Protein carbonyl determination by a rhodamine B hydrazide-based fluorometric assay

Georgiou, Christos D.; Zisimopoulos, Dimitrios; Argyropoulou, Vasiliki; Kalaitzopoulou, Electra; Ioannou, Panayiotis V.; Salachas, George; Grune, Tilman

doi:10.1016/j.redox.2018.04.017

Cited by 14 publications

(10 citation statements)

References 51 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…L=O can be quantified by a method under development by our lab. Extending to apoB100 oxidative modifications, of particular interest are the following: the OS-induced formation of dityrosines ( 89 ), and disulfides (between two Cys; -S-S-) ( 90 ), hydroperoxides (apoB100-OOH) ( 50 ), and carbonyls (apoB100-C=O) ( 91 , 92 ), the last three by methods developed by our lab. As illustrated in (right) panel, macrophages recognize and bind oxLDL with several scavenger receptors (SR, e.g., SR-A1, CD36, and LOX-1).…”

Section: Introductionmentioning

confidence: 99%

Methods on LDL particle isolation, characterization, and component fractionation for the development of novel specific oxidized LDL status markers for atherosclerotic disease risk assessment

et al. 2023

Self Cite

View full text Add to dashboard Cite

The present study uses simple, innovative methods to isolate, characterize and fractionate LDL in its main components for the study of specific oxidations on them that characterize oxidized low-density lipoprotein (oxLDL) status, as it causatively relates to atherosclerosis-associated cardiovascular disease (CVD) risk assessment. These methods are: (a) A simple, relatively time-short, low cost protocol for LDL isolation, to avoid shortcomings of the currently employed ultracentrifugation and affinity chromatography methodologies. (b) LDL purity verification by apoB100 SDS-PAGE analysis and by LDL particle size determination; the latter and its serum concentration are determined in the present study by a simple method more clinically feasible as marker of CVD risk assessment than nuclear magnetic resonance. (c) A protocol for LDL fractionation, for the first time, into its main protein/lipid components (apoB100, phospholipids, triglycerides, free cholesterol, and cholesteryl esters), as well as into LDL carotenoid/tocopherol content. (d) Protocols for the measurement, for the first time, of indicative specific LDL component oxidative modifications (cholesteryl ester-OOH, triglyceride-OOH, free cholesterol-OOH, phospholipid-OOH, apoB100-MDA, and apoB100-DiTyr) out of the many (known/unknown/under development) that collectively define oxLDL status, which contrasts with the current non-specific oxLDL status evaluation methods. The indicative oxLDL status markers, selected in the present study on the basis of expressing early oxidative stress-induced oxidative effects on LDL, are studied for the first time on patients with end stage kidney disease on maintenance hemodialysis, selected as an indicative model for atherosclerosis associated diseases. Isolating LDL and fractionating its protein and main lipid components, as well as its antioxidant arsenal comprised of carotenoids and tocopherols, paves the way for future studies to investigate all possible oxidative modifications responsible for turning LDL to oxLDL in association to their possible escaping from LDL’s internal antioxidant defense. This can lead to studies to identify those oxidative modifications of oxLDL (after their artificial generation on LDL), which are recognized by macrophages and convert them to foam cells, known to be responsible for the formation of atherosclerotic plaques that lead to the various CVDs.

show abstract

Section: Introductionmentioning

confidence: 99%

Methods on LDL particle isolation, characterization, and component fractionation for the development of novel specific oxidized LDL status markers for atherosclerotic disease risk assessment

et al. 2023

Self Cite

View full text Add to dashboard Cite

show abstract

“…It is also evident that the latter procedure is more notable in the hemisphere while cerebellum and brainstem only show late alterations at the 14-day post ligation group. This profile level difference between the OS markers PrMDA and PrC = O, may be attributed to the fact that MDA is produced by phospholipids that in a time fashion can peroxidize each other, as being in proximity in membrane structures especially in lipid-rich biological tissues such as brain, while protein carbonylation is considered as a slower phenomenon [ 13 ].The high sensitivity of the RBH fluorometric assay which was used for the PrC = O quantification, allowing detection limit as low as 0.4 pmol [ 14 ], confirms the start of the proteins oxidation even at 12 h post CBD ligation when compared to the control/sham group.…”

Section: Discussionmentioning

confidence: 99%

“…Protein carbonyls (PrC = O) are measured by the specific rhodamine B hydrazide (RBH)-based fluorometric assay, described elsewhere [ 14 ]. PrC = O result from the free radical-generated carbonyl groups (–C = O) on them.…”

Section: Methodsmentioning

confidence: 99%

Time progression and regional expression of brain oxidative stress induced by obstructive jaundice in rats

et al. 2022

Self Cite

View full text Add to dashboard Cite

Background Obstructive jaundice induces oxidative changes in the brain parenchyma and plays significant role in clinical manifestations of hepatic encephalopathy. We aim to study the progression of the brain oxidative status over time and the differences of its pattern over the hemispheres, the brainstem and the cerebellum. We use an experimental model in rats and measuring the oxidative stress (OS) specific biomarkers protein malondialdehyde (PrMDA) and protein carbonyls (PrC = O). Results Hyperbilirubinemia has been confirmed in all study groups as the result of common bile duct obstruction. We confirmed increase in both PrMDA and PrC = O biomarkers levels with different type of changes over time. We also confirmed that the oxidative process develops differently in each of the brain areas in study. Conclusions The present study confirms the progressive increase in OS in all brain areas studied using markers indicative of cumulative protein modification.

show abstract

“…Carbonyl groups can also be detected by hydrazides coupled with fluorescent labels [53]. Fluorescent hydrazides have been used for qualitative protein carbonyl assessment such as microscopy imaging but also gel-based proteomics [54].…”

Section: Introductionmentioning

confidence: 99%

Enhanced Carbonylation of Photosynthetic and Glycolytic Proteins in Antibiotic Timentin-Treated Tobacco In Vitro Shoot Culture

Andriūnaitė

Rugienius

Tamošiūnė

et al. 2022

Plants

View full text Add to dashboard Cite

Antibiotics are used in plant in vitro tissue culture to eliminate microbial contamination or for selection in genetic transformation. Antibiotic timentin has a relatively low cytotoxic effect on plant tissue culture; however, it could induce an enduring growth-inhibiting effect in tobacco in vitro shoot culture that persists after tissue transfer to a medium without antibiotic. The effect is associated with an increase in oxidative stress injury in plant tissues. In this study, we assessed changes of reactive oxygen species accumulation, protein expression, and oxidative protein modification response associated with enduring timentin treatment-induced growth suppression in tobacco (Nicotiana tabacum L.) in vitro shoot culture. The study revealed a gradual 1.7 and 1.9-fold increase in superoxide (O2•−) content at the later phase of the propagation cycle for treatment control (TC) and post-antibiotic treatment (PA) shoots; however, the O2•− accumulation pattern was different. For PA shoots, the increase in O2•− concentration occurred several days earlier, resulting in 1.2 to 1.4-fold higher O2•− concentration compared to TC during the period following the first week of cultivation. Although no protein expression differences were detectable between the TC and PA shoots by two-dimensional electrophoresis, the increase in O2•− concentration in PA shoots was associated with a 1.5-fold increase in protein carbonyl modification content after one week of cultivation, and protein carbonylation analysis revealed differential modification of 26 proteoforms involved in the biological processes of photosynthesis and glycolysis. The results imply that the timentin treatment-induced oxidative stress might be implicated in nontranslational cellular redox balance regulation, accelerates the development of senescence of the shoot culture, and contributes to the shoot growth-suppressing effect of antibiotic treatment.

show abstract

Protein carbonyl determination by a rhodamine B hydrazide-based fluorometric assay

Cited by 14 publications

References 51 publications

Methods on LDL particle isolation, characterization, and component fractionation for the development of novel specific oxidized LDL status markers for atherosclerotic disease risk assessment

Methods on LDL particle isolation, characterization, and component fractionation for the development of novel specific oxidized LDL status markers for atherosclerotic disease risk assessment

Time progression and regional expression of brain oxidative stress induced by obstructive jaundice in rats

Enhanced Carbonylation of Photosynthetic and Glycolytic Proteins in Antibiotic Timentin-Treated Tobacco In Vitro Shoot Culture

Contact Info

Product

Resources

About