Diabetes mellitus is associated with hyperglycemia and with accelerated non-enzymatic glycation, increased oxidative stress and free radical production. The aim of the present study was to evaluate the levels of proteins glycation and oxidation parameters, compare them between poorly and well controlled children with type 1 diabetes mellitus, and determine the impact of glycemic control on these parameters. Blood and serum were obtained from 81 patients with type 1 diabetes mellitus (DM1) (20 patients had long-term good glycemic control [GGC], 61 patients had long-term poor glycemic control [PGC]). Thirty-one healthy children were used as controls. Fructosamine (FAM) was determined by a spectrophotometric method, HbA1c was measured by LPLC, serum advanced glycation end-products (s-AGEs) were determined fluorimetrically, and advanced oxidation protein products (AOPP) were measured spectrophotometrically. We observed significantly higher FAM, HbA1c, s-AGEs and AOPP levels in the patients with DM1 compared with controls, and significantly higher FAM, HbA1c and sAGEs levels in the PGC group compared with the GGC group. AOPP was higher in the PGC group than in the GGC group, but not significantly. In the PGC group we observed significant correlations between HbA1c and HDL-C (r = -0.306, p = 0.01), HbA1c and s-AGEs (r = 0.486, p < 0.001), and HbA1c and AOPP (r = 0.447, p < 0.01). s-AGEs significantly correlated with triacylglycerols (TAG) (r = 0.537, p < 0.001) and AOPP with HDL-C (r = -0.336, p < 0.05), TAG (r = 0.739, p < 0.001) and s-AGEs (r = 0.577, p < 0.001). In conclusion, our results showed both glycative and oxidative stress are increased in the PGC diabetic group compared with controls, they are linked with glycemic control, and probably contribute to the development of diabetic complications. We suggest that the measurement of not only HbA1c but also s-AGEs and AOPP may be useful to predict the risk of development of diabetic complications.

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A new procedure was developed to determine in urine the concentrations of N(epsilon)-(carboxymethyl)lysine (CML) and N(epsilon)-(carboxyethyl)lysine (CEL), the major products of oxidative modification of glycated proteins, to assess levels of oxidative stress in physiological systems. The urine samples were acetonitrile-deproteinized, then derivatized by ethylchloroformate, and N(O,S)-ethoxycarbonyl ethyl esters of amino acids were analysed by isotope dilution gas chromatography/mass spectrometry. Recovery averaged 89%. Linearity was excellent (r = 0.998-0.999) in the 0.5-25 micromol/L range for CML and CEL. The limit of detection of this assay was 0.1 micromol/L (corresponding to 0.20 pmol of CML or CEL on column). Intra-day and inter-day precisions were likewise excellent, with relative standard deviations <4.63 and <6.15%, respectively. Accuracy of CML and CEL determination (15 micromol/L) was 2.9 and 5.9% of the estimated theoretical value. The time from obtaining the urine sample to determination of the concentration from the chromatographic peak was 80 min or less. This method is sensitive, reproducible, accurate, relatively cheap and very simple. It can be useful for laboratories involved in the diagnosis and monitoring of age-related chronic diseases.

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Abstract:Aim: In this study we tried to investigate the impact of glycemic control on parameters of glycation and infl ammation in overweight and obese patients with type 2 diabetes mellitus. Patients and methods: Markers of glycation (HbA 1c , AGEs; measured by HPLC and spectrofl uorimetry, resp.) and infl ammatory markers (IL-6, IL-8, TNF-α, MCP-1; measured by xMAP technology) were assessed in 69 patients with T2DM, of whom 32 were patients were with poor glycemic control (PGC group), 37 patients were with good glycemic control (GGC group) and 23 were healthy blood volunteers. Results: Our results showed that plasma levels of fl uorescent AGEs, IL-6, IL-8, TNF-α, and MCP-1 were signifi cantly increased in PGC and GGC groups in comparison with control group, while the levels were higher in PGC group in comparison with GGC group, but the difference was not signifi cant. We found a positive correlation between AGEs and MCP-1 and between TNF-α and creatinine in PGC group. We found signifi cantly decreased levels of glycated HbA 1c and AGEs in patients who used statins compared to patients who used fi brates. We observed benefi cial impact of treatment with oral antidiabetic (OAD) agents + insulin on levels of IL-8, TNF-α and TAG in comparison with treatment with insulin alone. Conclusions: Despite good glycemic compensation of patients with T2DM, levels of AGEs and infl ammatory markers remained signifi cantly elevated in comparison with healthy controls. There was a benefi cial impact of treatment with OAD agents + insulin in sense of lowering the low-grade infl ammation (Tab. 3, Fig. 7, Ref. 113). Text in PDF www.elis.sk.