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Glutathione and Ascorbate: The balance between oxidant and carbonyl stress in the aging human lens
Author(s) -
MONNIER VM,
FAN XJ
Publication year - 2013
Publication title -
acta ophthalmologica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.534
H-Index - 87
eISSN - 1755-3768
pISSN - 1755-375X
DOI - 10.1111/j.1755-3768.2013.3761.x
Subject(s) - crystallin , glycation , glutathione , chemistry , in vitro , incubation , disulfide bond , oxidative stress , lens (geology) , biochemistry , biophysics , protein aggregation , biology , enzyme , receptor , paleontology
Background: Two major processes appear to be at play in formation of ARNC. On the one hand advanced glycation end products (AGEs) modify lens crystallins, increasing thereby their susceptibility to oxidation via unfolding, exposure of SH‐groups and formation of UV‐photosensitizers, on the other hand SH groups form disulfides and high molecular weight aggregates that scatter light. In order to find out if there are critical SH residues in the latter process, we have generated a glutathione knockout(LEGSKO) mouse and an in vitro model of crystallin aggregation upon oxidation with H2O2.Methods: Crystallins from in vitro incubation or LEGSKO mouse were analyzed using ICAT labeling methods and 2D gel electrophoresis (2D‐PAGE) for disulfide analysis. Incubation of mouse lens homogenate (LH) with 5 mM H2O2 led to opacification within a few hours. Results: Analysis of LEGSKO crystallins by 2D‐PAGE revealed an age‐related shift from mostly intramolecular to intermolecular disulfide crosslinks which mimicked the pattern observed at 20 months in wild type lenses. All oxidation‐related disulfide forming sites in 13 crystallins were identified in vitro and found quasi concordant with those forming in the LEGSKO lens.