31P NMR spectroscopy and polarographic combined study of erythrocytes treated with 5-fluorouracil: cardiotoxicity-related changes in ATP, 2,3-BPG, and O2 metabolism.

Antineoplastic drug 5-fluorouracil (5-FU) frequently shows cardiotoxic effects, the mechanism of which has not yet been elucidated. The objective of the present study was to explore effects of 5-FU on metabolism of ATP, 2,3-BPG, and oxygen in erythrocytes and to relate these to the phenomenon of 5-FU cardiotoxicity. We determined that 5-FU induced rapid increase in O(2) consumption, which led to drastic changes in the metabolism of phosphate compounds in erythrocytes. Decrease in pO(2) provoked increase in production of 2,3-BPG and subsequent deoxygenation of oxyHb to deoxyHb. However, the most important effect of 5-FU on erythrocytes is severe decrease in the level of ATP. This could lead to a number of irreversible changes in erythrocyte structure and functioning, such as echinocytosis, increase in membrane fluidity, and non-functioning of membrane ion pumps. All these changes affect normal functioning of erythrocytes, leading to difficulties in oxygen transport and insufficient supply of oxygen to the heart, and pointing to the importance of studying the effects of antineoplastic drugs on intracellular metabolism of erythrocytes.

Detection of oxygen-centered radicals using EPR spin-trap DEPMPO: the effect of oxygen.

Studies of the ability of the EPR spin trap DEPMPO to detect both superoxide and hydroxyl radicals produced by systems in vitro and in vivo are presented. Experiments using free radical-generating systems confirmed the suitability of the EPR spin trapping technique but also revealed the existence of an undesirable conversion of DEPMPO/OOH into DEPMPO/OH adducts. The rate of conversion decreases with oxygenation, and the production of oxygen-centered radicals increases. However, this property of DEPMPO does not have a significant influence on its ability to independently detect radicals produced by plant plasma membranes. Since the adduct conversion appears to be rather slow compared to radical generation, we conclude that the DEPMPO spin-trap can be efficiently used for detection of oxygen-centered radicals produced by systems in vivo, as demonstrated for isolated plasma membranes.

Myopathy, muscle atrophy and tongue lipid composition in MuSK myasthenia gravis.

Myasthenia gravis (MG) associated with anti-muscle-specific tyrosine kinase (MuSK) antibodies differs in many aspects from typical presentation of acetylcholine receptor (AChR)-positive MG. Myopathy and muscle atrophy are observed in MuSK-positive MG patients, unlike AChR-positive patients with MG. That is why the aim of this study was to assess the presence of myopathy and muscle atrophy as well as the tongue lipid composition in our cohort of MuSK-positive MG patients. Clinical examination, electromyography (EMG) and proton magnetic resonance spectroscopy were performed in 31 MuSK-positive and 28 AChR-positive MG patients. Myopathic EMG was more frequent in MuSK compared to AChR MG patients. In AChR MG patients, myopathic EMG in facial muscles was more frequent after long-term corticosteroid treatment, which was not the case with MuSK-positive MG patients. Facial and/or tongue muscle atrophy was registered in 23 % of MuSK MG patients. Longer disease duration was observed in patients with clinical signs of tongue and/or facial muscle atrophy compared to those with normal tongue muscle. Intramyocellular lipid deposition in the tongue was present in 85.2 % of MuSK and 20 % of AChR MG patients. Female MuSK MG patients had more frequently electrophysiological signs of myopathy on the facial muscles and signs of intramyocellular lipid deposition in the tongue, compared to male patients with MuSK-positive MG. Myopathy, muscle atrophy and intramyocellular lipid deposition in the tongue are more frequent in MuSK-positive compared to AChR-positive MG patients.

Binding of doxyl stearic spin labels to human serum albumin: an EPR study.

The binding of spin-labeled fatty acids (SLFAs) to the human serum albumin (HSA) examined by electron paramagnetic resonance (EPR) spectroscopy was studied to evaluate the potential of the HSA/SLFA/EPR technique as a biomarking tool for cancer. A comparative study was performed on two spin labels with nitroxide groups attached at opposite ends of the fatty acid (FA) chain, 5-doxyl stearic (5-DS) and 16-doxyl stearic (16-DS) acid. The effects of incubation time, different [SLFA]/[HSA] molar ratios, ethanol, and temperature showed that the position of the nitroxide group produces certain differences in binding between the two SLFAs. Spectra for different [SLFA]/[HSA] molar ratios were decomposed into two spectral components, which correspond to the weakly and strongly bound SLFAs. The reduction of SLFA with ascorbate showed the existence of a two component process, fast and slow, confirming the decomposition results. Warfarin has no effect on the binding of the two SLFAs, whereas ibuprofen significantly decreases the binding of 5-DS and has no effect on 16-DS. Together, the results of this study indicate that both SLFAs, 5-DS and 16-DS, should be used for the study of HSA conformational changes in blood induced by various medical conditions.

Oxygen radicals produced by plant plasma membranes: an EPR spin-trap study.

Plant plasma membranes are known to produce superoxide radicals, while the production of the hydroxyl radical, previously detected in complex plant tissues, is thought to occur in the cell wall. The mechanism of production of superoxide radicals by plant plasma membranes is, however, under dispute. It is shown, using electron paramagnetic resonance spectroscopy with a 5-diethoxyphosphoryl-5-methyl-1-pyrroline N-oxide spin-trap capable of differentiating between radical species, that isolated purified plasma membranes from maize roots produce hydroxyl radicals besides superoxide radicals. The results argue in favour of superoxide production through an oxygen and diphenylene iodonium-sensitive, NADH-dependent superoxide synthase mechanism, as well as through other unidentified mechanism(s). The hydroxyl radical is produced by an oxygen-insensitive, NADH-stimulated mechanism, which is enhanced in membranes in which the superoxide synthase is incapacitated by substrate removal or inhibition.