Levels of F2-isoprostanes,F4-neuroprostanes,and total nitrate/nitrite in plasma and cerebrospinal fluid of patients with traumatic brain injury

Several events occurring during the secondary damage of traumatic brain injury (TBI) can cause oxidative stress. F₂-isoprostanes (F₂-IsoPs) and F₄-neuroprostanes (F₄-NPs) are specific lipid peroxidation markers generated from arachidonic acid and docosahexaenoic acid, respectively. In this study, we evaluated oxidative stress in patients with moderate and severe TBI. Since sedatives are routinely used to treat TBI patients and propofol has been considered an antioxidant, TBI patients were randomly treated with propofol or midazolam for 72 h postoperation. We postoperatively collected cerebrospinal fluid (CSF) and plasma from 15 TBI patients for 6–10 d and a single specimen of CSF or plasma from 11 controls. Compared with the controls, the TBI patients exhibited elevated levels of F₂-IsoPs and F₄-NPs in CSF throughout the postsurgery period regardless of the sedative used. Compared with the group of patients who received midazolam, those who received propofol exhibited markedly augmented levels of plasma F₂-IsoPs, which were associated with higher F₄-NPs levels and lower total nitrate/nitrite levels in CSF early in the postsurgery period. Furthermore, the higher CSF F₂-IsoPs levels correlated with 6-month and 12-month worse outcomes, which were graded according to the Glasgow Outcome Scale. The results demonstrate enhanced oxidative damage in the brain of TBI patients and the association of higher CSF levels of F₂-IsoPs with a poor outcome. Moreover, propofol treatment might promote lipid peroxidation in the circulation, despite possibly suppressing nitric oxide or peroxynitrite levels in CSF, because of the increased loading of the lipid components from the propofol infusion.     

F2-Isoprostanes in HDL are bound to neutral lipids and phospholipids

Low HDL cholesterol (HDL-C) is a risk factor for coronary artery disease (CAD). However, interventions that raise HDL-C have failed to reduce cardiovascular events. We previously reported that HDL is the main carrier of plasma F₂-isoprostanes (F₂-IsoPs) that are markers of oxidative stress formed upon oxidation of arachidonic acid. F₂-IsoPs are predominantly associated with phospholipids. However, there is evidence that F₂-IsoPs in the liver of rats treated with carbon tetrachloride associate with the neutral lipids. To date it is not known whether F₂-IsoPs are found in the neutral lipids in HDL in humans. Possible candidate neutral lipids include cholesteryl esters, triglycerides, diglycerides, and monoglycerides. This study aimed to identify the lipid classes within native and oxidized HDL that contain F₂-IsoPs. We showed that F₂-IsoPs in HDL are bound to neutral lipids as well as phospholipids. HDL-3 contained the highest concentration of F₂-IsoPs in all lipid classes before and after in vitro oxidation. Using targeted LC/MS and high resolution MS, we were unable to provide conclusive evidence for the presence of the synthesized standards 15(R)-15-F_2t-isoP cholesterol and 1-ent-15(RS)-15-F_2t-isoprostanoyl-sn-glycerol in the neutral lipids of HDL. Our findings show that oxidized lipids such as F₂-IsoPs are found in the core and surface of HDL. However, the exact molecular species remain to be definitively characterized. Future studies are required to determine whether the presence of F₂-IsoPs in neutral lipids alters HDL function.     

F2-Dihomo-isoprostanes and brain white matter damage in stage 1 Rett syndrome

Oxidative damage has been reported in Rett syndrome (RTT), a pervasive development disorder mainly caused up to 95% of cases by mutations in the X-linked methyl-CpG binding protein 2 (MeCP2) gene. We have recently synthesized F₂-Dihomo-isoprostanes (F₂-Dihomo-IsoP), peroxidation products from adrenic acid (C22:4 n − 6, AdA), a known component of myelin, and tested the potential value of F₂-Dihomo-IsoPs as a novel disease marker and its relationship with clinical presentation, and disease progression. F₂-Dihomo-IsoPs were determined by a gas chromatography/negative ion chemical ionization tandem mass spectrometry. The ent-7(RS)-F_2t-Dihomo-IsoP and 17-F_2t-Dihomo-IsoP were used as reference standards. The measured ions were the product ions at m/z 327 derived from the [M − 181]⁻ precursor ions (m/z 597) produced from both the derivatized ent-7(RS)-F_2t-Dihomo-IsoP and 17-F_2t-Dihomo-IsoP. Average plasma F₂-Dihomo-IsoP levels in RTT were about 1 order of magnitude higher than in healthy controls, being higher in typical RTT as compared to RTT variants, with a remarkable increase of about 2 orders of magnitude in patients at the earliest stage of the disease followed by a steady decrease during the natural clinical progression. These data indicate for the first time that quantification of F₂-Dihomo-IsoPs in plasma represents an early marker of the disease and may provide a better understanding of the pathogenic mechanisms behind the neurological regression in patients with RTT.     

Oxidative damage in Parkinson disease: Measurement using accurate biomarkers

Oxidative damage has been implicated in the pathogenesis of Parkinson disease (PD) but the literature data are confusing. Using products of lipid and DNA oxidation measured by accurate methods, we assessed the extent of oxidative damage in PD patients. The levels of plasma F₂-isoprostanes (F₂-IsoPs), hydroxyeicosatetraenoic acid products (HETEs), cholesterol oxidation products, neuroprostanes (F₄-NPs), phospholipase A₂ (PLA₂) and platelet activating factor–acetylhydrolase (PAF-AH) activities, urinary 8-hydroxy-2′-deoxyguanosine (8-OHdG), and serum high-sensitivity C-reactive protein were compared in 61 PD patients and 61 age-matched controls. The levels of plasma F₂-IsoPs, HETEs, 7β-and 27-hydroxycholesterol, 7-ketocholesterol, F₄-NPs, and urinary 8-OHdG were elevated, whereas the levels of plasma PLA₂ and PAF-AH activities were lower, in PD patients compared to controls (p <  0.05). The levels of plasma F₂-IsoPs, HETEs, and urinary 8-OHdG were higher in the early stages of PD (p trend <  0.05). There was a significant negative correlation between the cumulative intake of levodopa and urinary 8-OHdG (r =  ?0.305, p =  0.023) and plasma total HETEs (r =  ?0.285, p =  0.043). Oxidative damage markers are systemically elevated in PD, which may give clues about the relation of oxidative damage to the onset and progression of PD.     

Nonenzymatic lipid mediators,neuroprostanes, exert the antiarrhythmic properties of docosahexaenoic acid

Neuroprostanes are lipid mediators produced by nonenzymatic free radical peroxidation of docosahexaenoic acid (DHA). DHA is associated with a lower atherosclerosis risk, suggesting a beneficial role in cardiovascular diseases. The aim of this study was to investigate the influence of DHA peroxidation on its potentially antiarrhythmic properties (AAP) in isolated ventricular cardiomyocytes and in vivo in post-myocardial infarcted mice. Calcium imaging and biochemical experiments indicate that cardiac arrhythmias induced by isoproterenol are associated with Ca²⁺ leak from the sarcoplasmic reticulum after oxidation and phosphorylation of the type 2 ryanodine receptor (RyR2) leading to dissociation of the FKBP12.6/RyR2 complex. Both oxidized DHA and 4(RS)-4-F_4t-NeuroP prevented cellular arrhythmias and posttranslational modifications of the RyR2 leading to a stabilized FKBP12.6/RyR2 complex. DHA per se did not have AAP. The AAP of 4(RS)-4-F_4t-NeuroP was also observed in vivo. In this study, we challenged the paradigm that spontaneously formed oxygenated metabolites of lipids are undesirable as they are unconditionally toxic. This study reveals that the lipid mediator 4(RS)-4-F_4t-neuroprostane derived from nonenzymatic peroxidation of docosahexaenoic acid can counteract such deleterious effects through cardiac antiarrhythmic properties. Our findings demonstrate 4(RS)-4-F_4t-NeuroP as a mediator of the cardioprotective AAP of DHA. This discovery opens new perspectives for products of nonenzymatic oxidized ω3 polyunsaturated fatty acids as potent mediators in diseases that involve ryanodine complex destabilization such as ischemic events.     

Oxidative stress in Rett syndrome: Natural history,genotype, and variants

Abstract

Objectives

Rett syndrome (RTT) is an X-linked autism spectrum disorder caused by mutations in the MeCP2 gene in the great majority of cases. Evidence suggests a potential role of oxidative stress (OS) in its pathogenesis. Here, we investigated the potential value of OS markers (non-protein-bound iron (NPBI) and F₂-isoprostanes (F₂-IsoPs)) in explaining natural history, genotype-phenotype correlation, and clinical heterogeneity of RTT, and gauging the response to omega-3 polyunsaturated fatty acids (ω-3 PUFAs).

Methods

RTT patients (n = 113) and healthy controls were assayed for plasma NPBI and F₂-IsoPs, and intraerythrocyte NPBI. Forty-two patients with typical RTT were randomly assigned to ω-3 PUFAs supplementation for 12 months. NPBI was measured by HPLC and F₂-IsoPs using a gas chromatography/negative ion chemical ionization tandem mass spectrometry (GC/NICI-MS/MS) technique.

Results

F₂-IsoPs were significantly higher in the early stages as compared with the late natural progression of classic RTT. MeCP2 mutations related to more severe phenotypes exhibited higher OS marker levels than those of milder phenotypes. Higher OS markers were observed in typical RTT and early seizure variant as compared with the preserved speech and congenital variants. Significant reduction in OS markers levels and improvement of severity scores were observed after ω-3 PUFAs supplementation.

Discussion

OS is a key modulator of disease expression in RTT.     

Oxidative Stress Predicts All-Cause Mortality in HIV-Infected Patients

ObjectiveWe aimed to assess whether oxidative stress is a predictor of mortality in HIV-infected patients.MethodsWe conducted a nested case-control study in CoRIS, a contemporary, multicentre cohort of HIV-infected patients, antiretroviral-naïve at entry, launched in 2004. Cases were patients who died with available stored plasma samples collected. Two age and sex-matched controls for each case were selected. We measured F2-isoprostanes (F₂-IsoPs) and malondialdehyde (MDA) plasma levels in the first blood sample obtained after cohort engagement.Results54 cases and 93 controls were included. Median F₂-IsoPs and MDA levels were significantly higher in cases than in controls. When adjustment was performed for age, HIV-transmission category, CD4 cell count and HIV viral load at cohort entry, and subclinical inflammation measured with highly-sensitive C-reactive protein (hsCRP), the association of F₂-IsoPs with mortality remained significant (adjusted OR per 1 log₁₀ increase, 2.34 [1.23–4.47], P = 0.009). The association of MDA with mortality was attenuated after adjustment: adjusted OR (95% CI) per 1 log₁₀ increase, 2.05 [0.91–4.59], P = 0.080. Median hsCRP was also higher in cases, and it also proved to be an independent predictor of mortality in the adjusted analysis: OR (95% CI) per 1 log₁₀ increase, 1.39 (1.01–1.91), P = 0.043; and OR (95% CI) per 1 log₁₀ increase, 1.46 (1.07–1.99), P = 0.014, respectively, when adjustment included F₂-IsoPs and MDA.ConclusionOxidative stress is a predictor of all-cause mortality in HIV-infected patients. For plasma F₂-IsoPs, this association is independent of HIV-related factors and subclinical inflammation.     

Photoinhibition and xanthophyll cycle activity in bayberry (Myrica rubra) leaves induced by high irradiance

The effect of high irradiance (HI, photosynthetically active photon flux density of 1 300 μmol m⁻² s⁻¹) on net photosynthetic rate (P _N), chlorophyll fluorescence parameters, and xanthophyll cycle components were studied in fruit tree bayberry leaves. HI induced the photoinhibition and inactivation of photosystem 2 (PS2) reaction centres (RCs), which was characterized by decreased P _N, maximum yield of fluorescence after dark adaptation (F_m), photochemical efficiency of PS2 (F_v/F_m) and quantum yield of PS2 (Φ_PS2), and increased reduction state of Q_A (1-q_P) and non-photochemical quenching (NPQ). Initial fluorescence (F₀) showed a decrease after the first 2 h, and subsequently increased from the third hour exposure to HI. Furthermore, a greater increase in the ratio (F_i-F₀)/(F_p-F₀) which is an expression of the proportion of the Q_B non-reducing PS2 centres, whereas a remarked decrease in the slope of F_i to F_p which represents the rate of Q_A reduction was observed in leaves after HI exposure. Additionally, HI caused an increase in the pool size of the xanthophyll cycle pigments and sustained elevated contents of zeaxanthin (Z), antheraxanthin (A), and de-epoxidation state (DES) at the end of the irradiation period. During HI, decreased F_m, F_v/F_m, Φ_PS2, NPQ, slope of F_i to F_p, V+A+Z, and DES, and increased F₀, 1-q_P, ratio (F_i-F₀)/(F_p-F₀), and V were observed in dithiothreitol (DTT)-fed leaves compared to control ones under the same conditions. Hence photoinhibition caused by HI in bayberry was probably attributed to inactivation of PS2 RCs, and photoprotection from photodamage were mainly related to the xanthophyll cycle-dependent heat dissipation in excess photons.     

On the mechanism of the reconstitution of F1-depleted ATPase complex with purified F1: Possible conformational effects

The membrane sector (F₀) of H⁺-ATPase was prepared by trypsin and urea treatment of F₁-F₀ and reconstituted with purified F₁. The oligomycin sensitivity of the reconstituted F₁-F₀ complex obtained by treating F₁ or F₀ with Mg²⁺ before binding is much higher than that obtained without Mg²⁺ treatment. The greater change in the intrinsic fluorescence of the reconstituted F₁-F₀ complex obtained by Mg²⁺ treatment suggests that conformational changes may occur during the reconstitution. We deduce that Mg²⁺ binds to membrane lipids, thus decreasing membrane fluidity and changing the physical state of the lipids to provide a suitable microenvironment for conformational changes in F₀. The data also suggest that the conformational change in the F₀ portion of the F₁-F₀ complex can be transmitted to the F₁ portion, the conformation of which is in turn altered, resulting in the formation of an F₁-F₀ complex with high oligomycin sensitivity. On the other hand, Mg²⁺ may act on F₁ directly to induce a suitable conformational change which is then trnsmitted to F₀, resulting in the formation of an H⁺-ATPase with greater sensitivity to oligomycin.Abbreviations STED 0.25 M sucrose, 10 mM Tris-SO₄, 0.2 mM EDTA, and 1 mM dithiothreitol, pH 8.0 - NADH nicotinamide adenine dinucleotide, reduced form - olig. oligomycin - OSCP oligomycin sensitivity conferring protein - F₆ coupling factor 6 - F₁ coupling factor one (or F₁-ATPase) - F₁ ^{+Mg 2+} and F₁ ^{–Mg 2+} the F₁ treated and untreated with 1 mM Mg²⁺ respectively - F₀ the membrane sector proteins of the H⁺-ATPase - TUF₀ trypsin-urea – F₀ - EUF₀ EDTA-urea – F₀ - F₀ ^{+Mg 2+} and F₀ ^{–Mg 2+} the F₀ treated and untreated with 1 mM Mg²⁺ respectively - (F₁ · F₀)^{+Mg 2+} and (F₁ · F₀)^{–Mg 2+} the reconstituted F₁ · F₀ complex containing Mg²⁺-treated F₁ and F₀ and untreated F₁ and F₀ respectively - F₁ · F₀ ^{+Mg 2+} and F₁ · F₀ ^{–Mg 2+} the reconstituted H⁺-ATPase complex derived from the binding of purified F₁ to the F₀ treated and untreated with Mg²⁺ respectively - F₁ ^{+Mg 2+} · F₀ and F₁ ^{–Mg 2+} · F₀ the reconstituted H⁺-ATPase derived from the binding of F₀ to the purified F₁ treated and untreated with Mg²⁺ respectively     

Electrophilic cyclopentenone neuroprostanes are anti-inflammatory mediators formed from the peroxidation of the omega-3 polyunsaturated fatty acid docosahexaenoic acid

The omega-3 polyunsaturated fatty acid docosahexaenoic acid (DHA) possesses potent anti-inflammatory properties and has shown therapeutic benefit in numerous inflammatory diseases. However, the molecular mechanisms of these anti-inflammatory properties are poorly understood. DHA is highly susceptible to peroxidation, which yields an array of potentially bioactive lipid species. One class of compounds are cyclopentenone neuroprostanes (A(4)/J(4)-NPs), which are highly reactive and similar in structure to anti-inflammatory cyclopentenone prostaglandins. Here we show that a synthetic A(4)/J(4)-NP, 14-A(4)-NP (A(4)-NP), potently suppresses lipopolysaccharideinduced expression of inducible nitric-oxide synthase and cyclooxygenase-2 in macrophages. Furthermore, A(4)-NP blocks lipopolysaccharide-induced NF-kappaB activation via inhibition of Ikappa kinase-mediated phosphorylation of IkappaBalpha. Mutation on Ikappa kinase beta cysteine 179 markedly diminishes the effect of A(4)-NP, suggesting that A(4)-NP acts via thiol modification at this residue. Accordingly, the effects of A(4)-NP are independent of peroxisome proliferator-activated receptor-gamma and are dependent on an intact reactive cyclopentenone ring. Interestingly, free radical-mediated oxidation of DHA greatly enhances its anti-inflammatory potency, an effect that closely parallels the formation of A(4)/J(4)-NPs. Furthermore, chemical reduction or conjugation to glutathione, both of which eliminate the bioactivity of A(4)-NP, also abrogate the anti-inflammatory effects of oxidized DHA. Thus, we have demonstrated that A(4)/J(4)-NPs, formed via the oxidation of DHA, are potent inhibitors of NF-kappaB signaling and may contribute to the anti-inflammatory actions of DHA. These findings have implications for understanding the anti-inflammatory properties of omega-3 fatty acids, and elucidate novel interactions between lipid peroxidation products and inflammation.     

The dynamics of mitochondrial Ca fluxes

We have investigated the kinetics of mitochondrial Ca²⁺ influx and efflux and their dependence on cytosolic [Ca²⁺] and [Na⁺] using low-Ca²⁺-affinity aequorin. The rate of Ca²⁺ release from mitochondria increased linearly with mitochondrial [Ca²⁺] ([Ca²⁺]_M). Na⁺-dependent Ca²⁺ release was predominant al low [Ca²⁺]_M but saturated at [Ca²⁺]_M around 400 μM, while Na⁺-independent Ca²⁺ release was very slow at [Ca²⁺]_M below 200 μM, and then increased at higher [Ca²⁺]_M, perhaps through the opening of a new pathway. Half-maximal activation of Na⁺-dependent Ca²⁺ release occurred at 5-10 mM [Na⁺], within the physiological range of cytosolic [Na⁺]. Ca²⁺ entry rates were comparable in size to Ca²⁺ exit rates at cytosolic [Ca²⁺] ([Ca²⁺]_c) below 7 μM, but the rate of uptake was dramatically accelerated at higher [Ca²⁺]_c. As a consequence, the presence of [Na⁺] considerably reduced the rate of [Ca²⁺]_M increase at [Ca²⁺]_c below 7 μM, but its effect was hardly appreciable at 10 μM [Ca²⁺]_c. Exit rates were more dependent on the temperature than uptake rates, thus making the [Ca²⁺]_M transients to be much more prolonged at lower temperature. Our kinetic data suggest that mitochondria have little high affinity Ca²⁺ buffering, and comparison of our results with data on total mitochondrial Ca²⁺ fluxes indicate that the mitochondrial Ca²⁺ bound/Ca²⁺ free ratio is around 10- to 100-fold for most of the observed [Ca²⁺]_M range and suggest that massive phosphate precipitation can only occur when [Ca²⁺]_M reaches the millimolar range.     

Increases in cytosolic Ca2+ are not required for abscisic acid-inhibition of inward K+ currents in guard cells of Vicia faba L.

 The inward K⁺ channels (I_Kin) of guard cells are inhibited upon application of abscisic acid (ABA). It has been postulated that I_Kin inhibition requires an elevation in cytosolic free Ca²⁺ levels ([Ca²⁺]_c) because: (i) experimental increases in [Ca²⁺] _c can mimic the ABA effect, and; (ii) ABA can trigger an elevation of [Ca²⁺]_c in guard cells. However, not all guard cells respond to ABA with a [Ca²⁺]_c increase, and the magnitude of the increases that do occur is variable. Therefore, an obligate role for Ca²⁺ in the regulation of downstream effectors of ABA response, such as the I_Kin channels, remains in question. In this study, we developed a methodology for simultaneous patch clamping and confocal ratiometric Ca²⁺ imaging of Vicia faba L. guard-cell protoplasts. This allowed us to directly assess the relationship between ABA-induced changes in [Ca²⁺]_c and I_Kin inhibition. In the presence of extracellular Ca²⁺, the extent of [Ca²⁺]_c elevation correlated with the extent of I_Kin inhibition. However, upon chelation of either extracellular Ca²⁺, [Ca²⁺]_c, or both, extracellular Ca²⁺ and [Ca²⁺]_c, [Ca²⁺]_c elevation did not occur in response to ABA yet I_Kin currents were still strongly inhibited. These data illustrate that Ca²⁺-independent regulation is involved in ABA-inhibition of stomatal opening processes. Received: 17 September 1999 / Accepted: 26 October 1999     

A novel fluorescence chamber for the determination of volume changes in human CaSki cell cultures attached on filters

The objective of the study was to test the hypothesis that, in the cultured human cervical epithelium, CaSki, the effect of calcium mobilizing agents on transepithelial electrical conductance (G_TE), is the result of cell volume decrease. CaSki cells attached on filters were loaded with fura-2, and measurements of fluorescence at the isosbestic wavelength 360 nm (excitation/emission [F_360/510]) were made in a newly designed fluorescence chamber; this design allowed us also to determine changes in cytosolic calcium ([Ca²⁺]_i). The experimental conditions were similar to those used to measure changes in paracellular permeability in the Ussing chamber, and they enabled us to compare the time-course of changes in [Ca²⁺]_i, in F_360/510, and in G_TE. Hypertonicity increased, and hypotonicity decreased F_360/510 and G_TE, without having an effect on [Ca²⁺]_i, and the changes in F_360/510 and in G_TE correlated linearly. Metabolism, bleaching, and extrusion of intracellular fura-2 were minimal, indicating that the changes in F_360/510 reflect changes in dye concentration. Hypertonicity decreased, and hypotonicity increased the size of dispersed CaSki cells, suggesting that osmolarity-induced changes in F_360/510 reflect changes in size of the attached cells. Ionomycin increased [Ca²⁺]_i, F_360/510, and G_TE, but the increases in [Ca²⁺]_i preceded those in F_360/510 and G_TE. The calcium chelator BAPTA blocked the ionomycin-induced increase in [Ca²⁺]_i, F_360/510, and in G_TE. Preincubation with 4-acetamido-4′isothiocyanatostilbene-2,2′disulfonic acid (SITS) augmented the ionomycin-induced increase in [Ca²⁺]_i, but blocked the increases in F_360/510 and in G_TE. Pretreatment of cells with hypertonic solution abrogated the increases in F_360/510 and in G_TE in response to ionomycin, but had little effect on the ionomycin-induced increase in [Ca²⁺]_i. On the basis of these results we suggest that the ionomycin-induced increase in G_TE is mediated by [Ca²⁺]_i-dependent chloride secretion and osmotic water loss.     

Isoprostanes and isofurans as non-traditional risk factors for cardiovascular disease among Canadian Inuit

Abstract

Objectives: The aim of the present study was to investigate the potential importance of oxidative stress, measured by isoprostanes-related compounds, as non-traditional risk factor for cardiovascular disease. We planned to examine the relationship between concentrations of plasma F₂-isoprostanes (F₂-IsoPs), isofurans (IsoFs), measures of obesity and various cardiometabolic risk factors. Materials and methods: Cross-sectional study using a sub-sample from the population of a survey conducted in the summer and fall 2007 and 2008 by Canadian Coastguard Ship Amundsen in 36 Canadian Arctic Inuit communities. Subjects included a subset (n =?233) of a total study population (n =?2595) with a mean age 42.56 ± 15.39 years and body mass index 27.78 ± 5.65 kg/m². Plasma levels of F₂-IsoPs and IsoFs was determined by gas chromatography/negative ion chemical ionization/mass spectrometry (GC/NICI/MS) method; and their relationships to waist circumference (WC), blood pressure C reactive proteins (CRP), blood lipids and fasting glucose were assessed by multivariate analyses. Results: Plasma F₂-IsoPs correlated positively with CRP (r =.132, P =.048) and systolic blood pressure (SBP) (r =.157, P =.024) after adjustment for age, sex and body mass index. IsoFs correlated with WC (r =.190, P =.005) and SBP (r =.137, P =.048). F2-IsoPs were not found elevated in smokers (P =.034), whereas IsoFs were decreased in smokers (P =.001). WC, SBP and sex were found to be major correlates of oxidative stress in Canadian Inuit. Conclusions: Plasma measures of F₂-IsoPs and IsoFs increase with increased obesity and associated cardiometabolic risk factors, including CRP and blood pressure. Simultaneous measurement of IsoFs provides an advantageous mechanistic insight into oxidative stress not captured by F₂-IsoPs alone.     

Does radiotherapy increase oxidative stress? A study with nasopharyngeal cancer patients revealing anomalies in isoprostanes measurements

Abstract

This study aimed to examine if exposure to ionizing radiation during clinical radiotherapy (RT) causes increased oxidative damage. Seven patients with nasopharyngeal cancer (NPC) who underwent RT took part in this controlled-trial study. Blood and urine samples were obtained for F₂-isoprostanes (F₂-IsoPs) measurement. Urinary F₂-IsoPs levels were elevated pre-treatment and remained high (but did not increase) during treatment, but decreased to the normal range after treatment. Plasma F₂-IsoPs decreased significantly after the start of treatment before rising midway through treatment. Levels decreased significantly to below baseline following treatment. However, the patients were observed to have substantially lower levels of plasma esterified arachidonic acid (AA) residues than controls. The data shows that NPC is associated with elevated F₂-isoprostanes in urine and in plasma after correction for decreased AA levels. RT did not increase these levels and, indeed, was associated with falls in F₂-IsoPs. The validity and usefulness of correction of plasma F₂-IsoPs for lowered AA levels is discussed.     

Multiple pathways of charge recombination revealed by the temperature dependence of electron transfer kinetics in cyanobacterial photosystem I

The kinetics of charge recombination in Photosystem I P₇₀₀-F_A/F_B complexes and P₇₀₀-F_X cores lacking the terminal iron?sulfur clusters were studied over a temperatures range of 310 K to 4.2 K. Analysis of the charge recombination kinetics in this temperature range allowed the assignment of backward electron transfer from the different electron acceptors to P₇₀₀⁺. The kinetic and thermodynamic parameters of these recombination reactions were determined. The kinetics of all electron transfer reactions were activation-less below 170 K, the glass transition temperature of the water-glycerol solution. Above this temperature, recombination from [F_A/F_B]^? in P₇₀₀-F_A/F_B complexes was found to proceed along two pathways with different activation energies (E_a). The charge recombination via A_1A has an E_a of ~290 meV and is dominant at temperatures above ~280 K, whereas the direct recombination from F_X^? has an E_a of 22 meV and is prevalent in the 200 K to 270 K temperature range. Charge recombination from the F_X cluster becomes highly heterogeneous at temperatures below 200 K. The conformational mobility of Photosystem I was studied by molecular dynamics simulations. The F_X cluster was found to ‘swing’ by ~30° along the axis between the two sulfur atoms proximal to F_A/F_B. The partial rotation of F_X is accompanied by significant changes of electric potential within the iron?sulfur cluster, which may induce preferential electron localization at different atoms of the F_X cluster. These effects may account for the partial arrest of forward electron transfer and for the heterogeneity of charge recombination observed at the glass transition temperature.     

Crystal structure and properties of [TTF]2.5[V(mnt)3 ]·0.5CH2ClCH2Cl[TTF = tetrathiafulvalene and mnt = 1,2-dicyanoethylene-1,2-dithiolate(2−)]

The title salt was obtained by a reaction of [TTF]₃ [BF₄]₂ (TTF = tetrathiafulvalene) with [NMe₄]₂ [V(mnt)₃] [mnt = 1,2-dicyanoethylene-1,2- dithiolate(2−)] in a mixture of 1,2-dichloroethane and acetonitrile (3:2 ν/ν). A single crystal X-ray analysis of it revealed a TTF columnar structure consisting of both TTF⁰ and the TTF^·+ radical cation and the distorted octahedral geometry of the [V(mnt)₃]²⁻ anion. The salt crystallizes in a monoclinic system, space group C2/c with unit cell constants a = 25.428(3), b = 12.434(2), c = 25.477(3) Å, β = 92.428(3)° and Z = 8. The structure was solved by the direct method and refined, on the basis of 3854 [|F_o| > 3σ(F)] observed data, to an R value of 0.078. The salt e`xhibits electrical conductivity of 1.7 x 10⁻⁴ S cm⁻¹ at 25°C for a compacted pellet.     

Crystal structures of trisammonium bisperoxotetrafluoroniobate(V) and analogous tantalate(V)

(NH₄)₃[Nb(O₂)₂F₄] (I) and (NH₄)₃[Ta(O₂)₂F₄] (II) are isostructural, and belong to the cubic Fm3m space group with four molecules in the unit cell. The unit cell parameters are a = 9.4442(4) (I) and a = 9.4512(4) Å (II). The structures were solved by the Patterson method and were refined by the least-squares method to the conventional R factors of 0.036 for 86 reflections (I) and 0.043 for 103 reflections (II) (in both structures having I ? 2σ(I)). The disordered distributions of fluorine and peroxo oxygens with partially occupied sites are observed. The disordered NH₄⁺ tetrahedra appear in the structures.The metal atoms exhibit an octahedral coordination with two corners of a polyhedron at the centre of the peroxo bonds. Inter-atomic distances are NbF, 1.95(2), NbO, 1.94, TaF, 1.91(4) and TaO, 2.07 Å.The structures (I, II) are composed of [M^υ(O₂)₂-F₄]^3? octahedra and two symmetrically-independent ammonium cations connected by NH?O and NH?F hydrogen bonds. These two structures are compared with the structure of (NH₄)₃Ti(O₂)F₅].     

Effect of aluminum on cytoplasmic Ca2+ homeostasis in root hairs of Arabidopsis thaliana (L.)

Aluminum inhibition of root growth is a major world agricultural problem where the cause of toxicity has been linked to changes in cellular calcium homeostasis. Therefore, the effect of aluminum ions (Al) on changes in cytoplasmic free calcium concentration ([Ca²⁺]_c) was followed in root hairs of wild-type, Al-sensitive and Al-resistant mutants of Arabidopsis thaliana (L.) Heynh. Generally, Al exposure resulted in prolonged elevations in tip-localized [Ca²⁺]_c in both wild-type and Al-sensitive root hairs. However, these Al-induced increases in [Ca²⁺]_c were not tightly correlated with growth inhibition, occurring up to 15 min after Al had induced growth to stop. Also, in 32% of root hairs examined growth stopped without a detectable change in [Ca²⁺]_c. In contrast, Al-resistant mutants showed little growth inhibition in response to AlCl₃ exposure and in no case was a change in [Ca²⁺]_c observed. Of the other externally applied stresses tested (oxidative and mechanical stress), both were found to inhibit root hair growth, but only oxidative stress (H₂O₂, 10 μM) caused a prolonged rise in [Ca²⁺]_c similar to that induced by Al. Again this increase occurred after growth had been inhibited. The lack of a tight correlation between Al exposure, growth inhibition and altered [Ca²⁺]_c dynamics suggests that although exposure of root hairs to toxic levels of Al causes an alteration in cellular Ca²⁺ homeostasis, this may not be a required event for Al toxicity. The elevation in [Ca²⁺]_c induced by Al also strongly suggests that the phytotoxic action of Al in root hairs is not through blockage of Ca²⁺-permeable channels required for Ca²⁺ influx into the cytoplasm. Received: 24 October 1997 / Accepted: 6 March 1998     

Oxidative stress,erythrocyte ageing and plasma non-protein-bound iron in diabetic patients

Increased oxidative stress and decreased life span of erythrocytes (RBCs) are repeatedly reported in diabetes. In the aim to elucidate the mechanism of the latter, i.e. the events leading to erythrocyte ageing, this study determined in RBCs from diabetic patients iron release in a free desferrioxamine-chelatable form (DCI), methemoglobin (MetHb) formation, binding of autologous IgG to membrane proteins and in plasma non-protein-bound iron (NPBI), F₂-Isoprostanes (F₂-IsoPs) and advanced oxidation protein products (AOPP). DCI and MetHb were higher in diabetic RBCs than in controls and autologous IgG binding occurred in a much higher percentage of diabetic patients than controls. A significant correlation between DCI and IgG binding was found in diabetic RBCs. Plasma NPBI, esterified F₂-IsoPs and AOPP were higher in diabetic patients and a significant correlation was found between plasma NPBI and intra-erythrocyte DCI. The increased DCI and autologous IgG binding appear to be important factors in the accelerated removal of RBCs from the blood stream in diabetes and the increase in plasma NPBI could play an important role in the increased oxidative stress.