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We have recently reported that cell walls (CWs) of two closely related E. aerofaciens strains appear arthritogenic or nonarthritogenic when injected i.p. into the rats (Zhang et al. Rheumatology 2000). These strains have different structures of the CW peptidoglycan (PG). To further define what determines the arthritogenicity of these human intestinal bacteria, the tissue distribution of their CWs was compared. Muramic acid (MurNAc), a component of PG, was selected as a marker for bacterial CW as it is not synthesized by eukaryotic cells. Gas chromatography-mass spectrometry was applied to identify and quantify MurNAc. The results obtained indicate that the amount of MurNAc was much higher in the spleen and liver after injection of the arthritogenic CW than after injection of the nonarthritogenic CW. MurNAc was detected in synovial tissues and fluids from day 1 to day 28 after injection of the arthritogenic CW, but not after injection of the nonarthritogenic CW. This is probably due to the resistance of the arthritogenic CW against biodegradation; lysozyme and mutanolysin degraded the arthritogenic CW only by 21-34%, whereas the nonarthritogenic CW was degraded by 77-78%, both after 24 h incubation. Furthermore, degradation by mutanolysin significantly increased the capacity of the arthritogenic PG to stimulate rat macrophages to secrete TNF-α and MCP-1, whereas it dramatically decreased such a capacity of the nonarthritogenic PG, suggesting that peptides with proinflammatory activity are released from the arthritogenic PG. These results, obtained with an arthritogenic and nonarthritogenic strains of E. aerofaciens, indicate that capacity to resist biodegradation, leading to persistence in the tissues, and to release proinflammatory PG peptides, are crucial factors determining arthritogenicity or nonarthritogenicity of a bacterial CW.