Serum Levels of Advanced Glycation Endproducts and Other Markers of Protein Damage in Early Diabetic Nephropathy in Type 1 Diabetes

Division of Endocrinology, University of Toronto, Toronto, Canada.
PLoS ONE (Impact Factor: 3.23). 04/2012; 7(4):e35655. DOI: 10.1371/journal.pone.0035655
Source: PubMed


To determine the role of markers of plasma protein damage by glycation, oxidation and nitration in microalbuminuria onset or subsequent decline of glomerular filtration rate (termed "early GFR decline") in patients with type 1 diabetes.
From the 1(st) Joslin Kidney Study, we selected 30 patients with longstanding normoalbuminuria and 55 patients with new onset microalbuminuria. Patients with microalbuminuria had 8-12 years follow-up during which 33 had stable GFR and 22 early GFR decline. Mean baseline GFR(CYSTATIN C) was similar between the three groups. Glycation, oxidation and nitration markers were measured in protein and ultrafiltrate at baseline by liquid chromatography-tandem mass spectrometry using the most reliable methods currently available.
Though none were significantly different between patients with microalbuminuria with stable or early GFR decline, levels of 6 protein damage adduct residues of plasma protein and 4 related free adducts of plasma ultrafiltrate were significantly different in patients with microalbuminuria compared to normoalbuminuria controls. Three protein damage adduct residues were decreased and 3 increased in microalbuminuria while 3 free adducts were decreased and one increased in microalbuminuria. The most profound differences were of N-formylkynurenine (NFK) protein adduct residue and N(ω)-carboxymethylarginine (CMA) free adduct in which levels were markedly lower in microalbuminuria (P<0.001 for both).
Complex processes influence levels of plasma protein damage and related proteolysis product free adducts in type 1 diabetes and microalbuminuria. The effects observed point to the possibility that patients who have efficient mechanisms of disposal of damaged proteins might be at an increased risk of developing microalbuminuria but not early renal function decline. The findings support the concept that the mechanisms responsible for microalbuminuria may differ from the mechanisms involved in the initiation of early renal function decline.

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Available from: Monika A Niewczas
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    • "The molecular fingerprint of damage to major plasma proteins includes amino acids modified by chlorination, nitrosylation, oxidation, nitration, crosslinking (disulphide and Schiff base) formation, glycation, dialdehydic lipids such as malondialdehyde and 4-hydoxynonenal [28] [29] [30] [31] [32] [33] [34] [35]. The mechanisms underlying these modifications have been considered extensively elsewhere and are not considered here [36]. "
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    ABSTRACT: The presence and concentrations of modified proteins circulating in plasma depend on rates of protein synthesis, modification and clearance. In early studies, the proteins most frequently analysed for damage were those which were more abundant in plasma (e.g. albumin and immunoglobulins) which exist at up to 10 orders of magnitude higher concentrations than other plasma proteins e.g. cytokines. However, advances in analytical techniques using mass spectrometry and immuno-affinity purification methods, have facilitated analysis of less abundant, modified proteins and the nature of modifications at specific sites is now being characterised. The damaging reactive species that cause protein modifications in plasma principally arise from reactive oxygen species (ROS) produced by NADPH oxidases (NOX), nitric oxide synthases (NOS) and oxygenase activities; reactive nitrogen species (RNS) from myeloperoxidase (MPO) and NOS activities; and hypochlorous acid from MPO. Secondary damage to proteins may be caused by oxidized lipids and glucose autooxidation. In this review, we focus on redox regulatory control of those enzymes and processes which control protein maturation during synthesis, produce reactive species, repair and remove damaged plasma proteins. We have highlighted the potential for alterations in the extracellular redox compartment to regulate intracellular redox state and, conversely, for intracellular oxidative stress to alter the cellular secretome and composition of extracellular vesicles. Through secreted, redox-active regulatory molecules, changes in redox state may be transmitted to distant sites.
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    • "Because of their stability to acid conditions, CML and pentosidine are generally the most frequently measured AGEs. Despite having a concentration differing by over 100- fold in proteins, their levels are strongly correlated with one another and with concentrations of other AGEs in tissue proteins (Thornalley et al., 2003; Niwa, 2006; Perkins et al., 2012). On the other hand, CMC is present at approximately the same level as CML in muscle protein (Alt et al., 2004), while glucosepane is the most abundant cross-linked AGE formed in vivo in proteins (Biemel, Friedl, & Lederer, 2002b). "
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    ABSTRACT: The Maillard reaction includes a complex network of processes affecting food and biopharmaceutical products; it also occurs in living organisms and has been strictly related to cell aging, to the pathogenesis of several (chronic) diseases, such as diabetes, uremia, cataract, liver cirrhosis and various neurodegenerative pathologies, as well as to peritoneal dialysis treatment. Dozens of compounds are involved in this process, among which a number of protein-adducted derivatives that have been simplistically defined as early, intermediate and advanced glycation end-products. In the last decade, various bottom-up proteomic approaches have been successfully used for the identification of glycation/glycoxidation protein targets as well as for the characterization of the corresponding adducts, including assignment of the modified amino acids. This article provides an updated overview of the mass spectrometry-based procedures developed to this purpose, emphasizing their partial limits with respect to current proteomic approaches for the analysis of other post-translational modifications. These limitations are mainly related to the concomitant sheer diversity, chemical complexity, and variable abundance of the various derivatives to be characterized. Some challenges to scientists are finally proposed for future proteomic investigations to solve main drawbacks in this research field. © 2013 Wiley Periodicals, Inc. Mass Spec Rev 9999: 1-29, 2013.
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    • "Inefficient clearance of AGE-rich peptides and recirculation of these ‘toxic’ molecules might be responsible for vascular damage in diabetic patients. Recent studies have shown that in patients with chronic renal failure (CRF), reduced renal metabolism of AGEs likely accounts for the accumulation of AGEs in the serum [5], [50]. Further experiments with various pathological conditions are needed to establish whether AGE-rich diets can cause an accumulation of AGEs that is harmful to animal or human bodies. "
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    ABSTRACT: N(ε)-carboxymethyl-lysine (CML) is a major advanced glycation end-product (AGEs) widely found in foods. The aim of our study was to evaluate how exogenous CML-peptide is dynamically absorbed from the gastrointestinal tract and eliminated by renal tubular secretion using microPET imaging. The present study consisted of three investigations. In study I, we synthesized the imaging tracer (18)F-CML by reacting N-succinimidyl 4-(18)F-fluorobenzoate ((18)F-SFB) with CML. In study II, the biological activity of (18)F-CML was evaluated in RAW264.7 cells and HepG2 cells. In study III, the biodistribution and elimination of AGEs in ICR mice were studied in vivo following tail vein injection and intragastric administration of (18)F-CML. The formation of (18)F-CML was confirmed by comparing its retention time with the corresponding reference compound (19)F-CML. The radiochemical purity (RCP) of (18)F-CML was >95%, and it showed a stable character in vitro and in vivo. Uptake of (18)F-CML by RAW264.7 cells and HepG2 cells could be inhibited by unmodified CML. (18)F-CML was quickly distributed via the blood, and it was rapidly excreted through the kidneys 20 min after tail vein injection. However, (18)F-CML was only slightly absorbed following intragastric administration. After administration of (18)F-CML via a stomach tube, the radioactivity was completely localized in the stomach for the first 15 min. At 150 min post intragastric administration, intense accumulation of radioactivity in the intestines was still observed. PET technology is a powerful tool for the in vivo analysis of the gastrointestinal absorption of orally administered drugs. (18)F-CML is hardly absorbed by the gastrointestinal tract. It is rapidly distributed and eliminated from blood following intravenous administration. Thus, it may not be harmful to healthy bodies. Our study showed the feasibility of noninvasively imaging (18)F-labeled AGEs and was the first to describe CML-peptide gastrointestinal absorption by means of PET.
    Preview · Article · Mar 2013 · PLoS ONE
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