Depletion of reactive advanced glycation endproducts from diabetic uremic sera using a lysozyme-linked matrix.
Diabetic uremic sera contain excessive amounts of reactive advanced glycation endproducts (AGEs), which accelerate the vasculopathy of diabetes and end-stage renal disease. To capture in vivo-derived toxic AGEs, high affinity AGE-binding protein lysozyme (LZ) was linked to a Sepharose 4B matrix. Initial studies showed that > 80% of 125I-AGE-BSA was retained by the LZ matrix, compared with < 10% retained by a control matrix. More than 60% of AGE-lysine was captured by the LZ matrix, and the LZ-bound fraction retained immunoreactivity and cross-linking activity, but had little intrinsic fluorescence (370/440 nm). After passage through the LZ matrix, AGE levels in diabetic sera (0.37+/-0.04 U/mg) were significantly reduced to a level (0.09+/-0.01 U/mg; n = 10; P < 0. 0001) comparable with the level of normal human serum, whereas total protein absorption was < 3%. The AGE-enriched serum fraction exhibited cross-linking activity, which was completely prevented by aminoguanidine. Among numerous LZ-bound proteins in diabetic uremic sera, three major proteins "susceptible" to AGE modification were identified: the immunoglobulin G light chain, apolipoprotein J (clusterin/SP-40,40), and the complement 3b beta chain. These findings indicate that the LZ-linked AGE affinity column may serve as an efficient method for the depletion of toxic AGEs from sera, including specific AGE-modified proteins that may be linked to altered immunity, lipoprotein metabolism, and accelerated vasculopathy in renal failure patients with or without diabetes.
Mitsuhashi
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T
The Picower Institute for Medical Research, Manhasset, New York 11030, USA.
Li
Y M
YM
Fishbane
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Vlassara
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H
D000328:Adult; D000368:Aged; D000595:Amino Acid Sequence; D001798:Blood Proteins / Q000494:pharmacology; D003928:Diabetic Nephropathies / Q000097:blood*; D017127:Glycation End Products, Advanced / Q000097:blood* / Q000737:chemistry; D006801:Humans; D015151:Immunoblotting; D008875:Middle Aged; D008969:Molecular Sequence Data; D009113:Muramidase / Q000378:metabolism*; D012710:Serum Albumin, Bovine / Q000378:metabolism; D014511:Uremia / Q000097:blood
D001798:Blood Proteins; D017127:Glycation End Products, Advanced; D012710:Serum Albumin, Bovine; D009113:Muramidase