The enzymatic processes underlying the degradation of aggrecan in cartilage and the corresponding changes in the biomechanical properties of the tissue are an important part of the pathophysiology of osteoarthritis. Recent studies have demonstrated that the hexosamines glucosamine (GlcN) and mannosamine (ManN) can inhibit aggrecanase-mediated cleavage of aggrecan in IL-1-treated cartilage cultures. The term aggrecanase describes two or more members of the ADAMTS family of metalloproteinases whose glutamyl endopeptidase activity is known to be responsible for much of the aggrecan degradation seen in human arthritides. In this study we examined the effect of ManN and GlcN on aggrecanase-mediated degradation of aggrecan induced by IL-1α and the corresponding tissue mechanical properties in newborn bovine articular cartilage. After 6 days of culture in 10 ng/ml IL-1 plus ManN, mechanical testing of explants in confined compression demonstrated that ManN inhibited the IL-1α-induced degradation in tissue equilibrium modulus, dynamic stiffness, streaming potential, and hydraulic permeability, in a dose-dependent fashion, with peak inhibition (∼75–100% inhibition) reached by a concentration of 1.35 mM. Aggrecan from explants cultured in IL-1 was found by Western analysis to be almost entirely processed down to the G1-NITEGE³⁷³ end product. Addition of ManN or GlcN was found to produce 75–90% inhibition of this cleavage, but the proportion of aggrecan remaining in the tissue which was cleaved at aggrecanase sites in the chondroitin sulfate (CS)-rich region (Glu¹⁵⁰¹ and Glu¹⁶⁸⁷) was higher than with IL-1 alone. This result suggests that the preservation of mechanical properties by hexosamines in explants is primarily due to inhibition of cleavage at the Glu³⁷³ site in the interglobular domain. While the precise mechanism by which hexosamines function in this system is unclear, the present analysis suggests that the mechanical properties examined may be predominantly a function of electrostatic repulsion due to the charged CS chains in the tightly packed repetitive sequences of the CS-1 region.