The first application of terephthalate fluorescence for highly selective detection of hydroxyl radicals in thylakoid membranes

2007 ◽  
Vol 34 (12) ◽  
pp. 1105 ◽  
Author(s):  
Iva Šnyrychová ◽  
Éva Hideg

Possibilities and limitations of the detection of hydroxyl radicals via the conversion of terephthalate (TPA) into the strongly fluorescent hydroxyterephthalate were investigated in order to adapt this method for chlorophyll-containing samples. Using model chemical sources of various reactive oxygen species, we confirmed that TPA detects hydroxyl radicals very sensitively, but is not reactive to either hydrogen peroxide or superoxide radicals. As a new result, we showed that the conversion of TPA to hydroxyterephthalate cannot be induced by singlet oxygen, which may be produced in photosynthetic systems under stress. Until now, the TPA method has not been used in photosynthesis research, so necessary adaptations to minimise the effects of chlorophyll and buffering sugars on hydroxyl radical detection were also explored and optimal conditions for using the method in thylakoid preparations are suggested. Anticipating further plant physiology applications, usefulness of the TPA method was tested in a wider range of pH than reported earlier. To demonstrate that this simple and highly specific method can be used as an alternative approach for the detection of hydroxyl radicals in plant samples, we measured these radicals in isolated thylakoid membranes exposed to 312 nm ultraviolet radiation.

2019 ◽  
Author(s):  
Heta Mattila ◽  
Kumud B. Mishra ◽  
Iiris Kuusisto ◽  
Anamika Mishra ◽  
Kateřina Novotná ◽  
...  

AbstractTo understand the effects of low temperature and cold-acclimation on reactive oxygen species and photoinhibition of photosystem II (PSII), light-induced inactivation of PSII was measured at 22 and 4 °C from four Arabidopsis thaliana accessions (Rschew, Tenela, Columbia-0 and Coimbra) grown under optimal conditions. Photoinhibition was also measured at 4 °C from plants cold-acclimated at 4 °C for two weeks. Measurements were done in the absence and presence of lincomycin that blocks PSII repair, and PSII activity was assayed with the ratio of variable to maximum chlorophyll a fluorescence (FV/FM) and with light-saturated rate of oxygen evolution using a quinone acceptor. Of the non-acclimated accessions, Rschew was the most tolerant to photoinhibition and Coimbra the least; the rate constants of photoinhibition of the most sensitive accession were 1.3-1.9 times as high as those of the tolerant ones. The damaging reaction of photoinhibition in non-acclimated plants was slower or equal at 4 °C than at 22 °C. The rate constants of photoinhibition of cold-acclimated plants, at 4 °C, were 0.55 to 1.25 times as high as those of non-acclimated plants; the protective effect of cold-acclimation on photoinhibition was consistent in Columbia-0 and Coimbra whereas Rschew and Tenela were either slightly more tolerant or susceptible, depending on the method used to assay photoinhibition. Production of singlet oxygen, measured from thylakoid membranes isolated from non-acclimated and cold-acclimated plants, did not decrease due to cold-acclimation, nor did singlet oxygen production correlate with the rate of photoinhibition or with flavonol contents of the leaves.


2012 ◽  
Vol 48 (39) ◽  
pp. 4719 ◽  
Author(s):  
Manoj Kumar ◽  
Naresh Kumar ◽  
Vandana Bhalla ◽  
Parduman Raj Sharma ◽  
Yasrib Qurishi

2020 ◽  
Vol 44 (26) ◽  
pp. 11248-11255
Author(s):  
Vijayesh Kumar ◽  
Abhay Sachdev ◽  
Ishita Matai

A new dimension for the selective detection of short-lived ROS by an electroactive reduced graphene oxide–cerium oxide nanocomposite@cytochrome c hydrogel.


2004 ◽  
Vol 286 (5) ◽  
pp. C1152-C1158 ◽  
Author(s):  
A. McArdle ◽  
J. van der Meulen ◽  
G. L. Close ◽  
D. Pattwell ◽  
H. Van Remmen ◽  
...  

Contractions of skeletal muscles produce increases in concentrations of superoxide anions and activity of hydroxyl radicals in the extracellular space. The sources of these reactive oxygen species are not clear. We tested the hypothesis that, after a demanding isometric contraction protocol, the major source of superoxide and hydroxyl radical activity in the extracellular space of muscles is mitochondrial generation of superoxide anions and that, with a reduction in MnSOD activity, concentration of superoxide anions in the extracellular space is unchanged but concentration of hydroxyl radicals is decreased. For gastrocnemius muscles from adult (6–8 mo old) wild-type ( Sod2+/+) mice and knockout mice heterozygous for the MnSOD gene ( Sod2+/-), concentrations of superoxide anions and hydroxyl radical activity were measured in the extracellular space by microdialysis. A 15-min protocol of 180 isometric contractions induced a rapid, equivalent increase in reduction of cytochrome c as an index of superoxide anion concentrations in the extracellular space of Sod2+/+ and Sod2+/- mice, whereas hydroxyl radical activity measured by formation of 2,3-dihydroxybenzoate from salicylate increased only in the extracellular space of muscles of Sod2+/+ mice. The lack of a difference in increase in superoxide anion concentration in the extracellular space of Sod2+/+ and Sod2+/- mice after the contraction protocol supported the hypothesis that superoxide anions were not directly derived from mitochondria. In contrast, the data obtained suggest that the increase in hydroxyl radical concentration in the extracellular space of muscles from wild-type mice after the contraction protocol most likely results from degradation of hydrogen peroxide generated by MnSOD activity.


2016 ◽  
Vol 23 (12) ◽  
pp. 926-933 ◽  
Author(s):  
Siroon Bekkering ◽  
Bastiaan A. Blok ◽  
Leo A. B. Joosten ◽  
Niels P. Riksen ◽  
Reinout van Crevel ◽  
...  

ABSTRACTInnate immune memory, or trained immunity, has recently been described to be an important property of cells of the innate immune system. Due to the increased interest in this important new field of immunological investigation, we sought to determine the optimal conditions for anin vitroexperimental protocol of monocyte training using three of the most commonly used training stimuli from the literature: β-glucan, the bacillus Calmette-Guérin (BCG) vaccine, and oxidized low-density lipoprotein (oxLDL). We investigated and optimized a protocol of monocyte trained immunity induced by an initial training period with β-glucan, BCG, or oxLDL, followed by washing and resting of the cells and, thereafter, restimulation with secondary bacterial stimuli. The training and resting time intervals were varied to identify the optimal setting for the long-term induction of trained immunity. Trained immunity was assessed in terms of the secondary cytokine response, the production of reactive oxygen species, cell morphology, and induction of glycolysis. Monocytes primed with β-glucan, BCG, and oxLDL showed increased pro- and anti-inflammatory cytokine responses upon restimulation with nonrelated stimuli. Also, all three stimuli induced a switch to glycolysis (the Warburg effect). These effects were most pronounced when the training interval was 24 h and the resting time interval was 6 days. Training with BCG and oxLDL also led to the increased production of reactive oxygen species, whereas training with β-glucan led to the decreased production of reactive oxygen species. We describe the optimal conditions for anin vitroexperimental model with human primary monocytes for study of the induction of trained innate immunity by microbial and metabolic stimuli.


2010 ◽  
Vol 665 (1) ◽  
pp. 74-78 ◽  
Author(s):  
Shinya Yamaguchi ◽  
Naoya Kishikawa ◽  
Kaname Ohyama ◽  
Yoshihito Ohba ◽  
Maiko Kohno ◽  
...  

2015 ◽  
Vol 1130 ◽  
pp. 118-122 ◽  
Author(s):  
Sören Bellenberg ◽  
Dieu Huynh ◽  
Laura Castro ◽  
Maria Boretska ◽  
Wolfgang Sand ◽  
...  

Reactive oxygen species (ROS), such as hydrogen peroxide (H2O2), superoxide (O2-) and hydroxyl radicals (OH.) are known to be formed on the surface of metal sulfides in aqueous solution under oxic and anoxic conditions. Consequently bacteria which have not been adapted to their presence are metabolically inhibited [1], presumably due to the presence of these ROS. Pyrite-grown cells ofAcidithiobacillus ferrooxidansT, in contrast to iron (II)-grown cells, were able to oxidize iron (II)-ions or pyrite after 24 h starvation and contact with 1 mM externally added H2O2. In this study, similar results were obtained withAcidiferrobactersp. SPIII/3. However,Acidithiobacillus ferrivoransSS3 showed the highest tolerance towards contact with H2O2, whileLeptospirillum ferrooxidansDSM 2391 was most sensitive. Similar results were obtained after exposure to defined doses of gamma radiation, which cleaves water molecules and generates ROS. In this study members of the three aforementioned genera of mineral-oxidizing bacteria were compared regarding their ability to survive, colonize pyrite and to oxidize iron (II)-ions after exposure to different concentrations of H2O2. Pyrite colonization was studied after exposure to endogenous ROS formed on pyrite or after external addition of H2O2using confocal laser scanning microscopy (CLSM).


2003 ◽  
Vol 69 (11) ◽  
pp. 6500-6506 ◽  
Author(s):  
Paula A. Belinky ◽  
Nufar Flikshtein ◽  
Sergey Lechenko ◽  
Shimon Gepstein ◽  
Carlos G. Dosoretz

ABSTRACT We studied oxidative stress in lignin peroxidase (LIP)-producing cultures (cultures flushed with pure O2) of Phanerochaete chrysosporium by comparing levels of reactive oxygen species (ROS), cumulative oxidative damage, and antioxidant enzymes with those found in non-LIP-producing cultures (cultures grown with free exchange of atmospheric air [control cultures]). A significant increase in the intracellular peroxide concentration and the degree of oxidative damage to macromolecules, e.g., DNA, lipids, and proteins, was observed when the fungus was exposed to pure O2 gas. The specific activities of manganese superoxide dismutase, catalase, glutathione reductase, and glutathione peroxidase and the consumption of glutathione were all higher in cultures exposed to pure O2 (oxygenated cultures) than in cultures grown with atmospheric air. Significantly higher gene expression of the LIP-H2 isozyme occurred in the oxygenated cultures. A hydroxyl radical scavenger, dimethyl sulfoxide (50 mM), added to the culture every 12 h, completely abolished LIP expression at the mRNA and protein levels. This effect was confirmed by in situ generation of hydroxyl radicals via the Fenton reaction, which significantly enhanced LIP expression. The level of intracellular cyclic AMP (cAMP) was correlated with the starvation conditions regardless of the oxygenation regimen applied, and similar cAMP levels were obtained at high O2 concentrations and in cultures grown with atmospheric air. These results suggest that even though cAMP is a prerequisite for LIP expression, high levels of ROS, preferentially hydroxyl radicals, are required to trigger LIP synthesis. Thus, the induction of LIP expression by O2 is at least partially mediated by the intracellular ROS.


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