Formalin, an aqueous solution of formaldehyde, has been the gold standard for fixation of histological samples for over a century. Despite its considerable advantages, growing evidence points to objective toxicity, particularly highlighting its carcinogenicity and mutagenic effects. In 2016, the European Union proposed a ban, but a temporary permission was granted in consideration of its fundamental role in the medical-diagnostic field. In the present study, we tested an innovative fixative, glyoxal acid-free (GAF) (a glyoxal solution deprived of acids), which allows optimal tissue fixation at structural and molecular level combined with the absence of toxicity and carcinogenic activity. An open-label, non-inferiority, multicentric trial was performed comparing fixation of histological specimens with GAF fixative vs standard phosphate-buffered formalin (PBF), evaluating the morphological preservation and the diagnostic value with four binary score questions answered by both the central pathology reviewer and local center reviewers. The mean of total score in the GAF vs PBF fixative groups was 3.7 ± 0.5 vs 3.9 ± 0.3 for the central reviewer and 3.8 ± 0.5 vs 4.0 ± 0.1 for the local pathologist reviewers, respectively. In terms of median value, similar results were observed between the two fixative groups, with a median value of 4.0. Data collected indicate the non-inferiority of GAF as compared to PBF for all organs tested. The present clinical performance study, performed following the international standard for performance evaluation of in vitro diagnostic medical devices, highlights the capability of GAF to ensure both structural preservation and diagnostic value of the preparations.
- MeSH
- Tissue Fixation * methods MeSH
- Fixatives * chemistry MeSH
- Formaldehyde * chemistry MeSH
- Glyoxal * MeSH
- Humans MeSH
- Check Tag
- Humans MeSH
- Male MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
- Equivalence Trial MeSH
- Multicenter Study MeSH
- Comparative Study MeSH
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
- Anthraquinones * MeSH
- Anti-Inflammatory Agents pharmacology MeSH
- Glucosides * MeSH
- Cells, Cultured MeSH
- Humans MeSH
- Luteolin * MeSH
- Osteogenesis * physiology MeSH
- Magnesium Oxide MeSH
- Pyruvaldehyde * toxicity MeSH
- Reactive Oxygen Species MeSH
- Inflammation chemically induced drug therapy MeSH
- Dental Pulp MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
Stárnutí lze definovat jako progresivní, ireverzibilní ztrátu vitality a stoupající mortalitu nevyhnutelně přicházejícím s věkem. Jde o stochastický proces, nikoliv program. Biochemická podstata stárnutí spočívá v nemožnosti zabránit nahromadění náhodných chyb v důležitých biomolekulách, tj. v DNA a v bílkovinách. Svou roli v komplikovaném procesu stárnutí hrají reaktivní formy kyslíku, mitochondriální dysfunkce, reaktivní karbonyly, poškození genomové DNA včetně telomer, epigenetické alterace, buněčná senescence a selhání proteostázy. I když všechny tkáně lidského těla stárnou, biologické limity údržby a oprav nejvíce dopadají na buňky, které se nedělí. Jelikož klíčové orgány lidského organismu, mozek, srdce a kosterní sval jsou založeny na postmitotických buňkách, lidská bytost ve svém biologickém těle nesmrtelnosti nikdy dosáhnout nemůže. V rámci svých biologických limitů jsou organismy vybaveny důležitými adaptabilními mechanismy tělesné údržby a oprav, jako je stresem stimulovaná buněčná odolnost vůči stresu (hormeze), autofagie či reakce na poškození DNA. Vhled do těchto mechanismů poskytuje vysvětlení pro příznivé působení faktorů životního stylu, jako je kalorická restrikce a fyzická aktivita, a do budoucna může přinést i léky schopné stárnutí zpomalit.
Aging can be defined as progressive and irreversible loss of vitality and increasing mortality coming inescapably with age. It is a stochastic process, rather than a program. The biochemical essence of aging lies in impossibility to prevent accumulation of random errors in important biomolecules, such as DNA and proteins. In the complicated process of aging, there are roles for reactive oxygen species, mitochondrial dysfunction, reactive carbonyls, and damage to genomic DNA including telomeres, epigenetic alterations, cell senescence, and collapse of proteostasis. Although all tissues of human body age, the biological limits of maintenance and repair affect the non-dividing cells the most. As the key organs of human body, brain, heart, and skeletal muscle, are based on postmitotic cells, a human being in his biological body can never reach immortality. Within the biological limits, organisms are equipped with important adaptable mechanisms for body maintenance and repair, such as stress-induced cellular resistance to stress (hormesis), autophagy and DNA damage response. Insight into these mechanisms provides rationale for benefits of life style interventions, such as caloric restriction and physical exercise, and in future might even bring drugs that could slow aging.
- MeSH
- Proteostasis MeSH
- Humans MeSH
- Mitochondria MeSH
- DNA Damage MeSH
- Pyruvaldehyde MeSH
- Reactive Oxygen Species MeSH
- Aging * genetics MeSH
- Check Tag
- Humans MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
- Keywords
- EMPAGLIFLOZIN,
- MeSH
- Benzhydryl Compounds * administration & dosage pharmacology metabolism MeSH
- Diabetic Angiopathies drug therapy chemically induced metabolism MeSH
- Sodium-Glucose Transporter 2 Inhibitors administration & dosage pharmacology metabolism MeSH
- Glucosides * administration & dosage pharmacology metabolism MeSH
- Hypoglycemic Agents * administration & dosage pharmacology metabolism MeSH
- Clinical Studies as Topic MeSH
- Disease Models, Animal * MeSH
- Rats, Wistar MeSH
- Prediabetic State * drug therapy chemically induced metabolism MeSH
- Pyruvaldehyde administration & dosage adverse effects MeSH
- Sucrose adverse effects MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
A side effect of diabetes is formation of glycated proteins and, from them, production of advanced early glycation end products that could determine aberrant immune responses at the systemic level. We investigated a relevant aberrant post-translational modification (PTM) in diabetes based on synthetic peptides modified on the lysine side chain residues with 1-deoxyfructopyranosyl moiety as a possible modification related to glycation. The PTM peptides were used as molecular probes for detection of possible specific autoantibodies developed by diabetic patients. The PDC-E2(167-186) sequence from the pyruvate dehydrogenase complex was selected and tested as a candidate peptide for antibody detection. The structure-based designed type I' β-turn CSF114 peptide was also used as a synthetic scaffold. Twenty-seven consecutive type 1 diabetic patients and 29 healthy controls were recruited for the study. In principle, the 'chemical reverse approach', based on the use of patient sera to screen the synthetic modified peptides, leads to the identification of specific probes able to characterize highly specific autoantibodies as disease biomarkers of autoimmune disorders. Quite surprisingly, both peptides modified with the (1-deoxyfructosyl)-lysine did not lead to significant results. Both IgG and IgM differences between the two populations were not significant. These data can be rationalized considering that i) IgGs in diabetic subjects exhibit a high degree of glycation, leading to decreased functionality; ii) IgGs in diabetic subjects exhibit a privileged response vs proteins containing advanced glycation products (e.g., methylglyoxal, glyoxal, glucosone, hydroimidazolone, dihydroxyimidazolidine) and only a minor one with respect to (1-deoxyfructosyl)-lysine.
- MeSH
- Diabetes Mellitus, Type 1 metabolism MeSH
- Glycosylation MeSH
- Glyoxal metabolism MeSH
- Imidazoles metabolism MeSH
- Immunoassay MeSH
- Ketoses metabolism MeSH
- Humans MeSH
- Lysine chemistry metabolism MeSH
- Peptides chemistry metabolism MeSH
- Protein Processing, Post-Translational MeSH
- Glycation End Products, Advanced metabolism MeSH
- Pyruvaldehyde metabolism MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
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.
Reactive dicarbonyls stimulate production of advanced glycation endproducts, increase oxidative stress and inflammation and contribute to the development of vascular complications. We measured concentrations of dicarbonyls - methylglyoxal (MG), glyoxal (GL) and 3-deoxyglucosone (3-DG) - in the heart and kidney of a model of metabolic syndrome - hereditary hypertriglyceridemic rats (HHTg) and explored its modulation by metformin. Adult HHTg rats were fed a standard diet with or without metformin (300 mg/kg b.w.) and dicarbonyl levels and metabolic parameters were measured. HHTg rats had markedly elevated serum levels of triacylglycerols (p<0.001), FFA (p<0.01) and hepatic triacylglycerols (p<0.001) along with increased concentrations of reactive dicarbonyls in myocardium (MG: p<0.001; GL: p<0.01; 3-DG: p<0.01) and kidney cortex (MG: p<0.01). Metformin treatment significantly reduced reactive dicarbonyls in the myocardium (MG: p<0.05, GL: p<0.05, 3-DG: p<0.01) along with increase of myocardial concentrations of reduced glutathione (p<0.01) and glyoxalase 1 mRNA expression (p<0.05). Metformin did not have any significant effect on dicarbonyls, glutathione or on glyoxalase 1 expression in kidney cortex. Chronically elevated hypertriglyceridemia was associated with increased levels of dicarbonyls in heart and kidney. Beneficial effects of metformin on reactive dicarbonyls and glyoxalase in the heart could contribute to its cardioprotective effects.
- MeSH
- Deoxyglucose analogs & derivatives metabolism MeSH
- Diet MeSH
- Stress, Physiological MeSH
- Glutathione metabolism MeSH
- Glyoxal metabolism MeSH
- Hypertriglyceridemia drug therapy genetics physiopathology MeSH
- Hypoglycemic Agents therapeutic use MeSH
- Rats MeSH
- Lactoylglutathione Lyase metabolism MeSH
- Metformin therapeutic use MeSH
- Myocardium metabolism MeSH
- Rats, Wistar MeSH
- Pyruvaldehyde metabolism MeSH
- Animals MeSH
- Check Tag
- Rats MeSH
- Male MeSH
- Animals MeSH
- Publication type
- Journal Article 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
- Renal Insufficiency, Chronic blood pathology MeSH
- Diabetes Mellitus blood pathology MeSH
- Diabetic Nephropathies blood pathology MeSH
- Lactoylglutathione Lyase blood MeSH
- Middle Aged MeSH
- Humans MeSH
- Pyruvaldehyde blood MeSH
- Aged MeSH
- Case-Control Studies MeSH
- Check Tag
- Middle Aged MeSH
- Humans MeSH
- Male MeSH
- Aged MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
Neurodegenerative disorders, such as Alzheimer's disease (AD) and Parkinson's disease (PD), are increasing in prevalence. Currently, there are no effective and specific treatments for these disorders. Recently, positive effects of the orexigenic hormone ghrelin on memory and learning were demonstrated in mouse models of AD and PD. In this study, we tested the potential neuroprotective properties of a stable and long-lasting ghrelin analog, Dpr(3)ghrelin (Dpr(3)ghr), in SH-SY5Y neuroblastoma cells stressed with 1.2 mM methylglyoxal (MG), a toxic endogenous by-product of glycolysis, and we examined the impact of Dpr(3)ghr on apoptosis. Pre-treatment with both 10(-5) and 10(-7) M Dpr(3)ghr resulted in increased viability in SH-SY5Y cells (determined by MTT staining), as well as reduced cytotoxicity of MG in these cells (determined by LDH assay). Dpr(3)ghr increased viability by altering pro-apoptotic and viability markers: Bax was decreased, Bcl-2 was increased, and the Bax/Bcl-2 ratio was attenuated. The ghrelin receptor GHS-R1 and Dpr(3)ghr-induced activation of PBK/Akt were immuno-detected in SH-SY5Y cells to demonstrate the presence of GHS-R1 and GHS-R1 activation, respectively. We demonstrated that Dpr(3)ghr protected SH-SY5Y cells against MG-induced neurotoxicity and apoptosis. Our data suggest that stable ghrelin analogs may be candidates for the effective treatment of neurodegenerative disorders.
- MeSH
- Apoptosis drug effects MeSH
- Ghrelin analogs & derivatives pharmacology MeSH
- Glycolysis drug effects MeSH
- L-Lactate Dehydrogenase metabolism MeSH
- Humans MeSH
- MAP Kinase Signaling System drug effects MeSH
- Membrane Potential, Mitochondrial drug effects MeSH
- Cell Line, Tumor MeSH
- Neuroprotective Agents pharmacology MeSH
- Neurotoxicity Syndromes prevention & control MeSH
- Apoptosis Regulatory Proteins biosynthesis genetics MeSH
- Pyruvaldehyde toxicity MeSH
- Receptors, Ghrelin biosynthesis MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
An electrophoretic apparatus with a flow-gating interface has been developed, enabling hydrodynamic sequence injection of the sample into the separation capillary from the liquid flow by underpressure generated in the outlet electrophoretic vessel. The properties of the apparatus were tested on an artificial sample of an equimolar mixture of 100μM potassium and sodium ions and arginine. The repeatability of the injection of the tested ions expressed as RSD (in%) for the peak area, peak height and migration time was in the range 0.76-2.08, 0.18-0.68 and 0.28-0.48, respectively. Under optimum conditions, the apparatus was used for sequence monitoring of the reaction between the antidiabetic drug phenyl biguanide and the glycation agent methyl glyoxal. The reaction solution was continuously sampled by a microdialysis probe from a thermostated external vessel using a syringe pump at a flow rate of 3μLmin(-1) and was injected into a separation capillary at certain time intervals. The electrophoretic separation progressed in a capillary with an internal diameter of 50μm with a length of 11.5cm and was monitored using a contactless conductivity detector.
- MeSH
- Arginine MeSH
- Biguanides chemistry MeSH
- Time Factors MeSH
- Potassium MeSH
- Electrophoresis, Capillary instrumentation methods MeSH
- Hydrodynamics * MeSH
- Hypoglycemic Agents chemistry MeSH
- Microdialysis MeSH
- Pyruvaldehyde chemistry MeSH
- Solutions MeSH
- Sodium MeSH
- Publication type
- Journal Article MeSH
