draws together technologies ranging from immunocytochemistry to detailed proteoglycan biochemistry and will provide basic information about changes in skin extracellular matrices as a function of human age.
Specific Aims : 1. Identify monoclonal antibodies which react with carbohydrate moieties, which change in discrete layers of human skin as a function of age (focus on chondroitin and keratan sulfate epitopes). 2. Use these antibodies which are specific to discrete layers of skin to isolate and/or characterize the macromolecules which contain these epitopes. 3. Establish new, innovative techniques to characterize the detailed microchemistry of proteoglycans relevant to skin aging.
