Glyoxalase 1 overexpression improves neurovascular coupling and limits development of mild cognitive impairment in a mouse model of type 1 diabetes.

Diabetes is associated with cognitive impairment, but the underlying mechanism remains unclear. Methylglyoxal (MGO), a precursor to advanced glycation endproducts (AGEs), is elevated in diabetes and linked to microvascular dysfunction. In this study, overexpression of the MGO-detoxifying enzyme glyoxalase 1 (Glo1) was used in a mouse model of diabetes to explore whether MGO accumulation in diabetes causes cognitive impairment. Diabetes was induced with streptozotocin. Fasting blood glucose, cognitive function, cerebral blood flow, neurovascular coupling (NVC), Glo1 activity, MGO and AGEs were assessed. In diabetes, MGO-derived hydroimidazolone-1 increased in the cortex, and was decreased in Glo1-overexpressing mice compared to controls. Visuospatial memory was decreased in diabetes, but not in Glo1/diabetes. NVC response time was slightly increased in diabetes, and normalised in the Glo1-overexpressing group. No impact of diabetes or Glo1 overexpression on blood-brain barrier integrity or vascular density was observed. Diabetes induced a mild visuospatial memory impairment and slightly reduced NVC response speed and these effects were mitigated by Glo1. This study shows a link between MGO-related AGE accumulation and cerebrovascular/cognitive functions in diabetes. Modulation of the MGO-Glo1 pathway may be a novel intervention strategy in patients with diabetes who have cerebrovascular complications. KEY POINTS: Diabetes is associated with an increased risk of stroke, cognitive decline, depression and Alzheimer's disease, but the underlying mechanism remains unclear. Methylglyoxal (MGO), a highly reactive by-product of glycolysis, plays an important role in the development of diabetes-associated microvascular dysfunction in the periphery and is detoxified by the enzyme glyoxalase 1. Diabetes reduced visuospatial memory in mice and slowed the neurovascular coupling response speed, which was improved by overexpression of glyoxalase 1. MGO formation and MGO-derived advanced glycation endproduct (AGE) accumulation in the brain of diabetic mice are associated with a slight reduction in neurovascular coupling and mild cognitive impairment. The endogenous formation of MGO, and the accumulation of MGO-derived AGEs, might be a potential target in reducing the risk of vascular cognitive impairment in people with diabetes.

Increased methylglyoxal formation in plasma and tissues during a glucose tolerance test is derived from exogenous glucose.

The dicarbonyl compound methylglyoxal (MGO) is a major precursor in the formation of advanced glycation endproducts (AGEs). MGO and AGEs are increased in subjects with diabetes and are associated with fatal and nonfatal cardiovascular disease. Previously, we have shown that plasma MGO concentrations rapidly increase in the postprandial phase, with a higher increase in individuals with type 2 diabetes. In current study, we investigated whether postprandial MGO formation in plasma and tissues originates from exogenous glucose and whether the increased plasma MGO concentration leads to a fast formation of MGO-derived AGEs. We performed a stable isotope-labelled oral glucose tolerance test (OGTT) in 12 healthy males with universally labelled D(+)13C glucose. Analysis of plasma-labelled 13C3 MGO and glucose levels at 11 time-points during the OGTT revealed that the newly formed MGO during OGTT is completely derived from exogenous glucose. Moreover, a fast formation of protein-bound MGO-derived AGEs during the OGTT was observed. In accordance, ex-vivo incubation of MGO with plasma or albumin showed a rapid decrease in MGO and a fast increase in MGO-derived AGEs. In an intraperitoneal glucose tolerance test in C57BL/6J mice, we confirmed that the formation of postprandial MGO is derived from exogenous glucose in plasma and also showed in tissues that MGO is increased and this is also from exogenous glucose. Collectively, increased formation of MGO during a glucose tolerance test arises from exogenous glucose both in plasma and in tissues, and this leads to a fast formation of MGO-derived AGEs.

Pyridoxamine reduces methylglyoxal and markers of glycation and endothelial dysfunction, but does not improve insulin sensitivity or vascular function in abdominally obese individuals: A randomized double-blind placebo-controlled trial.

AIM: To investigate the effects of pyridoxamine (PM), a B6 vitamer and dicarbonyl scavenger, on glycation and a large panel of metabolic and vascular measurements in a randomized double-blind placebo-controlled trial in abdominally obese individuals. MATERIALS AND METHODS: Individuals (54% female; mean age 50 years; mean body mass index 32 kg/m2 ) were randomized to an 8-week intervention with either placebo (n = 36), 25 mg PM (n = 36) or 200 mg PM (n = 36). We assessed insulin sensitivity, ß-cell function, insulin-mediated microvascular recruitment, skin microvascular function, flow-mediated dilation, and plasma inflammation and endothelial function markers. PM metabolites, dicarbonyls and advanced glycation endproducts (AGEs) were measured using ultra-performance liquid chromatography tandem mass spectrometry. Treatment effects were evaluated by one-way ANCOVA. RESULTS: In the high PM dose group, we found a reduction of plasma methylglyoxal (MGO) and protein-bound Nδ-(5-hydro-5-methyl-4-imidazolon-2-yl)-ornithine (MG-H1), as compared to placebo. We found a reduction of the endothelial dysfunction marker soluble vascular cell adhesion molecule-1 (sVCAM-1) in the low and high PM dose group and of soluble intercellular adhesion molecule-1 (sICAM-1) in the high PM dose, as compared to placebo. We found no treatment effects on insulin sensitivity, vascular function or other functional outcome measurements. CONCLUSIONS: This study shows that PM is metabolically active and reduces MGO, AGEs, sVCAM-1 and sICAM-1, but does not affect insulin sensitivity and vascular function in abdominally obese individuals. The reduction in adhesion markers is promising because these are important in the pathogenesis of endothelial damage and atherosclerosis.

A Citrus and Pomegranate Complex Reduces Methylglyoxal in Healthy Elderly Subjects: Secondary Analysis of a Double-Blind Randomized Cross-Over Clinical Trial.

Reactive α-dicarbonyls (α-DCs), such as methylglyoxal (MGO), glyoxal (GO), and 3-deoxyglucosone (3-DG), are potent precursors in the formation of advanced glycation end products (AGEs). In particular, MGO and MGO-derived AGEs are thought to be involved in the development of vascular complications in diabetes. Experimental studies showed that citrus and pomegranate polyphenols can scavenge α-DCs. Therefore, the aim of this study was to evaluate the effect of a citrus and pomegranate complex (CPC) on the α-DCs plasma levels in a double-blind, placebo-controlled cross-over trial, where thirty-six elderly subjects were enrolled. They received either 500 mg of Citrus sinensis peel extract and 200 mg of Punica granatum concentrate in CPC capsules or placebo capsules for 4 weeks, with a 4-week washout period in between. For the determination of α-DCs concentrations, liquid chromatography tandem mass spectrometry was used. Following four weeks of CPC supplementation, plasma levels of MGO decreased by 9.8% (-18.7 nmol/L; 95% CI: -36.7, -0.7 nmol/L; p = 0.042). Our findings suggest that CPC supplementation may represent a promising strategy for mitigating the conditions associated with MGO involvement. This study was registered on clinicaltrials.gov as NCT03781999.

A 4-Week Diet Low or High in Advanced Glycation Endproducts Has Limited Impact on Gut Microbial Composition in Abdominally Obese Individuals: The deAGEing Trial.

Dietary advanced glycation endproducts (AGEs), abundantly present in Westernized diets, are linked to negative health outcomes, but their impact on the gut microbiota has not yet been well investigated in humans. We investigated the effects of a 4-week isocaloric and macronutrient-matched diet low or high in AGEs on the gut microbial composition of 70 abdominally obese individuals in a double-blind parallel-design randomized controlled trial (NCT03866343). Additionally, we investigated the cross-sectional associations between the habitual intake of dietary dicarbonyls, reactive precursors to AGEs, and the gut microbial composition, as assessed by 16S rRNA amplicon-based sequencing. Despite a marked percentage difference in AGE intake, we observed no differences in microbial richness and the general community structure. Only the Anaerostipes spp. had a relative abundance >0.5% and showed differential abundance (0.5 versus 1.11%; p = 0.028, after low- or high-AGE diet, respectively). While the habitual intake of dicarbonyls was not associated with microbial richness or a general community structure, the intake of 3-deoxyglucosone was especially associated with an abundance of several genera. Thus, a 4-week diet low or high in AGEs has a limited impact on the gut microbial composition of abdominally obese humans, paralleling its previously observed limited biological consequences. The effects of dietary dicarbonyls on the gut microbiota composition deserve further investigation.

Physical activity and markers of glycation in older individuals: data from a combined cross-sectional and randomized controlled trial (EXAMIN AGE).

BACKGROUND: Advanced glycation end products (AGEs) are protein modifications that are predominantly formed from dicarbonyl compounds that arise from glucose and lipid metabolism. AGEs and sedentary behavior have been identified as a driver of accelerated (vascular) aging. The effect of physical activity on AGE accumulation is unknown. Therefore, we investigated whether plasma AGEs and dicarbonyl levels are different across older individuals that were active or sedentary and whether plasma AGEs are affected by high-intensity interval training (HIIT). METHODS: We included healthy older active (HA, n=38, 44.7% female, 60.1 ± 7.7 years old) and healthy older sedentary (HS, n=36, 72.2% female, 60.0 ± 7.3 years old) individuals as well as older sedentary individuals with increased cardiovascular risk (SR, n=84, 50% female, 58.7 ± 6.6 years old). The SR group was randomized into a 12-week walking-based HIIT program or control group. We measured protein-bound and free plasma AGEs and dicarbonyls by ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) at baseline and after the HIIT intervention. RESULTS: Protein-bound AGE Nε-(carboxymethyl)lysine (CML) was lower in SR (2.6 ± 0.5 µmol/l) and HS (3.1 ± 0.5 µmol/l) than in HA (3.6 ± 0.6 µmol/l; P<0.05) and remained significantly lower after adjustment for several potential confounders. None of the other glycation markers were different between HS and HA. HIIT did not change plasma AGEs and dicarbonyls in SR. DISCUSSION: Although lifestyle interventions may act as important modulators of cardiovascular risk, HIIT is not a potent short-term intervention to reduce glycation in older individuals, underlining the need for other approaches, such as pharmacological agents, to reduce AGEs and lower cardiovascular risk in this population.

Quercetin, but Not Epicatechin, Decreases Plasma Concentrations of Methylglyoxal in Adults in a Randomized, Double-Blind, Placebo-Controlled, Crossover Trial with Pure Flavonoids.

Background: Methylglyoxal (MGO) is the most potent precursor of advanced glycation end products (AGEs). MGO and AGEs have been associated with diabetes, its complications, and other age-related diseases. Experimental studies have shown that the flavonoids quercetin and epicatechin are able to scavenge MGO and lower AGE formation. Objective: Data on the effects of these flavonoids on MGO and AGE concentrations in humans are not yet available. We therefore investigated the effect of quercetin and epicatechin on the concentrations of MGO and AGEs in a post hoc analysis. Methods: Thirty-seven apparently healthy, nonsmoking adults with a systolic blood pressure between 125 and 160 mm Hg at screening were included in a randomized, double-blind, placebo-controlled crossover trial. Participants ingested (-)-epicatechin (100 mg/d), quercetin 3-glucoside (160 mg/d), or placebo capsules for periods of 4 wk separated by 4-wk washout periods. Fasting blood samples were collected at the start and end of each intervention period. Liquid chromatography-tandem mass spectrometry was used to determine plasma concentrations of the dicarbonyl compounds MGO, glyoxal (GO), and 3-deoxyglucosone (3-DG) and free and protein-bound AGEs. Gene expression of glyoxalase 1 (GLO1), the enzyme involved in the degradation of MGO, was determined by either microarray or quantitative reverse transcriptase-polymerase chain reaction. Results: The treatment effect (Δtreatment - Δplacebo) of quercetin on MGO was -40.2 nmol/L (95% CI: -73.6, -6.8 nmol/L; P = 0.019), a decrease of 11% from baseline values, whereas GO, 3-DG, and free and protein-bound AGEs did not change significantly. Epicatechin did not affect the concentrations of dicarbonyls and free and protein-bound AGEs. We did not find a significant change in expression of GLO1. Conclusions: In apparently healthy (pre)hypertensive men and women, quercetin but not epicatechin decreased plasma MGO concentrations. Quercetin may potentially form a new treatment strategy for diseases in which MGO plays a pivotal role. This study was registered at clinicaltrials.gov as NCT01691404.

Advanced Glycation Endproducts Are Increased in the Animal Model of Multiple Sclerosis but Cannot Be Reduced by Pyridoxamine Treatment or Glyoxalase 1 Overexpression.

Multiple sclerosis (MS) is a demyelinating autoimmune disease of the central nervous system (CNS). The immune response in MS patients leads to the infiltration of immune cells in the CNS and their subsequent activation. Immune cell activation induces a switch towards glycolysis. During glycolysis, the dicarbonyl product methylglyoxal (MGO) is produced. MGO is a glycating agent that can rapidly form advanced glycation endproducts (AGEs). In turn, AGEs are able to induce inflammatory responses. The glyoxalase system is the endogenous defense system of the body to reduce the burden of MGO thereby reducing AGE formation. This system consists of glyoxalase-1 and glyoxalase-2 which are able to detoxify MGO to D-lactate. We investigated whether AGE levels are induced in experimental autoimmune encephalitis (EAE), an inflammatory animal model of MS. Twenty seven days post EAE induction, MGO and AGE (Nε-(carboxymethyl)lysine (CML), Nε-(carboxyethyl)lysine (CEL), 5-hydro-5-methylimidazolone (MG-H1)) levels were significantly increased in the spinal cord of mice subjected to EAE. Yet, pyridoxamine treatment and glyoxalase-1 overexpression were unable to counteract AGE production during EAE and did not influence the clinical course of EAE. In conclusion, AGEs levels increase during EAE in the spinal cord, but AGE-modifying treatments do not inhibit EAE-induced AGE production and do not affect disease progression.

Disparity in the micronutrient content of diets high or low in advanced glycation end products (AGEs) does not explain changes in insulin sensitivity.

We have previously shown that an isoenergetic low advanced glycation end products (AGEs) diet matched for macronutrient content improved insulin sensitivity compared to high AGE diet. Here, we evaluated the differences in micronutrient intake of these two dietary patterns and if they could explain differences in insulin sensitivity. Participants consumed the intervention diets each for 2 weeks with 4 weeks of habitual dietary intake (washout) in-between. Dietary analysis revealed that the high AGE diet contained greater levels of retinol equivalents (RE) (478.9 + 151.3 µg/day versus 329.0 + 170.0 µg/day; p < .006), vitamin A (806.3 + 223.5 (µg RE)/day versus 649.1 + 235.8 (µg RE)/day; p < .05) and thiamine (2.3 + 0.6 mg/day versus 1.6 + 0.4 mg/day; p = .014) compared to the low AGE diet. The changes in polyunsaturated fat, retinol, vitamin A and thiamine did not correlate with changes in insulin sensitivity (all p > .1) therefore are unlikely to explain observed changes in insulin sensitivity. (clinicaltrials.gov:NCT00422253).

Low 25-hydroxyvitamin D2 and 25-hydroxyvitamin D3 levels are independently associated with macroalbuminuria, but not with retinopathy and macrovascular disease in type 1 diabetes: the EURODIAB prospective complications study.

BACKGROUND: Low circulating levels of total vitamin D [25(OH)D] and 25(OH)D3 have been associated with vascular complications in few studies on individuals with type 1 diabetes. However, these measures are affected by UV light exposure. Circulating 25(OH)D2, however, solely represents dietary intake of vitamin D2, but its association with complications of diabetes is currently unknown. We investigated the associations between 25(OH)D2 and 25(OH)D3 and the prevalence of albuminuria, retinopathy and cardiovascular disease (CVD) in individuals with type 1 diabetes. METHODS: We measured circulating 25(OH)D2 and 25(OH)D3 in 532 individuals (40 ± 10 years old, 51 % men) with type 1 diabetes who participated in the EURODIAB Prospective Complications Study. Cross-sectional associations of 25(OH)D2 and 25(OH)D3 with albuminuria, retinopathy and CVD were assessed with multiple logistic regression analyses adjusted for age, sex, season, BMI, smoking, HbA1c, total-HDL-cholesterol-ratio, systolic blood pressure, antihypertensive medication, eGFR, physical activity, alcohol intake, albuminuria, retinopathy and CVD, as appropriate. RESULTS: Fully adjusted models revealed that 1 nmol/L higher 25(OH)D2 and 10 nmol/L higher 25(OH)D3 were associated with lower prevalence of macroalbuminuria with ORs (95 % CI) of 0.56 (0.43;0.74) and 0.82 (0.72;0.94), respectively. These vitamin D species were not independently associated with microalbuminuria, non-proliferative and proliferative retinopathy or CVD. CONCLUSIONS: In individuals with type 1 diabetes, both higher 25(OH)D2 and 25(OH)D3 are associated with a lower prevalence of macroalbuminuria, but not of retinopathy and CVD. Prospective studies are needed to further examine the associations between 25(OH)D2 and 25(OH)D3 and the development of microvascular complications and CVD in type 1 diabetes.

Higher levels of advanced glycation endproducts in human carotid atherosclerotic plaques are associated with a rupture-prone phenotype.

AIMS: Rupture-prone atherosclerotic plaques are characterized by inflammation and a large necrotic core. Inflammation is linked to high metabolic activity. Advanced glycation endproducts (AGEs) and their major precursor methylglyoxal are formed during high metabolic activity and can have detrimental effects on cellular function and may induce cell death. Therefore, we investigated whether plaque AGEs are increased in human carotid rupture-prone plaques and are associated with plaque inflammation and necrotic core formation. METHODS AND RESULTS: The protein-bound major methylglyoxal-derived AGE 5-hydro-5-methylimidazolone (MG-H1) and N(ε)-(carboxymethyl)lysine (CML) were measured in human carotid endarterectomy specimens (n = 75) with tandem mass spectrometry. MG-H1 and CML levels were associated with rupture-prone plaques, increased protein levels of the inflammatory mediators IL-8 and MCP-1 and with higher MMP-9 activity. Immunohistochemistry showed that AGEs accumulated predominantly in macrophages surrounding the necrotic core and co-localized with cleaved caspase-3. Intra-plaque comparison revealed that glyoxalase-1 (GLO-1), the major methylglyoxal-detoxifying enzyme, mRNA was decreased (-13%, P < 0.05) in ruptured compared with stable plaque segments. In line, in U937 monoctyes, we found reduced (GLO-1) activity (-38%, P < 0.05) and increased MGO (346%, P < 0.05) production after stimulation with the inflammatory mediator TNF. Direct incubation with methylglyoxal increased apoptosis up to two-fold. CONCLUSION: This is the first study showing that AGEs are associated with human rupture-prone plaques. Furthermore, this study suggests a cascade linking inflammation, reduced GLO-1, methylglyoxal- and AGE-accumulation, and subsequent apoptosis. Thereby, AGEs may act as mediators of the progression of stable to rupture-prone plaques, opening a window towards novel treatments and biomarkers to treat cardiovascular diseases.

Plasma advanced glycation end-products and skin autofluorescence are increased in COPD.

Chronic obstructive pulmonary disease (COPD) is associated with systemic inflammation and oxidative stress. These conditions may lead to the formation of advanced glycation end-products (AGEs). In this study we investigated in 88 COPD patients and 55 control subjects (80% ex-smokers) the association of the plasma protein-bound AGEs N(ε)-(carboxymethyl)lysine (CML), pentosidine, N(ε)-(carboxyethyl)lysine (CEL), and AGE accumulation in skin by skin autofluorescence (AFR), with lung function. Mean ± sd plasma CML was decreased (COPD 61.6 ± 15.6 nmol · mmol(-1) lysine, never-smokers 80.7 ± 19.8 nmol · mmol(-1) lysine and ex-smokers 82.9 ± 19.3 nmol · mmol(-1) lysine) and CEL (COPD 39.1 ± 10.9 nmol · mmol(-1) lysine, never-smokers 30.4 ± 5.0 nmol · mmol(-1) lysine and ex-smokers 27.7 ± 6.4 nmol · mmol(-1) lysine) and AFR (COPD 3.33 ± 0.67 arbitrary units (AU), never-smokers 2.24 ± 0.45 AU and ex-smokers 2.31 ± 0.47 AU) were increased in COPD patients compared to controls. Disease state was inversely associated with CML, and linearly associated with CEL and AFR. Performing regression analyses in the total group, CEL and AFR showed a negative association and CML a positive association with lung function, even after correction for potential confounders. In conclusion, CEL and AFR were negatively and CML was positively associated with disease state. In the total group only the AGEs showed an association with forced expiratory volume in 1 s. Our data suggest that AGEs are involved in the pathophysiology of COPD, although their exact role remains to be determined.

Association of plasma sRAGE, but not esRAGE with lung function impairment in COPD.

RATIONALE: Plasma soluble Receptor for Advanced Glycation End Product (sRAGE) is considered as a biomarker in COPD. The contribution of endogenous sRAGE (esRAGE) to the pool of plasma sRAGE and the implication of both markers in COPD pathogenesis is however not clear yet. The aim of the current study was therefore to measure plasma levels of esRAGE comparative to total sRAGE in patients with COPD and a control group. Further, we established the relations of esRAGE and total sRAGE with disease specific characteristics such as lung function and DLCO, and with different circulating AGEs. METHODS: Plasma levels of esRAGE and sRAGE were measured in an 88 patients with COPD and in 55 healthy controls. FEV1 (%predicted) and FEV1/VC (%) were measured in both groups; DLCO (%predicted) was measured in patients only. In this study population we previously reported that the AGE Nϵ-(carboxymethyl) lysine (CML) was decreased, Nϵ-(carboxyethyl) lysine (CEL) increased and pentosidine was not different in plasma of COPD patients compared to controls. RESULTS: Plasma esRAGE (COPD: 533.9 ± 412.4, CONTROLS: 848.7 ± 690.3 pg/ml; p = 0.000) was decreased in COPD compared to controls. No significant correlations were observed between plasma esRAGE levels and lung function parameters or plasma AGEs. A positive correlation was present between esRAGE and total sRAGE levels in the circulation. Confirming previous findings, total sRAGE (COPD: 512.6 ± 403.8, CONTROLS: 1834 ± 804.2 pg/ml; p < 0.001) was lower in patients compared to controls and was positively correlated FEV1 (r = 0.235, p = 0.032), FEV1/VC (r = 0.218, p = 0.047), and DLCO (r = 0.308, p = 0.006). sRAGE furthermore did show a significant positive association with CML (r = 0.321, p = 0.003). CONCLUSION: Although plasma esRAGE is decreased in COPD patients compared to controls, only total sRAGE showed a significant and independent association with FEV1, FEV1/VC and DLCO, indicating that total sRAGE but not esRAGE may serve as marker of COPD disease state and severity.

Quantification of glyoxal, methylglyoxal and 3-deoxyglucosone in blood and plasma by ultra performance liquid chromatography tandem mass spectrometry: evaluation of blood specimen.

BACKGROUND: The reactive α-oxoaldehydes glyoxal (GO), methylglyoxal (MGO) and 3-deoxyglucosone (3-DG) have been linked to diabetic complications and other age-related diseases. Numerous techniques have been described for the quantification of α-oxoaldehydes in blood or plasma, although with several shortcomings such as the need of large sample volume, elaborate extraction steps or long run-times during analysis. Therefore, we developed and evaluated an improved method including sample preparation, for the quantification of these α-oxoaldehydes in blood and plasma with ultra performance liquid chromatography tandem mass spectrometry (UPLC MS/MS). METHODS: EDTA plasma and whole blood samples were deproteinized using perchloric acid (PCA) and subsequently derivatized with o-phenylenediamine (oPD). GO, MGO and 3-DG concentrations were determined using stable isotope dilution UPLC MS/MS with a run-to-run time of 8 min. Stability of α-oxoaldehyde concentrations in plasma and whole blood during storage was tested. The concentration of GO, MGO and 3-DG was measured in EDTA plasma of non-diabetic controls and patients with type 2 diabetes (T2DM). RESULTS: Calibration curves of GO, MGO and 3-DG were linear throughout selected ranges. Recoveries of these α-oxoaldehydes were between 95% and 104%. Intra- and inter-assay CVs were between 2% and 14%. CONCLUSIONS: To obtain stable and reliable α-oxoaldehyde concentrations, immediate centrifugation of blood after blood sampling is essential and the use of EDTA as anticoagulant is preferable. Moreover, immediate precipitation of plasma protein with PCA stabilized α-oxoaldehyde concentrations for at least 120 min. With the use of the developed method, we found increased plasma concentrations of GO, MGO and 3-DG in T2DM as compared with non-diabetic controls.

Methylglyoxal mediates the association between 2-hour plasma glucose and HbA1c with inflammation: The Maastricht Study.

CONTEXT: Glucose excursions in persons with diabetes may drive chronic inflammation. Methylglyoxal (MGO) is formed from glucose, is elevated in persons with diabetes, and is a potent glycating agent linked with inflammation. OBJECTIVE: We investigated whether glucose excursions are associated with low-grade inflammation and whether MGO mediates this association. DESIGN: We used data from The Maastricht Study, an extensive phenotyping study into the etiology of type 2 diabetes and its complications. PARTICIPANTS: Data of 3017 participants, who underwent an oral glucose tolerance test and where data on MGO levels and inflammation were available, were used. MAIN OUTCOME MEASURES: Linear regression analyses, adjusted for potential confounders, evaluated associations between fasting plasma glucose (FPG), 2-hours plasma glucose (2h-PG) and HbA1c and low-grade inflammation (stdß, [95% confidence interval]), calculated from plasma concentrations of C-reactive protein, serum amyloid A, interleukin-6, interleukin-8, tumor necrosis factor and soluble intercellular adhesion molecule-1. Mediation analyses investigated whether MGO mediated these associations. RESULTS: 2h-PG (0.172 [0.110; 0.234]) and HbA1c (0.148 [0.101; 0.196]), but not FPG (0.049 [-0.002; 0.100]), were associated with low-grade inflammation. 2h-PG and HbA1c were also associated with 2h-MGO (0.471 [0.407; 0.534], and 0.244 [0.195; 0.294], respectively). Furthermore, 2h-MGO was independently and positively associated with low-grade inflammation (0.078 [0.037, 0.120]). 2h-MGO mediated 23% of the association between 2h-PG and inflammation, and 16% of the association between HbA1c and inflammation. CONCLUSIONS: MGO mediates the association between post-load glucose excursions and HbA1c with inflammation, providing evidence for a role of postprandial MGO formation to hyperglycemia-induced low-grade inflammation.

Placental Methylglyoxal in Preeclampsia: Vascular and Biomarker Implications.

BACKGROUND: Preeclampsia is a multifaceted syndrome that includes maternal vascular dysfunction. We hypothesize that increased placental glycolysis and hypoxia in preeclampsia lead to increased levels of methylglyoxal (MGO), consequently causing vascular dysfunction. METHODS: Plasma samples and placentas were collected from uncomplicated and preeclampsia pregnancies. Uncomplicated placentas and trophoblast cells (BeWo) were exposed to hypoxia. The reactive dicarbonyl MGO and advanced glycation end products (Nε-(carboxymethyl)lysine [CML], Nε-(carboxyethyl)lysine [CEL], and MGO-derived hydroimidazolone [MG-H]) were quantified using liquid chromatography-tandem mass spectrometry. The activity of GLO1 (glyoxalase-1), that is, the enzyme detoxifying MGO, was measured. The impact of MGO on vascular function was evaluated using wire/pressure myography. The therapeutic potential of the MGO-quencher quercetin and mitochondrial-specific antioxidant mitoquinone mesylate (MitoQ) was explored. RESULTS: MGO, CML, CEL, and MG-H2 levels were elevated in preeclampsia-placentas (+36%, +36%, +25%, and +22%, respectively). Reduced GLO1 activity was observed in preeclampsia-placentas (-12%) and hypoxia-exposed placentas (-16%). Hypoxia-induced MGO accumulation in placentas was mitigated by the MGO-quencher quercetin. Trophoblast cells were identified as the primary source of MGO. Reduced GLO1 activity was also observed in hypoxia-exposed BeWo cells (-26%). Maternal plasma concentrations of CML and the MGO-derived MG-H1 increased as early as 12 weeks of gestation (+16% and +17%, respectively). MGO impaired endothelial barrier function, an effect mitigated by MitoQ, and heightened vascular responsiveness to thromboxane A2. CONCLUSIONS: This study reveals the accumulation of placental MGO in preeclampsia and upon exposure to hypoxia, demonstrates how MGO can contribute to vascular impairment, and highlights plasma CML and MG-H1 levels as promising early biomarkers for preeclampsia.

Increased Levels of Circulating Methylglyoxal Have No Consequence for Cerebral Microvascular Integrity and Cognitive Function in Young Healthy Mice.

Diabetes and other age-related diseases are associated with an increased risk of cognitive impairment, but the underlying mechanisms remain poorly understood. Methylglyoxal (MGO), a by-product of glycolysis and a major precursor in the formation of advanced glycation end-products (AGEs), is increased in individuals with diabetes and other age-related diseases and is associated with microvascular dysfunction. We now investigated whether increased levels of circulating MGO can lead to cerebral microvascular dysfunction, blood-brain barrier (BBB) dysfunction, and cognitive impairment. Mice were supplemented or not with 50 mM MGO in drinking water for 13 weeks. Plasma and cortical MGO and MGO-derived AGEs were measured with UPLC-MS/MS. Peripheral and cerebral microvascular integrity and inflammation were investigated. Cerebral blood flow and neurovascular coupling were investigated with laser speckle contrast imaging, and cognitive tests were performed. We found a 2-fold increase in plasma MGO and an increase in MGO-derived AGEs in plasma and cortex. Increased plasma MGO did not lead to cerebral microvascular dysfunction, inflammation, or cognitive decline. This study shows that increased concentrations of plasma MGO are not associated with cerebral microvascular dysfunction and cognitive impairment in healthy mice. Future research should focus on the role of endogenously formed MGO in cognitive impairment.

Plasma levels of advanced glycation endproducts are associated with type 1 diabetes and coronary artery calcification.

BACKGROUND: Advanced glycation endproducts (AGEs) may play a role in the development of coronary artery calcification (CAC) in type 1 diabetes (T1DM). We studied plasma AGEs in association with T1DM and CAC, and whether or not the latter association could be explained by low-grade inflammation (LGI) or endothelial dysfunction (ED). METHODS: We studied 165 individuals with and 169 without T1DM. CAC was quantified in a CAC score based on CT-scanning. Plasma levels of protein-bound pentosidine, Nϵ-(carboxymethyl)lysine (CML) and Nϵ-(carboxyethyl)lysine (CEL) were measured with HPLC/UPLC with fluorescence detection or tandem-mass spectrometry. Tetrahydropyrimidine (THP) was measured with ELISA, as were HsCRP, and sVCAM-1 and vWF, as markers for LGI and ED, respectively. Associations were analyzed with ANCOVA and adjusted for age, sex, BMI, waist-to-hip ratio, smoking, blood pressure, lipid profile, eGFR and T1DM. RESULTS: Individuals with T1DM had higher plasma levels of pentosidine, CML and THP compared with controls; means (95% CI) were 0.69 (0.65-0.73) vs. 0.51 (0.48-0.54) nmol/mmol LYS, p < 0.001; 105 (102-107) vs. 93 (90-95) nmol/mmol LYS, p < 0.001; and 126 (118-134) vs. 113 (106-120) U/mL, p = 0.03, respectively. Levels of pentosidine were higher in individuals with T1DM with a moderate to high compared with a low CAC score, means (95% CI) were 0.81 (0.70-0.93) vs. 0.67 (0.63-0.71) nmol/mmol LYS, p = 0.03, respectively. This difference was not attenuated by adjustment for LGI or ED. CONCLUSIONS: We found a positive association between pentosidine and CAC in T1DM. These results may indicate that AGEs are possibly involved in the development of CAC in individuals with T1DM.

Effects of fructose added to an oral glucose tolerance test on plasma glucose excursions in healthy adults.

Background and objective: Previous experimental studies have shown that fructose interacts with glucose metabolism by increasing hepatic glucose uptake. However, human studies investigating the effects of small ('catalytic') amounts of fructose, added to an oral glucose load, on plasma glucose levels remain inconclusive. The aim of this study, therefore, was to repeat and extend these previous studies by examining the plasma glucose response during a 75 g oral glucose tolerance test (OGTT) with the addition of different doses of fructose. Methods: Healthy adults (n = 13) received an OGTT without addition of fructose and OGTTs with addition of different doses of fructose (1, 2, 5, 7.5 and 15 g) in a random order, on six separate occasions. Plasma glucose levels were measured every 15 min for 120 min during the study. Findings: The plasma glucose incremental area under the curve (iAUC) of the OGTT without addition of fructose was not significantly different from any OGTT with fructose (p ≥ 0.2 for all fructose doses). Similar results were observed when these data were clustered with data from a similar, previous study (pooled mean difference: 10.6; 95%CI: 45.0; 23.8 for plasma glucose iAUC of the OGTT without addition of fructose versus an OGTT with 5 g fructose; fixed-effect meta-analysis, n = 38). Of interest, serum fructose increased from 4.8 µmol/L (interquartile range: 4.1-5.9) at baseline to 5.3 µmol/L (interquartile range: 4.8-7.5) at T = 60 min during an OGTT without addition of fructose (p = 0.002). Conclusion: Low doses of fructose added to an OGTT do not affect plasma glucose levels in healthy adults. The role of endogenous fructose production, as a potential explanation of these null-findings, deserves further investigation.

The Intake of Dicarbonyls and Advanced Glycation Endproducts as Part of the Habitual Diet Is Not Associated with Intestinal Inflammation in Inflammatory Bowel Disease and Irritable Bowel Syndrome Patients.

A Western diet comprises high levels of dicarbonyls and advanced glycation endproducts (AGEs), which may contribute to flares and symptoms in inflammatory bowel disease (IBD) and irritable bowel syndrome (IBS). We therefore investigated the intake of dietary dicarbonyls and AGEs in IBD and IBS patients as part of the habitual diet, and their association with intestinal inflammation. Food frequency questionnaires from 238 IBD, 261 IBS as well as 195 healthy control (HC) subjects were used to calculate the intake of dicarbonyls methylglyoxal, glyoxal, and 3-deoxyglucosone, and of the AGEs Nε-(carboxymethyl)lysine, Nε-(1-carboxyethyl)lysine and methylglyoxal-derived hydroimidazolone-1. Intestinal inflammation was assessed using faecal calprotectin. The absolute dietary intake of all dicarbonyls and AGEs was higher in IBD and HC as compared to IBS (all p < 0.05). However, after energy-adjustment, only glyoxal was lower in IBD versus IBS and HC (p < 0.05). Faecal calprotectin was not significantly associated with dietary dicarbonyls and AGEs in either of the subgroups. The absolute intake of methylglyoxal was significantly higher in patients with low (<15 µg/g) compared to moderate calprotectin levels (15−<50 µg/g, p = 0.031). The concentrations of dietary dicarbonyls and AGEs generally present in the diet of Dutch patients with IBD or IBS are not associated with intestinal inflammation, although potential harmful effects might be counteracted by anti-inflammatory components in the food matrix.