Glucosamine represents one of the most commonly used over-the- counter drugs to treat osteoarthritis. However, the molecular mechanisms involved in the realization of its anti-arthritic activities are still poorly understood. Recently, we discovered that glucosamine (GlcN) and its derivative, N-acetylglucosamine (GlcNAc), express a broad range of anti-inflammatory activities in cultured human articular chondrocytes stimulated with interleukin-1beta (IL-1beta). These activities include inhibition of IL-beta induced nitric oxide (NO) production, IL-6 and COX-2 synthesis. In contrast, GlcNAc does not affect constitutively expressed COX-1 protein. These data have allowed us to generate a working hypothesis proposing that GlcN and its derivatives may beneficially affect the impaired cartilage metabolism in OA, which eventually could result in restoration of extracellular cartilage matrix and chondrocyte homeostasis. Therefore, the goal of the proposed project is to analyze the influence of GlcN and GlcNAc on homeostasis of normal and osteoarthritic human articular chondrocytes and to study the effect of these agents on quantitative and qualitative changes in extracellular matrix glycosaminoglycans. The aims of the study are: 1) to study the effect of GlcN and GlcNAc on production of glycosaminoglycans, including hyaluronic acid, chondroitin-6 sulfate, chondroitin-4 sulfate, dermatan sulfate, and keratan sulfate, in cultured human chondrocytes isolated from normal and osteoarthritic joints; 2) to investigate in vitro chondroprotective effect of GlcNAc on glycosaminoglycan degradation induced by IL-1beta; 3) to study the effect of GlcNAc on the activity of selected lysosomal glycosidases in cultured human articular chondrocytes stimulated with IL-1beta; 4) to correlate the biological activities of GlcN and GlcNAc with the age of chondrocyte donors; 5) to analyze the effect of G1cN and GlcNAc on chondrocyte apoptosis; 6) to examine the effect of G1cN and GlcNAc on TGFbeta and IGF-1 production in cultured human articular chondrocytes. Results of the study will broaden our understanding of the mechanisms of chondroprotection, and will serve as a starting platform for the identification of new molecular targets involved in the pathogenesis of osteoarthritis. The potential practical value of this project is that the obtained experimental data could be used as a basis for the design of a new class of the pharmacological agents capable of preventing or may be even restoring cartilage loss in the joints affected by arthritis.