Reactive oxygen species play a key role in cellular homeostasis and redox signaling at physiological levels, where excessive production affects the function and integrity of macromolecules, specifically proteins. Therefore, it is important to define radical-mediated proteotoxic stress in macrophages and identify target protein to prevent tissue dysfunction. A well employed, THP-1 cell line was utilized as in vitro model to study immune response and herein we employ immuno-spin trapping technique to investigate radical-mediated protein oxidation in macrophages. Hydroxyl radical formation along macrophage differentiation was confirmed by electron paramagnetic resonance along with confocal laser scanning microscopy using hydroxyphenyl fluorescein. Lipid peroxidation product, malondialdehyde, generated under experimental conditions as detected using swallow-tailed perylene derivative fluorescence observed by confocal laser scanning microscopy and high-performance liquid chromatography, respectively. The results obtained from this study warrant further corroboration and study of specific proteins involved in the macrophage activation and their role in inflammations.
Singlet oxygen (1O2) is formed by triplet-triplet energy transfer from triplet chlorophyll to O2 via Type II photosensitization reaction in photosystem II (PSII). Formation of triplet chlorophyll is associated with the change in spin state of the excited electron and recombination of triplet radical pair in the PSII antenna complex and reaction center, respectively. Here, we have provided evidence for the formation of 1O2 by decomposition of protein hydroperoxide in PSII membranes deprived of Mn4O5Ca complex. Protein hydroperoxide is formed by protein oxidation initiated by highly oxidizing chlorophyll cation radical and hydroxyl radical formed by Type I photosensitization reaction. Under highly oxidizing conditions, protein hydroperoxide is oxidized to protein peroxyl radical which either cyclizes to dioxetane or recombines with another protein peroxyl radical to tetroxide. These highly unstable intermediates decompose to triplet carbonyls which transfer energy to O2 forming 1O2. Data presented in this study show for the first time that 1O2 is formed by decomposition of protein hydroperoxide in PSII membranes deprived of Mn4O5Ca complex.
- MeSH
- Chlorophyll metabolism MeSH
- Electron Spin Resonance Spectroscopy methods MeSH
- Photosystem II Protein Complex metabolism MeSH
- Oxygen metabolism MeSH
- Oxidation-Reduction MeSH
- Hydrogen Peroxide metabolism MeSH
- Peroxides metabolism MeSH
- Energy Transfer physiology MeSH
- Singlet Oxygen metabolism MeSH
- Light MeSH
- Light-Harvesting Protein Complexes metabolism MeSH
- Publication type
- Journal Article MeSH
Effect of UV irradiation on free radicals in different types of melanins and melanin complexes with diamagnetic Cd(II) and paramagnetic Cu(II) was examined by the use of electron paramagnetic resonance (EPR) spectroscopy. The aim of this studies was to compare o-semiquinone free radicals formation in two model eumelanins synthesized from 3,4-dihydroxyphenylalanine (DOPA) and tyrosine in the presence of tyrosinase, and in synthetic pheomelanin, under exposition on ultraviolet, because of the important role of free radicals and melanins in human organism. UV may change free radical concentrations in melanin. Changes in EPR spectra of DOPA-melanin-Cd(II) and DOPA-melanin-Cu(II) complexes after UV irradiation were determined. Diamagnetic Cd(II) strongly increased free radical concentrations in DOPA-melanin. UV irradiation during 30 and 60 min slightly increased and decreased free radical concentrations in DOPA-melanin-Cd(II) complexes, respectively. Paramagnetic Cu(II) quenched free radical lines of DOPA-melanin, and only the Cu(II) signals were detected for both UV-irradiated and nonirradiated samples. Free radical concentration in both eumelanins increased after UV irradiation, but it decreased in irradiated pheomelanin. EPR spectra of free radicals in the studied samples were homogeneously broadened. Slow spin-lattice relaxation processes exist in all the examined melanins and DOPA-melanin-Cd(II) complexes. Fast spin-lattice relaxation processes characterized Cu(II) in DOPA-melanin-Cu(II) complexes.
- MeSH
- Dihydroxyphenylalanine radiation effects MeSH
- Electron Spin Resonance Spectroscopy * methods utilization MeSH
- Epidermis immunology radiation effects MeSH
- Humans MeSH
- Melanins * isolation & purification radiation effects MeSH
- Melanocytes immunology radiation effects MeSH
- Statistics as Topic MeSH
- Monophenol Monooxygenase biosynthesis radiation effects MeSH
- Ultraviolet Rays * diagnostic use adverse effects MeSH
- Free Radicals isolation & purification adverse effects MeSH
- Check Tag
- Humans MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
Gamma- and neutron doses in an experimental reactor were measured using alanine/electron spin resonance (ESR) spectrometry. The absorbed dose in alanine was decomposed into contributions caused by gamma and neutron radiation using neutron kerma factors. To overcome a low sensitivity of the alanine/ESR response to thermal neutrons, a novel method has been proposed for the assessment of a thermal neutron flux using the (14)N(n,p) (14)C reaction on nitrogen present in alanine and subsequent measurement of (14)C by liquid scintillation counting (LSC).
- MeSH
- Alanine chemistry radiation effects MeSH
- Electron Spin Resonance Spectroscopy methods statistics & numerical data MeSH
- Nuclear Power Plants standards MeSH
- Neutrons * MeSH
- Carbon Radioisotopes analysis MeSH
- Radiometry methods statistics & numerical data MeSH
- Scintillation Counting methods statistics & numerical data MeSH
- Dose-Response Relationship, Radiation MeSH
- Gamma Rays * MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Magnetická rezonance (MR) je dlouho považována za slibnou zobrazovací metodu k posouzení lokalizace a lokálního stagingu karcinomu prostaty (KP). Cílem práce je prezentovat moderní multiparametickou MR a ostatní zobrazovací metody, které by mohli zlepšit detekci a lokalizaci KP. MR prostaty byla zavedena do klinické praxe v polovině osmdesátých let 20. století. Od té doby prošla značným vývojem až k nynější multiparametické MR prostaty, která zahrnuje kombinaci anatomického zobrazení a funkčních a metabolických informací. MR je přínosem v diagnostice karcinomu prostaty zejména v případě suspektního klinického či laboratorního nálezu, který není potvrzen předchozí biopsií prostaty. K ostatní moderním zobrazovacím metodám prostaty patří transrektální powerDoppler sonografie s 3D rekonstrukcí (3D-PDS) , transrektální sonografie s podáním echokontrastní látky (Contrast-Enhanced Ultrasound - CEUS), HistoScanning? či elastografie.
Magnetic resonance (MR) has long been considered a promising imaging method to assess the localization and local staging of prostate cancer (CP). The aim of this study is to present modern multiparametic MR and other imaging modalities that could lead to improvement in detection and localization of CP. MR of the prostate was introduced into clinical practice in the mid- 1980s. Since then, there have been significant developments to the present multiparametric MR, which includes a combination of anatomical and functional displays as well as metabolic information. MR is helpful in the diagnosis of prostate cancer, particularly in the cases of suspected clinical or laboratory finding which have not been confirmed by prostate biopsies. The other modern imaging methods include prostate transrectal power Doppler sonography with 3D reconstruction (3D-PDS ), transrectal sonography with echocontrast substances (Contrast - Enhanced Ultrasound, CEUS) HistoScanning? or elastography Currently, multiparametric MR is considered to be the optimal preoperative imaging method for evaluating localization and local staging of prostate cancer.
- MeSH
- Diagnostic Imaging * methods MeSH
- Elasticity Imaging Techniques methods MeSH
- Electron Spin Resonance Spectroscopy contraindications methods MeSH
- Contrast Media diagnostic use MeSH
- Humans MeSH
- Magnetic Resonance Imaging, Cine methods MeSH
- Magnetic Resonance Spectroscopy methods MeSH
- Bone Neoplasms secondary MeSH
- Prostatic Neoplasms * radiography ultrasonography MeSH
- Signal-To-Noise Ratio MeSH
- Positron-Emission Tomography methods MeSH
- Radionuclide Imaging methods MeSH
- Ultrasonography methods MeSH
- Imaging, Three-Dimensional MeSH
- Check Tag
- Humans MeSH
- Publication type
- Review MeSH
Melanins are polymorphous and multifunctional biopolymers with a relatively high concentration of free radicals. EPR spectroscopy was used to study o-semiquinone free radicals in model eumelanins synthesized from 3,4-dihydroxyphenylalanine (DOPA) and tyrosine in the presence of tyrosinase, and melanins isolated from A-375 and G-361 human melanoma malignum cells exposed to two compounds: 5,7-dimethoxycoumarin (DMC) and valproic acid (VPA). Changes were determined in the concentrations of free radicals in the individual melanins from tumour cells treated with DMC and VPA. A strong decrease in the concentrations of free radicals characterizes melanins isolated from tumour cells treated together with DMC and VPA. Slow spin-lattice relaxation processes were noted in the melanins tested with homogeneous broadened EPR spectra. The EPR technique may be useful not only for the elucidation of free radicals in melanins from A-375 and G-361 cells treated with VPA and DMC but it could also be applied to establish the relationship between melanin type and the malignancy of melanoma malignum.
- Keywords
- 5,7-dimethoxycoumarin,
- MeSH
- Biopolymers analysis MeSH
- Electron Spin Resonance Spectroscopy * methods statistics & numerical data utilization MeSH
- Coumarins pharmacology MeSH
- Valproic Acid pharmacology MeSH
- Humans MeSH
- Melanins * analysis MeSH
- Melanoma MeSH
- Cell Line, Tumor cytology drug effects MeSH
- Free Radicals * analysis MeSH
- Check Tag
- Humans MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
When photosystem II (PSII) is exposed to excess light, singlet oxygen ((1)O(2)) formed by the interaction of molecular oxygen with triplet chlorophyll. Triplet chlorophyll is formed by the charge recombination of triplet radical pair (3)[P680(•+)Pheo(•-)] in the acceptor-side photoinhibition of PSII. Here, we provide evidence on the formation of (1)O(2) in the donor side photoinhibition of PSII. Light-induced (1)O(2) production in Tris-treated PSII membranes was studied by electron paramagnetic resonance (EPR) spin-trapping spectroscopy, as monitored by TEMPONE EPR signal. Light-induced formation of carbon-centered radicals (R(•)) was observed by POBN-R adduct EPR signal. Increased oxidation of organic molecules at high pH enhanced the formation of TEMPONE and POBN-R adduct EPR signals in Tris-treated PSII membranes. Interestingly, the scavenging of R(•) by propyl gallate significantly suppressed (1)O(2). Based on our results, it is concluded that (1)O(2) formation correlates with R(•) formation on the donor side of PSII due to oxidation of organic molecules (lipids and proteins) by long-lived P680(•+)/TyrZ(•). It is proposed here that the Russell mechanism for the recombination of two peroxyl radicals formed by the interaction of R(•) with molecular oxygen is a plausible mechanism for (1)O(2) formation in the donor side photoinhibition of PSII.
- MeSH
- Models, Chemical MeSH
- Electron Spin Resonance Spectroscopy methods MeSH
- Photochemistry methods MeSH
- Photosystem II Protein Complex physiology MeSH
- Hydrogen-Ion Concentration MeSH
- Oxygen chemistry MeSH
- Propyl Gallate chemistry MeSH
- Singlet Oxygen * MeSH
- Spin Trapping methods MeSH
- Spinacia oleracea MeSH
- Light MeSH
- Synechococcus metabolism MeSH
- Carbon chemistry MeSH
- Free Radicals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Hydroxyl radical (HO•) production in photosystem II (PSII) was studied by electron paramagnetic resonance (EPR) spin-trapping technique. It is demonstrated here that the exposure of PSII membranes to heat stress (40 °C) results in HO• formation, as monitored by the formation of EMPO-OH adduct EPR signal. The presence of different exogenous halides significantly suppressed the EMPO-OH adduct EPR signal in PSII membranes under heat stress. The addition of exogenous acetate and blocker of chloride channel suppressed the EMPO-OH adduct EPR signal, whereas the blocker of calcium channel did not affect the EMPO-OH adduct EPR signal. Heat-induced hydrogen peroxide (H₂O₂) production was studied by amplex red fluorescent assay. The presence of exogenous halides, acetate and chloride blocker showed the suppression of H₂O₂ production in PSII membranes under heat stress. Based on our results, it is proposed that the formation of HO• under heat stress is linked to uncontrolled accessibility of water to the water-splitting manganese complex caused by the release of chloride ion on the electron donor side of PSII. Uncontrolled water accessibility to the water-splitting manganese complex causes the formation of H₂O₂ due to improper water oxidation, which leads to the formation of HO• via the Fenton reaction under heat stress.
- MeSH
- Chlorides metabolism MeSH
- Electron Spin Resonance Spectroscopy methods MeSH
- Photosystem II Protein Complex chemistry metabolism MeSH
- Hydroxyl Radical metabolism MeSH
- Oxidation-Reduction MeSH
- Hydrogen Peroxide pharmacology MeSH
- Spin Trapping methods MeSH
- Spinacia oleracea chemistry MeSH
- Heating MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Pressurized hot water extraction (PHWE) was employed to prepare extracts from dried grape skin of two wine grape varieties (St. Laurent and Alibernet) at various temperatures (from 40 up to 120°C) and amounts of sample (0.5, 1.0 and 1.5 g). To assess the antioxidant activity of the extracts, electron paramagnetic resonance (EPR) spectroscopy was applied involving DPPH and ABTS(+) assays. Other extract characteristics including HPLC profile of anthocyanins and total phenolic compound content were obtained as well. PHWE has also been compared with earlier results of extractions of the same grape skin samples with compressed methanol and compressed ethanol under the conditions of pressurized fluid extraction (PFE). From this comparison, PHWE emerges as the more benign and efficient extraction method to recover valuable phenolic antioxidants from grape skins for the prospective use in functional food supplements.
- MeSH
- Antioxidants chemistry MeSH
- Chemical Fractionation methods MeSH
- Electron Spin Resonance Spectroscopy methods MeSH
- Plant Epidermis chemistry MeSH
- Plant Extracts chemistry MeSH
- Spectrophotometry, Ultraviolet MeSH
- Pressure MeSH
- Vitis chemistry MeSH
- Water chemistry MeSH
- Hot Temperature MeSH
- Chromatography, High Pressure Liquid MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Electron paramagnetic resonance (EPR) spin trapping spectroscopy is an important method used in free radical research; however, its application in biological systems is hindered by EPR silencing of spin adducts. Previous studies in superoxide-generating chemical systems have shown that spin adducts can be partially stabilized by cyclodextrins. In this work, for the first time, this proposed protective effect of cyclodextrins is investigated in a real biological sample-in isolated thylakoid membranes and photosystem II (PSII) particles with EMPO as a spin trap. It is shown that (i) randomly methylated beta-cyclodextrin and 2-hydroxypropyl-beta-cyclodextrin form inclusion complexes with EMPO-superoxide adducts (EMPO-OOH), (ii) both cyclodextrins increase the intensity of the EMPO-OOH EPR signal in PSII particles up to five times, (iii) higher EMPO-OOH EPR signal intensity is a result of increased stability of EMPO-OOH, and (iv) the extent of the protection of EMPO-OOH adduct provided by cyclodextrins is different in thylakoids and PSII particles. Along with the spin trapping data, the toxicity of cyclodextrins is also discussed with particular focus on photosynthetic preparations. The presented data show that both tested cyclodextrins can be used as valuable tools to improve the sensitivity of spin trapping in biological samples.
- MeSH
- beta-Cyclodextrins pharmacology MeSH
- Cyclodextrins pharmacology MeSH
- Electron Spin Resonance Spectroscopy methods MeSH
- Photosystem II Protein Complex chemistry isolation & purification metabolism MeSH
- Pyrroles MeSH
- Sensitivity and Specificity MeSH
- Spin Trapping MeSH
- Spinacia oleracea MeSH
- Superoxides chemistry metabolism MeSH
- In Vitro Techniques MeSH
- Thylakoids chemistry drug effects metabolism MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
