Involvement of Hydrogen Peroxide in Collagen Cross-linking by High Glucose in Vitro and in Vivo

• Published in: The Journal of biological chemistry 1996-05, Vol.271 (22), p.12964-12971
• Main Authors: Elgawish, A, Glomb, M, Friedlander, M, Monnier, V M
• Format: Article
• Language: English
• Subjects: Animals; Antioxidants - chemistry; Collagen - chemistry; Cross-Linking Reagents - chemistry; Diabetes Mellitus, Experimental - chemically induced; Diabetes Mellitus, Experimental - metabolism; Female; Glucose - chemistry; Glucose - metabolism; Hydrogen Peroxide - chemistry; In Vitro Techniques; Kinetics; Maillard Reaction; Oxidative Stress; Rats; Rats, Sprague-Dawley; Streptozocin; Tendons - chemistry; Tendons - transplantation
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.271.22.12964

The Maillard reaction has been implicated in cross-linking and fluorescence formation of collagen exposed to high glucose in vitro . However, several pharmacologic agents, whose action seems unrelated to pathways of nonenzymatic glycation, have been demonstrated to prevent cross-linking in diabetes. To clarify this discrepancy, kinetic changes in glycation, glycoxidation (carboxymethyllysine, CML), and cross-linking (measured as tendon breaking time, TBT) were evaluated in rat tail tendons incubated in 5 and 30 m M glucose in vitro and in tendons implanted in vivo into diabetic rat peritoneal cavity. In vitro , rates were found to be both O 2 - and glucose-dependent. Tendon preglycation and presence of added 2 m M glycosylamine and Amadori compounds (Amadori product of glucose and propylamine) catalyzed these changes in a primarily O 2 -dependent manner. In the presence of Amadori compounds, kinetic changes were dramatically increased and were preventable by addition of catalase to the medium. Tendons implanted into diabetic rat peritoneum became more rapidly glycoxidized and cross-linked when implanted at day 30 from diabetes onset (high tissue glycation) compared to day 3 (low tissue glycation) in spite of similar glycation kinetics, suggesting a mechanistic dissociation between glycation, glycoxidation, and cross-linking in diabetes. Indeed, intraperitoneal injection of catalase and other antioxidants dramatically suppressed cross-linking, fluorescence formation, and, to some extent, glycoxidation, without affecting glycation. This study confirms the role of oxidative stress in protein cross-linking by the Maillard reaction in vitro and provides the first evidence for a role of H 2 O 2 in cross-linking in diabetes. Whereas Amadori products are a potent source of H 2 O 2 formation in vitro , their precise contribution to H 2 O 2 generation and the actual role of Maillard reaction products in collagen cross-linking in diabetes requires further investigation.