An in vitro study on the role of metal catalyzed oxidation in glycation and crosslinking of collagen.
The present investigation was carried out to understand the effect of metal catalyzed oxidation on glycation and crosslinking of collagen. Tail tendons obtained from rats weighing 200-225 g were incubated with glucose (250 mM) and increasing concentrations of copper ions (5, 25, 50 and 100 microM) under physiological conditions of temperature and pH. Early glycation, crosslinking and late glycation (fluorescence) of collagen samples were analyzed periodically. Early glycation was estimated by phenol sulfuric acid method, and the crosslinking was assessed by pepsin and cyanogen bromide digestion. A concentration-dependent effect of metal ions on the rate of glycation and crosslinking of collagen was observed. Tendon collagen incubated with glucose and 100 microM copper ions showed 80% reduction in pepsin digestion within seven days, indicating extensive crosslinking, whereas collagen incubated with glucose alone for the same period showed only 7% reduction. The presence of metal ions in the incubation medium accelerated the development of Maillard reaction fluorescence on collagen, and the increase was dependent on the concentration of metal ions used. The metal chelator diethylene triamine penta-acetate significantly prevented the increase in collagen crosslinking by glucose and copper ions. Free radical scavengers benzoate and mannitol effectively prevented the increased crosslinking and browning of collagen by glucose. The results indicate that the metal catalyzed oxidation reactions play a major role in the crosslinking of collagen by glucose. It is also suggested that the prevention of increased oxidative stress in diabetes may prevent the accelerated advanced glycation and crosslinking of collagen.
Sajithlal
G B
GB
Department of Biochemistry, Central Leather Research Institute, Adyar, Chennai, India.
Chithra
P
P
Chandrakasan
G
G
D000818:Animals; D002384:Catalysis; D003094:Collagen / Q000737:chemistry / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D016166:Free Radical Scavengers / Q000494:pharmacology; D005947:Glucose / Q000378:metabolism*; D017127:Glycation End Products, Advanced; D017665:Hydroxyl Radical; D008297:Male; D008670:Metals / Q000378:metabolism*; D010084:Oxidation-Reduction; D010449:Peptide Mapping; D051381:Rats; D017208:Rats, Wistar
D016166:Free Radical Scavengers; D017127:Glycation End Products, Advanced; D008670:Metals; D017665:Hydroxyl Radical; D003094:Collagen; D005947:Glucose