Plasma Proteins Modified by Advanced Glycation End Products (AGEs) Reveal Site-specific Susceptibilities to Glycemic Control in Patients with Type 2 Diabetes

Abstract

Protein glycation refers to the reversible reaction between aldoses (or ketoses) and amino groups yielding relatively stable Amadori (or Heyns) products. Consecutive oxidative cleavage reactions of these products or the reaction of amino groups with other reactive substances (e.g. ␣-dicarbonyls) yield advanced glycation end products (AGEs) that can alter the structures and functions of proteins. AGEs have been identified in all organisms, and their contents appear to rise with some diseases, such as diabetes and obesity. Here, we report a pilot study using highly sensitive and specific proteomics approach to identify and quantify AGE modification sites in plasma proteins by reversed phase HPLC mass spectrometry in tryptic plasma digests. In total, 19 AGE modification sites corresponding to 11 proteins were identified in patients with type 2 diabetes mellitus under poor glycemic control. The modification degrees of 15 modification sites did not differ among cohorts of normoglycemic lean or obese and type 2 diabetes mellitus patients under good and poor glycemic control. The contents of two amide-AGEs in human serum albumin and apolipoprotein A-II were significantly higher in patients with poor glycemic control, although the plasma levels of both proteins were similar among all plasma samples. These two modification sites might be useful to predict long term, AGE-related complications in diabetic patients, such as impaired vision, increased arterial stiffness, or decreased kidney function.The reaction between reducing sugars (i.e. aldoses or ketoses) and amines is termed glycation or nonenzymatic glycosylation (1). The initially formed aldimines or ketimines can rearrange forming relatively stable Amadori or Heyns products, respectively. Consecutive rearrangements and oxidations yield advanced glycation end products (AGEs) 3 (2-6), a structurally heterogeneous group of compounds including cross-linked proteins (1). Alternatively, ␣-dicarbonyls generated by Amadori degradation (7,8), saccharide autoxidation (9, 10), lipid peroxidation (11, 12), and enzymatic reactions (13-15) can modify the side chains of lysine and arginine residues (16 -18). Degradation of glucose-derived Amadori products, for example, yields 1-deoxyglucosone (7, 19) and its tautomers, which upon nucleophilic attack by a N ⑀ -amino group of a lysine residue undergo hydrolytic ␤-cleavage yielding amide-AGEs, such as N ⑀ -acetyl-, formyl-, and glycerinyl-lysine ( Fig. 1) (20). Generally, AGEs can alter the structure of proteins contributing to diabetic retinopathy (21, 22), increased arterial stiffness (23, 24), or impaired renal function (25) along with aging and diabetes. Furthermore, receptor binding-induced activation of proinflammatory signaling pathways is associated with the development and progression of atherosclerosis (26) and nephropathy (25).AGEs have been localized in biological tissues by immunohistochemistry (27-29) and quantified with ELISA using monoclonal antibodies recognizing methylglyoxal-derived hydroimidazolone (30), imidazol...

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