Formation of Nε-(carboxymethyl)lysine in inflammatory and noninflammatory conditions of nerve and muscle and in inflammatory cells in vitro

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We studied the occurrence of Nε-(carboxymethyl)lysine (CML), the major in vivo Maillard product in nerve and muscle in various inflammatory and noninflammatory diseases using a previously characterised specific CML antiserum. We found an increased CML staining in the perineurium of biopsies from B12-deficient, diabetic and vasculitic neuropathy and to a lesser extent, in alcoholic neuropathy and in mononuclear inflammatory cells. In diabetic neuropathy CML, the receptor for advanced glycation endproducts (RAGE) and nuclear factor kappa B (activated NFκB) were colocalised indicating possible interaction. In myositis, intracellular CML depositions were observed in infiltrating monocytes, myoblasts and necrotic fibers and Western blot analysis showed a marked increase of CML-modified proteins. To evaluate the impact of inflammation-associated cells on CML formation, we designed in vitro experiments using granulocytes and the monocytic cell line Mono-Mac6 (MM6). Oxidative burst of zymosan-stimulated granulocytes led to CML modification of extracellular proteins within 60–120 min. Predifferentiated MM6 cells, exerting NADPH oxidase activity but lacking myeloperoxidase activity, showed CML modification of distinct intracellular proteins within 1–3 days upon stimulation. Our findings suggest that CML modification occurs in vivo and in vitro when oxidative stress from e.g. inflammatory cells is present, indicating that CML may represent a marker for oxidative stress.