Methylglyoxal
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The great research interest in the quantification of reactive carbonyl compounds (RCCs), such as methylglyoxal (MGO) in biological and environmental samples, is reflected by the fact that several publications have described specific strategies to perform this task. Thus, many reagents have also been reported for the derivatization of RCCs to effectively detect and quantify the resulting compounds using sensitive techniques such as liquid chromatography coupled with mass spectrometry (LC-MS). However, the choice of the derivatization protocol is not always clear, and a comparative evaluation is not feasible because detection limits from separate reports and determined with different instruments are hardly comparable. Consequently, for a systematic comparison, we tested 21 agents in one experimental setup for derivatization of RCCs prior to LC-MS analysis. This consisted of seven commonly employed reagents and 14 similar reagents, three of which were designed and synthesized by us. All reagents were probed for analytical responsiveness of the derivatives and stability of the reaction mixtures. The results showed that derivatives of 4-methoxyphenylenediamine and 3-methoxyphenylhydrazine-reported here for the first time for derivatization of RCCs-provided a particularly high responsiveness with ESI-MS detection. We applied the protocol to investigate MGO contamination of laboratory water and show successful quantification in a lipoxidation experiment. In summary, our results provide valuable information for scientists in establishing accurate analysis of RCCs.
PURPOSE: The aim of this study was to investigate whether luteoloside, a flavonoid, could protect human dental pulp cells (HDPCs) against inflammation and oxidative stress induced by methylglyoxal (MGO), one of the advanced glycated end products (AGE) substances. METHODS: HDPCs were stimulated with MGO and treated with luteoloside. MTT assay was used to determine cell viability. Protein expression was measured via western blotting. Reactive oxygen species (ROS) were measured with a Muse Cell Analyzer. Alkaline phosphatase activity (ALP) and Alizarin red staining were used for mineralization assay. RESULTS: Luteoloside down-regulated the expression of inflammatory molecules such as ICAM-1, VCAM-1, TNF-α, IL-1β, MMP-2, MMP-9, and COX-2 in MGO-induced HDPCs without showing any cytotoxicity. It attenuated ROS formation and enhanced osteogenic differentiation such as ALP activity and Alizarin red staining in MGO-induced HDPCs. Overall, luteoloside showed protective actions against inflammation and oxidative stress in HDPCs induced by MGO through its anti-inflammatory, anti-oxidative, and osteogenic activities by down-regulating p-JNK in the MAPK pathway. CONCLUSION: These results suggest that luteoloside might be a potential adjunctive therapeutic agent for treating pulpal pathological conditions in patients with diabetes mellitus.
- MeSH
- anthrachinony * MeSH
- antiflogistika farmakologie MeSH
- glukosidy * MeSH
- kultivované buňky MeSH
- lidé MeSH
- luteolin * MeSH
- osteogeneze * fyziologie MeSH
- oxid hořečnatý MeSH
- pyruvaldehyd * toxicita MeSH
- reaktivní formy kyslíku MeSH
- zánět chemicky indukované farmakoterapie MeSH
- zubní dřeň MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Glycation is a process closely related to the aging and pathogenesis of diabetic complications. In this process, reactive α-dicarbonyl compounds (e.g., methylglyoxal) cause protein modification accompanied with potential loss of their biological activity and persistence of damaged molecules in tissues. We suppose that glutathione S-transferases (GSTs), a group of cytosolic biotransformation enzymes, may be modified by glycation in vivo, which would provide a rationale of its use as a model protein for studying glycation reactions. Glycation of GST by methylglyoxal, fructose, and glucose in vitro was studied. The course of protein glycation was evaluated using the following criteria: enzyme activity, formation of advanced glycation end-products using fluorescence and western blotting, amine content, protein conformation, cross linking and aggregation, and changes in molecular charge of GST. The ongoing glycation by methylglyoxal 2 mM resulted in pronounced decrease in the GST activity. It also led to the loss of 14 primary amino groups, which was accompanied by changes in protein mobility during native polyacrylamide gel electrophoresis. Formation of cross links with molecular weight of 75 kDa was observed. Obtained results can contribute to understanding of changes, which proceed in metabolism of xenobiotics during diabetes mellitus and ageing.
- MeSH
- diabetes mellitus enzymologie MeSH
- fruktosa chemie metabolismus MeSH
- glukosa chemie metabolismus MeSH
- glutathiontransferasa chemie metabolismus MeSH
- katalýza MeSH
- lidé MeSH
- produkty pokročilé glykace chemie metabolismus MeSH
- pyruvaldehyd chemie metabolismus MeSH
- stárnutí metabolismus MeSH
- xenobiotika chemie metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
Glycation is a process closely related to the aging and pathogenesis of diabetic complications. Reactive alpha-dicarbonyl compounds (e.g., methylglyoxal) are formed during middle stage of glycation reaction. Compounds that would inhibit the glycation process have been seeked for years. The objective of this study was to investigate the inhibitory effect of hydroxycitric (0.25-2.5 mM) and uric acid (0.4-1.2 mM) on middle stage of protein glycation in vitro using the model containing aspartate aminotransferase (AST) and 0.5 mM methylglyoxal. Hydroxycitric acid, at all tested concentrations, reduced AST activity decrease and formation of fluorescent AGEs during incubation of the enzyme with methylglyoxal at 37 degrees C. This compound also prevented formation of high-molecular weight protein cross-links and changes in molecular charge of AST caused by glycation. Uric acid showed no positive anti-glycation activity. The results support the hypothesis that hydroxycitric acid has beneficial effects in controlling protein glycation.
- MeSH
- aspartátaminotransferasy chemie metabolismus MeSH
- citráty farmakologie MeSH
- fluorescence MeSH
- glykosylace účinky léků MeSH
- kvarterní struktura proteinů MeSH
- kyselina močová farmakologie MeSH
- ornithin analogy a deriváty metabolismus MeSH
- produkty pokročilé glykace metabolismus MeSH
- pyrimidiny metabolismus MeSH
- pyruvaldehyd farmakologie MeSH
- reagencia zkříženě vázaná farmakologie MeSH
- Sus scrofa MeSH
- western blotting MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- práce podpořená grantem MeSH
- Klíčová slova
- EMPAGLIFLOZIN,
- MeSH
- benzhydrylové sloučeniny * aplikace a dávkování farmakologie metabolismus MeSH
- diabetické angiopatie farmakoterapie chemicky indukované metabolismus MeSH
- glifloziny aplikace a dávkování farmakologie metabolismus MeSH
- glukosidy * aplikace a dávkování farmakologie metabolismus MeSH
- hypoglykemika * aplikace a dávkování farmakologie metabolismus MeSH
- klinická studie jako téma MeSH
- modely nemocí na zvířatech * MeSH
- potkani Wistar MeSH
- prediabetes * farmakoterapie chemicky indukované metabolismus MeSH
- pyruvaldehyd aplikace a dávkování škodlivé účinky MeSH
- sacharosa škodlivé účinky MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- práce podpořená grantem MeSH
Proteins are subject to oxidative modification and the formation of adducts with a broad spectrum of reactive species via enzymatic and non-enzymatic mechanisms. Here we report that in vitro non-enzymatic methylglyoxal (MGO) binding causes the inhibition and formation of MGO advanced glycation end-products (MAGEs) in Na+/K+-ATPase (NKA). Concretely, MGO adducts with NKA amino acid residues (mainly Arg) and Nε-(carboxymethyl)lysine (CML) formation were found. MGO is not only an inhibitor for solubilized NKA (IC50=91±16μM), but also for reconstituted NKA in the lipid bilayer environment, which was clearly demonstrated using a DPPC/DPPE liposome model in the presence or absence of the NKA-selective inhibitor ouabain. High-resolution mass spectrometric analysis of a tryptic digest of NKA isolated from pig (Sus scrofa) kidney indicates that the intracellular α-subunit is naturally (post-translationally) modified by MGO in vivo. In contrast to this, the β-subunit could only be modified by MGO artificially, and the transmembrane part of the protein did not undergo MGO binding under the experimental setup used. As with bovine serum albumin, serving as the water-soluble model, we also demonstrated a high binding capacity of MGO to water-poorly soluble NKA using a multi-spectral methodology based on electroanalytical, immunochemical and fluorimetric tools. In addition, a partial suppression of the MGO-mediated inhibitory effect could be observed in the presence of aminoguanidine (pimagedine), a glycation suppressor and MGO-scavenger. All the results here were obtained with the X-ray structure of NKA in the E1 conformation (3WGV) and could be used in the further interpretation of the functionality of this key enzyme in the presence of highly-reactive metabolic side-products, glycation agents and generally under oxidative stress conditions.
- MeSH
- guanidiny farmakologie MeSH
- hmotnostní spektrometrie MeSH
- konformace proteinů MeSH
- krystalografie rentgenová MeSH
- ledviny metabolismus MeSH
- ouabain farmakologie MeSH
- oxidační stres MeSH
- produkty pokročilé glykace chemie metabolismus MeSH
- pyruvaldehyd chemie metabolismus MeSH
- sérový albumin hovězí metabolismus MeSH
- skot MeSH
- sodíko-draslíková ATPasa antagonisté a inhibitory chemie metabolismus MeSH
- Sus scrofa MeSH
- vazba proteinů MeSH
- zvířata MeSH
- Check Tag
- skot MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Excessive methylglyoxal (MG) production contributes to metabolic and vascular changes by increasing inflammatory processes, disturbing regulatory mechanisms and exacerbating tissue dysfunction. MG accumulation in adipocytes leads to structural and functional changes. We used transcriptome analysis to investigate the effect of MG on metabolic changes in the visceral adipose tissue of hereditary hypetriglyceridaemic rats, a non-obese model of metabolic syndrome. Compared to controls, 4-week intragastric MG administration impaired glucose tolerance (p < 0.05) and increased glycaemia (p < 0.01) and serum levels of MCP-1 and TNFα (p < 0.05), but had no effect on serum adiponectin or leptin. Adipose tissue insulin sensitivity and lipolysis were impaired (p < 0.05) in MG-treated rats. In addition, MG reduced the expression of transcription factor Nrf2 (p < 0.01), which controls antioxidant and lipogenic genes. Increased expression of Mcp-1 and TNFα (p < 0.05) together with activation of the SAPK/JNK signaling pathway can promote chronic inflammation in adipose tissue. Transcriptome network analysis revealed the over-representation of genes involved in insulin signaling (Irs1, Igf2, Ide), lipid metabolism (Nr1d1, Lpin1, Lrpap1) and angiogenesis (Dusp10, Tp53inp1).
- Publikační typ
- časopisecké články MeSH
Methylglyoxal production is increased in diabetes. Methylglyoxal is efficiently detoxified by enzyme glyoxalase 1 (GLO1). The aim was to study the effect of diabetic and CKD milieu on (a) GLO1 gene expression in peripheral blood mononuclear cells; (b) GLO1 protein levels in whole blood; and (c) GLO1 activity in RBCs in vivo in diabetic vs. non-diabetic subjects with normal or slightly reduced vs. considerably reduced renal function (CKD1-2 vs. CKD3-4). A total of 83 subjects were included in the study. Gene expression was measured using real-time PCR, and protein levels were quantified using Western blotting. Erythrocyte GLO1 activity was measured spectrophotometrically. GLO1 gene expression was significantly higher in subjects with CKD1-2 compared to CKD3-4. GLO1 protein level was lower in diabetics than in non-diabetics. GLO1 activity in RBCs differed between the four groups being significantly higher in diabetics with CKD1-2 vs. healthy subjects and vs. nondiabeticsfig with CKD3-4. GLO1 activity was significantly higher in diabetics compared to nondiabetics. In conclusion, both diabetes and CKD affects the glyoxalase system. It appears that CKD in advanced stages has prevailing and suppressive effects compared to hyperglycaemia. CKD decreases GLO1 gene expression and protein levels (together with diabetes) without concomitant changes of GLO1 activity.
- MeSH
- chronická renální insuficience krev patologie MeSH
- diabetes mellitus krev patologie MeSH
- diabetické nefropatie krev patologie MeSH
- laktoylglutathionlyasa krev MeSH
- lidé středního věku MeSH
- lidé MeSH
- pyruvaldehyd krev MeSH
- senioři MeSH
- studie případů a kontrol MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
A genetic locus controlling the electrophoretic mobility of a methylglyoxal dehydrogenase (EC 1.2.1.23) in the rat is described. The locus, designated Mgd1, is expressed in liver and kidney. Inbred rat strains have fixed either allele Mgd1a or allele Mgd1b. Codominant expression is observed in heterozygotes, providing evidence for a tetrameric enzyme structure. Backcross progenies showed the expected 1:1 segregation ratio, and there is evidence that Mgd1 is linked to Pep3 and Fh1 on chromosome 13. There is also evidence for two additional methylglyoxal dehydrogenases: Mgd2, present in liver and kidney, and Mgd3, present only in heart.
- MeSH
- aldehydoxidoreduktasy * genetika MeSH
- alely * MeSH
- dominantní geny MeSH
- ektroforéza na škrobovém gelu MeSH
- geny MeSH
- izoenzymy analýza MeSH
- játra enzymologie MeSH
- konformace proteinů MeSH
- křížení genetické MeSH
- krysa rodu rattus * genetika metabolismus MeSH
- ledviny enzymologie MeSH
- myokard enzymologie MeSH
- orgánová specificita MeSH
- polymorfismus genetický * MeSH
- svalové proteiny genetika MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus * genetika metabolismus MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- práce podpořená grantem MeSH
- srovnávací studie MeSH
