Hypertrophic scars are serious derangements in healing after orofacial injury that are defined largely by the abnormal accumulation of extracellular matrix. The long-term objectives of this work are to determine why extracellular matrix accumulates in hypertrophic scars and to establish how such accumulation can be prevented. The project focuses on a ubiquitous component of human connective tissue matrix known as hyaluronate (hyaluronic acid, hyaluronan). Abnormal hyaluronate accumulation in scars is believed to occur because of a disequilibrium between hyaluronate production and breakdown. The cause is unknown; however, differences in the capacity of fibroblasts to bind, internalize and degrade hyaluronate may represent a key distinction between normal versus abnormal scar tissues.
The specific aims of this project are: (1) To visualize, by light and electron microscopic autoradiography, the binding and internalization of radiolabeled hyaluronate to human cutaneous fibroblasts derived from normal skin, normal scar and hypertrophic scar; (2) To establish that hyaluronate binding to fibroblasts is a receptor-mediated phenomenon and to determine whether binding affinity for hyaluronate differs between fibroblasts derived from normal versus abnormal tissues; and (3) To compare fibroblasts derived from normal skin, normal scar and hypertrophic scar with regard to the presence of CD44, a putative cell-surface receptor for hyaluronate, and to explore the involvement of CD44 and hyaluronate binding region protein (HABR) in the adhesion of hyaluronate to the fibroblastic cell surface. The methodology involves exposing fibroblasts to radioactively-labeled hyaluronate. The binding and internalization of the labeled hyaluronate will then be visualized directly through light and electron microscopic autoradiography. Detailed receptor binding studies will be conducted. Two molecules (CD44 and HABR) are promising candidates as cell-surface hyaluronate receptors. These will be studied in depth using immunocytochemical (single and double labeling), immunoelectron microscopic, and Western blotting procedures.
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