Abstract
Purpose :
Advanced glycation end products (AGEs)-induced inflammation is involved in the pathogenesis of age related macular degeneration (AMD).To investigate biochemical and structural effect of FAOD in aged retina by measuring autofluorescence (AF), size and spectral changes of Drüsen. Defining fluorescent and non-fluorescent AGEs substrates of FAOD.
Methods :
After AGE modification of porcine retinas (n=20) with 25-mM glycolaldehyde, retinas were treated with FAOD (3.84 U/mL) or inactive mutant enzyme. AGEs were quantified by Maillard-type AF measurements (excitation 365 nm, emission 390–700 nm) by microspectrometry. Sections of three donor eyes with AMD (Biobank Antwerp, Belgium, ID: BE 71030031000) were treated with FAOD or PBS. Biochemically selective visualizations of Drüsen were compared by Fourier transform (FT)-Near Infrared (NIR) transmission microspectra (12,000 to 4000 cm−1). Spectral data were analyzed by principal component and partial least squares-discriminant analysis. Mixtures containing AGEs were digested with FAOD (3.83 U/L) and subjected to gel permeation chromatography, ultra-high performance liquid chromatography, high-resolution mass spectrometry (HR-MS) and tandem MS fragmentation.
Results :
Glycated porcine retinas showed a significant reduction in AF (43% ± 4%, P=0.001) following FAOD treatment but not FAOD mutant enzyme. FAOD treatment of sections of donor eyes with AMD reduced mean size of Drusen to 45 ± 21 % by NIR microspectroscopy (P<0.005). Hotelling’s plot showed a clear distinction between control- and FAOD treated Drüsen. Almost complete disappearance of Arginine-derived AGEs ornithine, fructosylarginine/glucosylarginine, and imidazolone A as well as lysine-based AGEs fructosyllysine, carboxyethyllysine (CEL) and carboxymethyllysine (CML) from AGE mixtures was observed by chromatography and MS.
Conclusions :
On both human and porcine glycolaldehyde-treated retinas, FAOD reduces AF caused by AGEs. On sections of AMD patients, size of drusen decreases after FAOD treatment. FAOD treatment results in breakdown of AGEs, shown by UV fluorescence, NIR microspectroscopy and gel permeation chromatography. Enzymatic digestion of common AGEs reveals a broad spectrum of substrates for FAOD. Deglycation by FAOD is worthwhile exploring as a novel treatment in GA patients.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.