Immunochemical Detection of Advanced Glycation End Products in Lens Crystallins From Streptozocin-Induced Diabetic Rat

Abstract

To reassess the significance of AGEs in cataract formation in diabetic animals, we measured amounts of AGEs in lens crystallins from STZ-induced diabetic animals with a newly developed ELISA. Lenses were removed at 5 and 20 wk after STZ injection. In 20-wk diabetic rats, all lenses were cataractous but not in control rats. In 20-wk diabetic compared with control rats, significant increases were observed in AGEs (172.3 ± 18.3 vs. 14.3 ± 1.7 All, P < 0.01) and fluorescence (2.04 ± 0.22 vs. 1.27 ± 0.10 AU, P < 0.05). The amounts of AGEs in lens crystallins, measured by the ELISA, were > 12-fold higher in diabetic rats. In agreement with earlier studies, we found that fluorescence in lens crystallins increased by 61% in diabetic rats. In 5-wk diabetic rats, all lenses were noncataractous. In 5-wk diabetic compared with control rats, significant increases were observed in AGEs (84.1 ± 7.7 vs. 9.4 ± 1.5 AU, P < 0.01) and fluorescence (1.45 ± 0.06 vs. 1.05 ± 0.06 AU, P < 0.01). Analysis of the AGE content by ELISA showed that accumulation of AGEs in diabetic lens crystallins does markedly occur with time, and a large amount of AGEs exists in the diabetic (cataractous) lens crystallins. The disproportionate elevation of AGEs, measured by the ELISA, compared with fluorescence suggests that the actual levels of AGEs in cataractous lens crystallins from diabetic animals are higher than previously anticipated, and nonfluorescent AGEs may exist in diabetic lens crystallins. With this premise, our data suggest that a newly developed ELISA for the detection of AGEs in tissue proteins may be a powerful tool for investigating the role of the advanced Maillard reaction in complications of diabetes and aging.