Neurodegenerative diseases (NDD) such as Alzheimer's (AD) and Parkinson's disease (PD) are distinct clinical entities, however, the aggregation of key neuronal proteins, presumably leading to ...neuronal demise appears to represent a common mechanism. It has become evident, that advanced glycation end products (AGEs) trigger the accumulation of such modified proteins, which eventually contributes to pathological aspect of NDDs. Increased levels of AGEs are found in amyloid plaques in AD brains and in both advanced and early PD (incidental Lewy body disease). The molecular mechanisms by which AGE dependent modifications may modulate the susceptibility towards NDDs, however, remain enigmatic and it is unclear, whether AGEs may serve as biomarker of NDD. In the present study, we examined AGEs (CML: Carboxymethyllysine and CEL: Carboxyethyllysine), markers of oxidative stress and micronutrients in the plasma of PD and AD patients and controls. As compared to healthy controls, AD females displayed lower levels of CEL while higher levels of CML were found in AD and PD patients. A somewhat similar pattern was observed for protein carbonyls (PC), revealing lower values exclusively in AD females, whereas AD males displayed significantly higher values compared to healthy controls and PD. Sex-specific differences were also observed for other relevant markers such as malondialdehyde, 3-nitrotyrosine, γ -tocopherols, retinol, plasma proteins and α-carotene, while α-tocopherols, β-carotene, lutein/zeaxanthin, β-cryptoxanthin and lycopene showed no relevant association. Taken together, our study suggests yet unappreciated differences of the distribution of AGEs among the sexes in NDD. We therefore suggest to make a clear distinction between sexes when analyzing oxidative (AGEs)-related stress and carbonyl-related stress and vitamins.
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•As compared to healthy controls, AD females displayed lower levels of CEL (carboxyethyllysine) while higher levels of CML (carboxymethyllysine) were found in AD and PD patients.•A somewhat similar pattern was observed for protein carbonyls (PC), revealing lower values exclusively in AD females, whereas AD males displayed significantly higher values compared to healthy controls and PD. Presumed carbonlyated residues were identified the in the major proteins associated with neurodegenerative diseases.•Sex-specific differences were also observed for other relevant markers such as malondialdehyde, 3-nitrotyrosine, γ-tocopherols, retinol, plasma proteins and α-carotene, while α-tocopherols, β-carotene, lutein/zeaxanthin, β-cryptoxanthin and lycopene showed no relevant association.
Supplemental oxygen is delivered to critically ill patients who require mechanical ventilation. Oxidative stress is a potential complication of oxygen therapy, resulting in damage to essential ...biomolecules such as proteins, lipids, and nucleic acids. Whether plasma levels of oxidative stress biomarkers vary based on how liberally oxygen therapy is applied during mechanical ventilation is unknown.
We carried out an oxidative stress substudy nested within a large multi-centre randomized controlled trial in which critically ill adults were randomized to receive either conservative oxygen therapy or standard oxygen therapy. Blood samples were collected at enrolment, and daily thereafter for up to three days. The antioxidant ascorbate (vitamin C) was assessed using HPLC with electrochemical detection and protein oxidation using a sensitive protein carbonyl ELISA. We also assessed whether critically ill patients with different disease states exhibited varying levels of oxidative stress biomarkers.
A total of 125 patients were included. Mean ascorbate concentrations decreased over time (from 25 ± 9 μmol/L to 14 ± 2 μmol/L, p < 0.001), however, there was no significant difference between the conservative oxygen group and standard care (p = 0.2), despite a significantly lower partial pressure of oxygen (PaO2) in the conservative oxygen group (p = 0.03). Protein carbonyl concentrations increased over time (from 208 ± 30 μmol/L to 249 ± 29 μmol/L; p = 0.016), however, there was no significant difference between the conservative and standard oxygen groups (p = 0.3). Patients with sepsis had significantly higher protein carbonyl concentrations than the other critically ill patients (293 ± 92 μmol/L vs 184 ± 24 μmol/L, p = 0.03). Within the septic subgroup, there were no significant differences in protein carbonyl concentrations between the two interventions (p = 0.4).
Conservative oxygen therapy does not alter systemic markers of oxidative stress in critically ill ventilated patients compared with standard oxygen therapy. Patients with sepsis exhibited elevated protein carbonyls compared with the other critically ill patients implying increased oxidative stress in this patient subgroup.
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•Antioxidant (ascorbate) concentrations are low in critically ill ventilated patients.•Protein carbonyls are elevated in septic patients relative to other critically ill patients.•Arterial partial pressure of oxygen (PaO2) is decreased with conservative oxygen therapy.•Oxidative stress biomarkers do not vary between conservative and standard oxygen therapy.
In this study, 154 pectoralis major breast muscles from three chicken age groups were separated into different severity levels of wooden breast condition by manual palpation to investigate the ...changes of lipid and protein oxidation and citrate synthase activity. TBARS values, carbonyl contents and loss of free thiol groups were greater (P < 0.05) in the superficial (ventral) part of severe wooden breast samples, whereas no clear change was found in the deep (dorsal) part. Mitochondrial citrate synthase activity declined with increasing severity of wooden breast myopathy. Both bird age and wooden breast degree showed significant main effects on these indicators, while no interaction effect was observed. Moreover, there were significant correlations between lipid and protein oxidation and citrate synthase activity. Here, we propose a hypothesis that lipid and protein damages and mitochondrial dysfunction is linked to the aggravation of the wooden breast myopathy.
•Increased severity of wooden breast increases lipid and protein oxidation.•Increased severity of wooden breast increases mitochondrial dysfunction.•Superficial parts of breast fillets are more vulnerable to wooden breast myopathy.•Accumulation of oxidative stress is associated with wooden breast development.
Modification of proteins in infant milk formula (IF) is of major concern to the dairy industry and consumers. Thermal treatment is required for microbiological safety, but heat, light, metal-ions and ...other factors may induce oxidative damage, and be a health risk. In this study protein modifications in IFs were quantified. IFs contained both reducible (disulphide) and non-reducible (di-tyrosine, lanthionine, lysinoalanine) protein cross-links. Dehydroalanine and the cross-linked species lanthionine and lysinoalanine were detected. Protein carbonyls were detected predominantly on high molecular mass materials. Oxidation products of phenylalanine (m-tyrosine), tryptophan (N-formylkynurenine, kynurenine, 3-hydroxykynurenine), tyrosine (di-tyrosine) and methionine (methionine sulphoxide) were detected, consistent with amino acid modification. Higher levels of most of the markers of protein modification were present in the hydrolysed protein brand, when compared to the conventional IF samples, indicative of increased damage during additional processing. Significant levels of racemised (D-) amino acids were present. These data indicate that amino acids in proteins in IFs are modified to a significant extent during manufacture, with hydrolysed IF being particularly prone.
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Dostopno za:
BFBNIB, DOBA, GIS, IJS, IZUM, KILJ, KISLJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
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•Curing of meat reduces lipid- and protein oxidation during in vitro digestion.•Oxidative stress and inflammation were not affected in cured meat-fed rats.•Cured meat consumption ...increased colonic Ruminococcaceae relative abundances.•Fecal acetaldehyde and diacetyl levels were increased in beef-fed rats.•Fecal CS2 levels were increased in rats consuming cured beef vs. fresh chicken.
Mechanisms explaining epidemiological associations between red (processed) meat consumption and chronic disease risk are not yet elucidated, but may involve oxidative reactions, microbial composition alterations, inflammation and/or the formation of toxic bacterial metabolites. First, in vitro gastrointestinal digestion of 23 cooked beef-lard minces, to which varying doses of nitrite salt (range 0–40 g/kg) and sodium ascorbate (range 0–2 g/kg) were added, showed that nitrite salt decreased protein carbonylation up to 3-fold, and inhibited lipid oxidation, demonstrated by up to 4-fold lower levels of ‘thiobarbituric acid reactive substances’, 32-fold lower 4-hydroxynonenal, and 21-fold lower hexanal values. The use of ascorbate increased the antioxidant effect of low nitrite salt levels, whereas it slightly increased protein carbonylation at higher doses of nitrite salt. The addition of a low dose of ascorbate without nitrite salt slightly promoted oxidation during digestion, whereas higher doses had varying antioxidant effects. Second, 40 rats were fed a diet of cooked chicken- or beef-lard minces, either or not cured, for three weeks. Beef, compared to chicken, consumption increased lipid oxidation (2- to 4-fold) during digestion, and gut protein fermentation (cecal iso-butyrate, (iso-)valerate, and fecal indole, cresol), but oxidative stress and inflammation were generally not affected. Cured, compared to fresh, meat consumption significantly increased stomach protein carbonylation (+16%), colonic Ruminococcaceae (2.1-fold) and cecal propionate (+18%), whereas it decreased cecal butyrate (-25%), fecal phenol (-69%) and dimethyl disulfide (-61%) levels. Fecal acetaldehyde and diacetyl levels were increased in beef-fed rats by 2.8-fold and 5.9-fold respectively, and fecal carbon disulfide was 4-fold higher in rats consuming cured beef vs. fresh chicken. Given their known toxicity, the role of acetaldehyde and carbon disulfide in the relation between meat consumption and health should be investigated in future studies.
Protein oxidation is involved in regulatory physiological events as well as in damage to tissues and is thought to play a key role in the pathophysiology of diseases and in the aging process. ...Protein-bound carbonyls represent a marker of global protein oxidation, as they are generated by multiple different reactive oxygen species in blood, tissues and cells. Sample preparation and stabilization are key steps in the accurate quantification of oxidation-related products and examination of physiological/pathological processes. This review therefore focuses on the sample preparation processes used in the most relevant methods to detect protein carbonyls after derivatization with 2,4-dinitrophenylhydrazine with an emphasis on measurement in plasma, cells, organ homogenates, isolated proteins and organelles. Sample preparation, derivatization conditions and protein handling are presented for the spectrophotometric and HPLC method as well as for immunoblotting and ELISA. An extensive overview covering these methods in previously published articles is given for researchers who plan to measure protein carbonyls in different samples.
•A novel DNPH-based method was developed.•The novel method increased protein solubility before DNPH labeling.•More abundant carbonyl labeling compared to traditional the method was obtained.•The ...novel method labeled more carbonyls in soluble as well as insoluble fractions.•Examples of muscle carbonyl levels determined with traditional and novel method.
Protein oxidation is considered an ongoing deteriorative process during storage of fresh and processed meat. Carbonyl compounds have traditionally been detected spectrophotometrically after derivatization with 2,4-dinitrophenylhydrazine (DNPH) to form protein-bound hydrazones with absorbance at 370nm. Here we describe a novel DNPH-based method to quantify protein carbonylation in muscle and meat. The additional steps of the novel method aimed at increasing the protein solubility and inducing protein unfolding before labeling with DNPH. Compared to the traditional method, the new procedure reflected an increased protein carbonylation level measuring overall two to fourfold more carbonyls in muscles from different species as well as in soluble, salt-soluble and insoluble protein fractions. The study suggested that protein unfolding is a more important phenomenon than solubilization for increased DNPH labeling. The novel method resulted in three to fourfold larger carbonyl content determined in chicken, pork and beef (2.8, 3.6 and 3.1nmol/mg of protein, respectively).
Septic shock is a life-threatening dysregulated response to severe infection and is associated with elevated oxidative stress. We aimed to assess protein carbonyls in critically ill patients with ...different sources of sepsis and determine the effect of vitamin C intervention on protein carbonyl concentrations.
Critically ill patients with septic shock (n = 40) were recruited, and sources of sepsis and ICU severity scores were recorded. The patients were randomised to receive either intravenous vitamin C (100 mg/kg body weight/day) or placebo infusions. Blood samples were collected at baseline and daily for up to three days for measurement of cell counts, vitamin C concentrations, protein carbonyls, C-reactive protein, and myeloperoxidase concentrations.
Protein carbonyl concentrations increased 2.2-fold in the cohort over the duration of the study (from 169 to 369 pmol/mg protein; p = 0.03). There were significant correlations between protein carbonyl concentrations and ICU severity scores (APACHE III r = 0.47 and SOFA r = 0.37; p < 0.05) at baseline. At study admission, the patients with pneumonia had nearly 3-fold higher protein carbonyl concentrations relative to the patients with other sources of sepsis (435 vs 157 pmol/mg protein, p < 0.0001). The septic patients had deficient vitamin C status at baseline (9.8 ± 1.4 μmol/L). This increased to 456 ± 90 μmol/L following three days of intravenous vitamin C intervention. Vitamin C intervention did not attenuate the increase in protein carbonyl concentrations.
Circulating protein carbonyls are specifically elevated in critically ill patients with pneumonia relative to other sources of sepsis. The reasons for this are currently unclear and may indicate a mechanism unique to pulmonary sources of sepsis. Intravenous vitamin C administration did not attenuate the increase in protein carbonyls over time.
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•Protein carbonyls increase over time in critically ill patients with septic shock.•Protein carbonyls correlate with ICU severity scores (APACHE III and SOFA).•Patients with pneumonia have 3-fold higher protein carbonyl concentrations.•Intravenous vitamin C administration does not attenuate increase in protein carbonyls.
In the body, nanoparticles can be systemically distributed and then may affect secondary target organs, such as the central nervous system (CNS). Putative adverse effects on the CNS are rarely ...investigated to date. Here, we used a mixed primary cell model consisting mainly of neurons and astrocytes and a minor proportion of oligodendrocytes to analyze the effects of well-characterized 20 and 40 nm silver nanoparticles (SNP). Similar gold nanoparticles served as control and proved inert for all endpoints tested. SNP induced a strong size-dependent cytotoxicity. Additionally, in the low concentration range (up to 10 μg/ml of SNP), the further differentiated cultures were more sensitive to SNP treatment. For detailed studies, we used low/medium dose concentrations (up to 20 μg/ml) and found strong oxidative stress responses. Reactive oxygen species (ROS) were detected along with the formation of protein carbonyls and the induction of heme oxygenase-1. We observed an acute calcium response, which clearly preceded oxidative stress responses. ROS formation was reduced by antioxidants, whereas the calcium response could not be alleviated by antioxidants. Finally, we looked into the responses of neurons and astrocytes separately. Astrocytes were much more vulnerable to SNP treatment compared with neurons. Consistently, SNP were mainly taken up by astrocytes and not by neurons. Immunofluorescence studies of mixed cell cultures indicated stronger effects on astrocyte morphology. Altogether, we can demonstrate strong effects of SNP associated with calcium dysregulation and ROS formation in primary neural cells, which were detectable already at moderate dosages.
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