We have reported that Fn fragments (Fn-f), which have been detected in synovial fluids of osteoarthritis and rheumatoid arthritis patients, can potently cause cartilage chondrolysis and depress proteoglycan (PG) synthesis in cartilage tissue cultured as explants. Amino-terminal 29-kDa, gelatin-binding 50-kDa, and integrin-binding 140-kDa Fn-f are active. In order to investigate the mode of action and devise means of blocking the damage mediated by all Fn-f, we have tested the effects of various analogs resembling the integrin binding sequence, Arg-Gly-Asp-Ser, on blocking Fn-f-mediated chondrolysis. The analog peptides, Gly-Arg-Ala-Asp-Ser-Pro-Lys and Arg-Phe-Asp-Ser, at concentrations as low as 1 microM, blocked the effects of all three Fn-f on cartilage degradation, while the native sequence peptide, Arg-Gly-Asp-Ser, had very low Fn-f-blocking activity and by itself caused cartilage damage. Random sequence peptides dissimilar to the analog sequences were inactive as inhibitors as well as was a sequence analog, Phe-Asp-Arg-Ser, related to the Arg-Phe-Asp-Ser inhibitor. The analog inhibitory peptides decreased rates of Fn-f-mediated PG degradation and release from cartilage and decreased Fn-f-mediated PG synthesis depression. The analog inhibitory peptides alone had no detectable effect on cartilage PG degradation or PG synthesis rates. These data show that the chondrolytic activities of integrin-binding and nonbinding Fn-f can be blocked by synthetic peptide analogs of the Arg-Gly-Asp-Ser sequence and suggest that these peptides may be useful for blocking other activities of Fn-f.