muramidase and Hyperglycemia

Metabolic syndrome and type 2 diabetes (T2DM) resulting from sustained hyperglycemia are considered as risk factors for cardiovascular disease (CVD) but the mechanism for their contribution to cardiopathogenesis is not well understood. Hyperglycemia induces nonenzymatic glycation of protein-yielding advanced glycation end products (AGE), which are postulated to stimulate interleukin-6 (IL-6) expression, triggering the liver to secrete tissue necrosis factor alpha (TNF-alpha) and C-reactive protein (CRP) that contribute to CVD pathogenesis. Although the high prevalence of periodontitis among individuals with diabetes is well known by dental researchers, it is relatively unrecognized in the medical community. The expression of the same proinflammatory mediators implicated in hyperglycemia (i.e., IL-6, TNF-alpha, and CRP) have been reported to be associated with periodontal disease and increased risk for CVD. We will review published evidence related to these 2 pathways and offer a consensus.

Topics: Carbohydrate Metabolism; Cardiovascular Diseases; Diabetes Mellitus, Type 2; Endothelium, Vascular; Focal Infection, Dental; Glycation End Products, Advanced; Humans; Hyperglycemia; Inflammation Mediators; Metabolic Syndrome; Muramidase; Periodontitis; Salivary Proteins and Peptides

Nonenzymatic modification of proteins in hyperglycemia is a major mechanism causing diabetic complications. These modifications can have pathogenic consequences when they target active site residues, thus affecting protein function. In the present study, we examined the role of glucose autoxidation in functional protein damage using lysozyme and RGD-α3NC1 domain of collagen IV as model proteins in vitro. We demonstrated that glucose autoxidation induced inhibition of lysozyme activity as well as NC1 domain binding to α(V)β(3) integrin receptor via modification of critical arginine residues by reactive carbonyl species (RCS) glyoxal (GO) and methylglyoxal while nonoxidative glucose adduction to the protein did not affect protein function. The role of RCS in protein damage was confirmed using pyridoxamine which blocked glucose autoxidation and RCS production, thus protecting protein function, even in the presence of high concentrations of glucose. Glucose autoxidation may cause protein damage in vivo since increased levels of GO-derived modifications of arginine residues were detected within the assembly interface of collagen IV NC1 domains isolated from renal ECM of diabetic rats. Since arginine residues are frequently present within protein active sites, glucose autoxidation may be a common mechanism contributing to ECM protein functional damage in hyperglycemia and oxidative environment. Our data also point out the pitfalls in functional studies, particularly in cell culture experiments, that involve glucose treatment but do not take into account toxic effects of RCS derived from glucose autoxidation.

Topics: Amino Acid Motifs; Animals; Arginine; Collagen Type IV; Diabetes Mellitus, Experimental; Glucose; Glyoxal; Hyperglycemia; Male; Micrococcus; Muramidase; Protein Carbonylation; Protein Structure, Tertiary; Proteins; Pyruvaldehyde; Random Allocation; Rats; Rats, Sprague-Dawley

Nonenzymatic modification of proteins is one of the key pathogenic factors in diabetic complications. Uncovering the mechanisms of protein damage caused by glucose is fundamental to understanding this pathogenesis and in the development of new therapies. We investigated whether the mechanism involving reactive oxygen species can propagate protein damage in glycation reactions beyond the classical modifications of lysine and arginine residues. We have demonstrated that glucose can cause specific oxidative modification of tryptophan residues in lysozyme and inhibit lysozyme activity. Furthermore, modification of tryptophan residues was also induced by purified albumin-Amadori, a ribose-derived model glycation intermediate. The AGE inhibitor pyridoxamine (PM) prevented the tryptophan modification, whereas another AGE inhibitor and strong carbonyl scavenger, aminoguanidine, was ineffective. PM specifically inhibited generation of hydroxyl radical from albumin-Amadori and protected tryptophan from oxidation by hydroxyl radical species. We conclude that oxidative degradation of either glucose or the protein-Amadori intermediate causes oxidative modification of protein tryptophan residues via hydroxyl radical and can affect protein function under physiologically relevant conditions. This oxidative stress-induced structural and functional protein damage can be ameliorated by PM via sequestration of catalytic metal ions and scavenging of hydroxyl radical, a mechanism that may contribute to the reported therapeutic effects of PM in the complications of diabetes.

Topics: Animals; Chickens; Glycation End Products, Advanced; Hydroxyl Radical; Hyperglycemia; Models, Biological; Models, Chemical; Muramidase; Oxidative Stress; Proteins; Pyridoxamine; Reactive Oxygen Species; Spectrophotometry, Ultraviolet; Superoxides; Tryptophan

Although the etiology of essential hypertension is not clearly understood, endothelial dysfunction from chronic infection and/or impaired glucose metabolism may be involved. We hypothesized that salivary lysozyme, a marker for oral infection and hyperglycemia, might display a significant relationship with hypertension, an early stage of cardiovascular disease. Logistic regression analyses of the Kuopio Oral Health and Heart Study demonstrated that persons with higher lysozyme levels were more likely to have hypertension, after adjustment for age, gender, smoking, BMI, diabetes, the ratio of total cholesterol to HDL cholesterol, and C-reactive protein. The exposure to increasing quartiles of lysozyme was associated with adjusted Odds Ratios for the outcome, hypertension, 1.00 (referent), 1.25, 1.42, and 2.56 (linear trend p < 0.003). When we restricted the sample to the individuals without heart disease (N = 250), we observed a non-significant trend for increasing odds. Our hypothesis--"high salivary lysozyme levels are associated with the odds of hypertension"--was confirmed.

Topics: Aged; Biomarkers; Case-Control Studies; Cohort Studies; Coronary Artery Disease; Cross-Sectional Studies; Female; Finland; Humans; Hyperglycemia; Hypertension; Male; Middle Aged; Muramidase; Odds Ratio; Reference Values; Regression Analysis; Saliva; Statistics, Nonparametric

Hyperglycaemia causes increased protein glycation and the formation of advanced glycation endproducts which underlie the complications of diabetes and ageing. Glycation is accompanied by metal-catalysed oxidation of glucose and Amadori products to form free radicals capable of protein fragmentation. Aged garlic extract is a potent antioxidant with established lipid-lowering effects attributed largely to a key ingredient called S-allyl cysteine. This study investigated the ability of aged garlic extract and S-allyl cysteine to inhibit advanced glycation in vitro. Bovine serum albumin (BSA) was glycated in the presence of Cu(2+) ions and different concentrations of aged garlic extract and protein fragmentation was examined by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). Lysozyme was glycated by glucose or methylglyoxal in the presence of different concentrations of aged garlic extract or S-allyl cysteine with subsequent analysis of glycation-derived crosslinking using SDS-PAGE. Amadori-rich protein was prepared by dialysing lysozyme that had been glycated by ribose for 24 h. This ribated lysozyme was reincubated and the effects of aged garlic extract, S-allyl cysteine and pyridoxamine on glycation-induced crosslinking was monitored. Aged garlic extract inhibited metal-catalysed protein fragmentation. Both aged garlic extract and S-allyl cysteine inhibited formation of glucose and methylglyoxal derived advanced glycation endproducts and showed potent Amadorin activity when compared to pyridoxamine. S-allyl cysteine inhibited formation of carboxymethyllysine (CML), a non-crosslinked advanced glycation endproduct derived from oxidative processes. Further studies are required to assess whether aged garlic extract and S-allyl cysteine can protect against the harmful effects of glycation and free radicals in diabetes and ageing.

Topics: Aging; Antioxidants; Cysteine; Diabetes Complications; Electrophoresis, Polyacrylamide Gel; Free Radicals; Garlic; Glucose; Glycation End Products, Advanced; Humans; Hyperglycemia; Muramidase; Peptide Fragments; Plant Extracts; Pyridoxamine; Pyruvaldehyde; Ribose

After a hyperglycaemic episode, glycated proteins remain in the body until removed by protein turnover. We have shown that in the absence of free sugar, such proteins can crosslink to native proteins, forming mixed dimers. They can also induce native proteins to crosslink into homodimers, presumably by release of a soluble crosslinking agent. Similar reactions in vivo could be responsible for the deposition of serum proteins in diabetic kidney, nerve and other tissues. Exposure to glycating sugar for brief periods, or a low concentration, still produced glycated protein capable of crosslinking to other proteins under sugar-free conditions. These crosslinks are nonfluorescent, unlike the advanced glycation endproducts usually observed.

Topics: Cross-Linking Reagents; Diabetes Mellitus; Electrophoresis, Polyacrylamide Gel; Female; Fructose; Glycation End Products, Advanced; Glycosylation; Humans; Hyperglycemia; Lactoglobulins; Macromolecular Substances; Muramidase; Spectrometry, Fluorescence