Abstract
Purpose: :
To determine whether kynurenines and lens pigments irradiated by UVA-light induce oxidation of ascorbic acid (ASC) and initiate the formation of advanced glycation end products (AGEs) in lens proteins
Methods: :
ASC oxidation by UVA photolysis (100 mW/cm2) of kynurenines or enzyme digested water insoluble proteins was assessed by spectrophotometry and HPLC. The irradiated and non-irradiated samples were incubated with human lens proteins or amino acids at pH 7.4 and 37oC to determine formation of AGEs. AGEs were quantified by HPLC.
Results: :
UVA photolysis of mixtures containing kynurenine (N-formyl kynurenine or kynurenine or 3OH-Kynurenine) and ASC led to the formation of DHA, formation of which was marginally affected by glutathione. ASC oxidation was also apparent in the UVA irradiated water-insoluble protein digest and it increased with time of irradiation (up to 2 hrs). ASC oxidation was 4-fold higher with cataractous lens protein digests than with age-matched normal lens digests; the percent oxidation was 12.62±4.2 (mean±SEM) for normal and 49.12±7.35 for cataractous lens protein digests (p<0.05). Incubation of UVA-irradiated kynurenine-modified lens proteins or UV-irradiated mixtures of ASC and kynurenines with Nα-BOC-lysine and Nα-BOC-arginine for 3 days at 37oC resulted in the formation of pentosidine, an AGE present in the human lens. These data show that UVA photolysis of kynurenines and lens pigments leads to oxidation of ASC, which results in AGE formation in lens proteins.
Conclusions: :
This study uncovers a novel pathway of AGE formation in the lens, which involves kynurenine and lens pigment-mediated oxidation of ASC.
Keywords: cataract • radiation damage: light/UV • protein modifications-post translational