We propose to investigate the hypothesis that the chronic tissue damage associated with long-term diabetes mellitus may arise in part from in situ immune complex formation by accumulated immunoglobulins and/or antigens covalently bound to long-lived structural proteins which have undergone excessive nonenzymatic glycosylation with subsequent generation of reactive carbonyls. In the diabetic kidney, such immobilized immune complexes may serve as a chronic stimulus for tissue destruction by macrophages.
The specific aims of the studies described in this proposal are to: I) Characterize the covalent trapping of lysine and proteins by nonenzymatically glycosylated collagen (chemical and kinetic studies), using soluble collagen immobilized on Agarose. The effect of time and reactant concentrations will be evaluated, and binding will be assessed both in vitro and in vivo; II) Quantitate in situ formation of immune complexes by antigen or antibody bound covalently to nonenzymatically glycosylated collagen, using radioiodinated soluble proteins; II) Investigate the effect of immune complexes bound to glycosylated collagen on macrophage tissue-injury mechanisms. Phagocytosis will be evaluated using antibody-sensitized sheep erythrocytes labelled with 51Cr, and hydrolytic enzyme secretion will be investigated by measuring selected glycohydrolases, neutral proteases, and plasminogen activator; IV) Evaluate the effect of diabetes and high in vitro glucose concentration on macrophage response to nonenzymatically glycosylated collagen-bound immune complexes. Results from these studies may provide a basis for future development of new therapeutic agents useful in the treatment of diabetic complications.
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