Metabolic dysfunction of endothelial cells in hyperglycemia contributes to the development of vascular complications of diabetes where increased reactive glycating agent, methylglyoxal (MG), is involved. We assessed if increased MG glycation induced proteotoxic stress, identifying related metabolic drivers and protein targets. Human aortal endothelial cells (HAECs) were incubated in high glucose concentration (20 mM versus 5 mM control) in vitro for 3–6 days. Flux of glucose metabolism, MG formation and glycation and changes in cytosolic protein abundances, MG modification and proteotoxic responses were assessed. Similar studies were performed with human microvascular endothelial HMEC-1 cells where similar outcomes were observed. HAECs exposed to high glucose concentration showed increased cellular concentration of MG (2.27 ± 0.21 versus 1.28 ± 0.03 pmol/10 6 cells, P < 0.01) and formation of MG-modified proteins (24.0 ± 3.7 versus 14.1 ± 3.2 pmol/10 6 cells/day; P < 0.001). In proteomics analysis, high glucose concentration increased proteins of the heat shock response – indicating activation of the unfolded protein response (UPR) with downstream inflammatory and pro-thrombotic responses. Proteins susceptible to MG modification were enriched in protein folding, protein synthesis, serine/threonine kinase signalling, glycolysis and gluconeogenesis. MG was increased in high glucose by increased flux of MG formation linked to increased glucose metabolism mediated by proteolytic stabilisation and increase of hexokinase-2 (HK-2); later potentiated by proteolytic down regulation of glyoxalase 1 (Glo1) - the major enzyme of MG metabolism. Silencing of Glo1, selectively increasing MG, activated the UPR similarly. Silencing of HK-2 prevented increased glucose metabolism and MG formation. trans -Resveratrol and hesperetin combination (tRES-HESP) corrected increased MG and glucose metabolism by increasing expression of Glo1 and decreasing expression of HK-2. Increased MG glycation activates the UPR in endothelial cells and thereby may contribute to endothelial cell dysfunction in diabetic vascular disease where tRES-HESP may provide effective therapy.

BACKGROUNDDental caries and obesity are multifactorial diseases with diet being a common contributory factor.OBJECTIVEThe main purpose of the present study was to investigate the association between dental caries and obesity among special care female school children in Makkah City, Saudi Arabia.DESIGNAnalytical cross-sectional study.SETTINGSpecial schools in Makkah City.STUDY POPULATION AND METHODSSchools were chosen by lottery and female children were randomly selected. Dental caries detection was performed according to the World Health Organization criteria. The medical evaluation assessed the body mass index (BMI).MAIN OUTCOME MEASURESWith appropriate sample weighting, relationships between dmft/DMFT (decayed, missing, filled teeth for deciduous and permanent dentition) and obesity were assessed using multilevel logistic regression.RESULTSIn 275 special care children, the prevalence of dental caries was 56.7 percent. The mean dmft and DMFT scores for the entire study population were 3.9 (4.8) and 3.2 (4.1), respectively. Forty percent of children were mentally retarded, 22.2% presented with deafness, blindness or both, 18.9% presented with Down syndrome and 14.9% were autistic. From the total sample, the mean BMI was 20.2 (2.8). When adjusted for covariates, the logistic regression model showed strong association between caries and obesity (adjusted odds ratio=2.9; 95% CI=1.2–4.9).CONCLUSIONThis study demonstrated a significant association between caries frequency and overweight/ obesity in special care school children.LIMITATIONSSince the data was cross-sectional, causal relationships cannot be established and the observed association could be due to other unexplored factors. Because of cultural and ethical consideration, including segregation of gender in Saudi Arabia, only female children were included in the present study, which limited the findings.

Protein glycation in biological systems occurs predominantly on lysine, arginine and N-terminal residues of proteins. Major quantitative glycation adducts are found at mean extents of modification of 1–5 mol percent of proteins. These are glucose-derived fructosamine on lysine and N-terminal residues of proteins, methylglyoxal-derived hydroimidazolone on arginine residues and Nε-carboxymethyl-lysine residues mainly formed by the oxidative degradation of fructosamine. Total glycation adducts of different types are quantified by stable isotopic dilution analysis liquid chromatography-tandem mass spectrometry (LC-MS/MS) in multiple reaction monitoring mode. Metabolism of glycated proteins is followed by LC-MS/MS of glycation free adducts as minor components of the amino acid metabolome. Glycated proteins and sites of modification within them – amino acid residues modified by the glycating agent moiety - are identified and quantified by label-free and stable isotope labelling with amino acids in cell culture (SILAC) high resolution mass spectrometry. Sites of glycation by glucose and methylglyoxal in selected proteins are listed. Key issues in applying proteomics techniques to analysis of glycated proteins are: (i) avoiding compromise of analysis by formation, loss and relocation of glycation adducts in pre-analytic processing; (ii) specificity of immunoaffinity enrichment procedures, (iii) maximizing protein sequence coverage in mass spectrometric analysis for detection of glycation sites, and (iv) development of bioinformatics tools for prediction of protein glycation sites. Protein glycation studies have important applications in biology, ageing and translational medicine – particularly on studies of obesity, diabetes, cardiovascular disease, renal failure, neurological disorders and cancer. Mass spectrometric analysis of glycated proteins has yet to find widespread use clinically. Future use in health screening, disease diagnosis and therapeutic monitoring, and drug and functional food development is expected. A protocol for high resolution mass spectrometry proteomics of glycated proteins is given.