fructosyl-lysine and Diabetes-Mellitus

Ever since the discovery of the Maillard reaction in 1912 and the discovery of the interaction between advanced glycation end-products and cellular receptors, impressive progress has been made in the knowledge of nonenzymatic browning of proteins in vivo. This reaction which leads to the accumulation of random damage in extracellular proteins is known to have deleterious effects on biological function, and is associated with aging and complication in chronic diseases. Despite a controlled membrane permeability and a protective regulation of the cells, intracellular proteins are also altered by the Maillard reaction. Two main factors, protein turnover and the concentration of carbonyls, are involved in the rate of formation of the Maillard products. This paper reviews the key milestones of the discovery of the Maillard reaction in vivo, better known as glycation, and the factors which are likely to affect it.

Topics: Aging; Amino Acids; Diabetes Mellitus; Extracellular Matrix; Food; Free Radicals; Glucose; Glycation End Products, Advanced; Humans; Lysine; Maillard Reaction; Monosaccharides; Oxidation-Reduction; Protein Carbonylation; Proteins; Receptor for Advanced Glycation End Products

Hyperglycemic conditions of diabetes accelerate protein modifications by glucose leading to the accumulation of advanced glycation end-products (AGEs). We have investigated the conversion of protein-Amadori intermediate to protein-AGE and the mechanism of its inhibition by pyridoxamine (PM), a potent AGE inhibitor that has been shown to prevent diabetic complications in animal models. During incubation of proteins with physiological diabetic concentrations of glucose, PM prevented the degradation of the protein glycation intermediate identified as fructosyllysine (Amadori) by 13C NMR using [2-13C]-enriched glucose. Subsequent removal of glucose and PM led to conversion of protein-Amadori to AGE Nepsilon-carboxymethyllysine (CML). We utilized this inhibition of post-Amadori reactions by PM to isolate protein-Amadori intermediate and to study the inhibitory effect of PM on its degradation to protein-CML. We first tested the hypothesis that PM blocks Amadori-to-CML conversion by interfering with the catalytic role of redox metal ions that are required for this glycoxidative reaction. Support for this hypothesis was obtained by examining structural analogs of PM in which its known bidentate metal ion binding sites were modified and by determining the effect of endogenous metal ions on PM inhibition. We also tested the alternative hypothesis that the inhibitory mechanism involves formation of covalent adducts between PM and protein-Amadori. However, our 13C NMR studies demonstrated that PM does not react with the Amadori. Because the mechanism of interference with redox metal catalysis is operative under the conditions closely mimicking the diabetic state, it may contribute significantly to PM efficacy in preventing diabetic complications in vivo. Inhibition of protein-Amadori degradation by PM also provides a simple procedure for the isolation of protein-Amadori intermediate, prepared at physiological levels of glucose for relevancy, to study both the biological effects and the chemistry of post-Amadori pathways of AGE formation.

Topics: Animals; Carbon Isotopes; Diabetes Mellitus; Glucose; Glycation End Products, Advanced; Humans; Lysine; Metals; Nuclear Magnetic Resonance, Biomolecular; Oxidation-Reduction; Protein Processing, Post-Translational; Proteins; Pyridoxamine

The glycoxidation products Nepsilon-(carboxymethyl)lysine and pentosidine increase in skin collagen with age and at an accelerated rate in diabetes. Their age-adjusted concentrations in skin collagen are correlated with the severity of diabetic complications. To determine the relative roles of increased glycation and/or oxidation in the accelerated formation of glycoxidation products in diabetes, we measured levels of amino acid oxidation products, distinct from glycoxidative modifications of amino acids, as independent indicators of oxidative stress and damage to collagen in aging and diabetes. We show that ortho-tyrosine and methionine sulfoxide are formed in concert with Nepsilon-(carboxymethyl)lysine and pentosidine during glycoxidation of collagen in vitro, and that they also increase with age in human skin collagen. The age-adjusted levels of these oxidized amino acids in collagen was the same in diabetic and nondiabetic subjects, arguing that diabetes per se does not cause an increase in oxidative stress or damage to extracellular matrix proteins. These results provide evidence for an age-dependent increase in oxidative damage to collagen and support previous conclusions that the increase in glycoxidation products in skin collagen in diabetes can be explained by the increase in glycemia alone, without invoking a generalized, diabetes-dependent increase in oxidative stress.

Topics: Adolescent; Adult; Age Factors; Aged; Aged, 80 and over; Arginine; Child; Child, Preschool; Collagen; Diabetes Mellitus; Glucose; Humans; Infant; Infant, Newborn; Lysine; Methionine; Middle Aged; Oxidative Stress; Skin; Time Factors; Tyrosine

3-Deoxyglucosone (3-DG) is a reactive dicarbonyl sugar thought to be a key intermediate in the nonenzymatic polymerization and browning of proteins by glucose. 3-DG may be formed in vivo from fructose, fructose 3-phosphate, or Amadori adducts to protein, such as N epsilon-fructoselysine (FL), all of which are known to be elevated in body fluids or tissues in diabetes. Modification of proteins by 3-DG formed in vivo is thought to be limited by enzymatic reduction of 3-DG to less reactive species, such as 3-deoxyfructose (3-DF). In this study, we have measured 3-DF, as a metabolic fingerprint of 3-DG, in plasma and urine from a group of diabetic patients and control subjects. Plasma and urinary 3-DF concentrations were significantly increased in the diabetic compared with the control population (0.853 +/- 0.189 vs. 0.494 +/- 0.072 microM, P < 0.001, and 69.9 +/- 44.2 vs. 38.7 +/- 16.1 nmol/mg creatinine, P < 0.001, respectively). Plasma and urinary 3-DF concentrations correlated strongly with one another, with HbA1c (P < 0.005 in all cases), and with urinary FL (P < 0.02 and P = 0.005, respectively). The overall increase in 3-DF concentrations in plasma and urine in diabetes and their correlation with other indexes of glycemic control suggest that increased amounts of 3-DG are formed in the body during hyperglycemia in diabetes and then metabolized to 3-DF. These observations are consistent with a role for increased formation of the dicarbonyl sugar 3-DG in the accelerated browning of tissue proteins in diabetes.

Topics: Adolescent; Adult; Aged; Biomarkers; Blood Glucose; Diabetes Mellitus; Diabetes Mellitus, Type 1; Diabetes Mellitus, Type 2; Glycated Hemoglobin; Humans; Ketoses; Lysine; Middle Aged; Reference Values; Regression Analysis

Fifty-five serum samples from 99 Type 1 and 71 serum samples from 81 Type 2 diabetic patients (56% and 88%, respectively) brought about a 1.5-3.5-fold increase in total cholesterol content of cultured human intimal aortic cells. This atherogenic effect did not correlate with patient's age, diabetes duration or plasma lipid levels, and was mainly due to low density lipoprotein (LDL). Cholesterol accumulation in cells incubated with LDL highly correlated with that in cells exposed to corresponding patient's serum (r = 0.872 and r = 0.811, P < 0.0001, in Type 1 and Type 2 diabetic patients, respectively). In LDL from diabetic patients the sialic acid content was decreased by an average of 30% (P < 0.05), as compared with healthy subjects, and the fructosyl lysine content was increased by an average of 25% (P < 0.05). Atherogenic effect of patients' LDL significantly correlated with their fructosyl lysine content (P < 0.0001) and negatively correlated with sialic acid content (P < 0.0001). Two LDL fractions were further separated from the total LDL preparation by affinity chromatography on Ricinus communis agglutinin-agarose. The bound (desialylated) LDL fraction was characterized by an increased fructosyl lysine content and the altered neutral lipid and phospholipid composition, while non-bound (sialylated) LDL fraction did not differ from normal LDL. Desialylated, but not sialylated, LDL fraction induced massive cholesterol accumulation in cultured cells. In conclusion, the cholesterol accumulating effect of diabetic patients' blood sera is mainly related to atherogenic low density lipoprotein fraction, which is modified in various ways--by increased non-enzymatic glycosylation, desialylation and alterations in lipid composition. This multiple-modified LDL may contribute to the premature atherosclerosis development in diabetes mellitus.

Topics: Adolescent; Adult; Aorta; Arteriosclerosis; Cells, Cultured; Cholesterol; Diabetes Mellitus; Female; Humans; Lipids; Lipoproteins, LDL; Lysine; Male; Middle Aged; N-Acetylneuraminic Acid; Phospholipids; Sialic Acids; Tunica Intima

We studied 11 diabetic patients, all of whom had severe atherothrombotic disease, and 11 normal controls. Overall glycation was assessed by the extent of incorporation of [3H]-NaBH4 into fructosyl lysine separated from whole platelet proteins following aminoacid analysis. Fructosyl lysine represented 5.7% +/- 1.0 S.D. of the total radioactivity in the normal whole platelet samples. Increased glycation was observed in platelets from 5 of the 11 diabetics. Platelet glycation did not correlate with glycation of hemoglobin or albumin. The pattern of glycation of various platelet proteins in whole platelets, as determined by the incorporation of [3H]-NaBH4 into electrophoretically separated proteins did not display selectivity, although myosin and glycoproteins IIb and IIIa showed relatively increased levels of [3H]-NaBH4 incorporation. Artificially glycated platelet membranes exhibited glycation mainly in proteins corresponding to the electrophoretic mobility of myosin, glycoproteins IIb and IIIa.

Topics: Amino Acids; Blood Platelets; Blood Proteins; Borohydrides; Diabetes Mellitus; Glycated Hemoglobin; Glycated Serum Albumin; Glycation End Products, Advanced; Glycosylation; Humans; Hydrolysis; Lysine; Serum Albumin