Changes in urinary dinor dihydro F(2)-isoprostane metabolite concentrations, a marker of oxidative stress, during and following asthma exacerbations

To investigate changes in oxidant stress during and following acute asthma exacerbations, this study measured 2,3-dinor-5,6-dihydro-15-F(2t)-IsoP (F(2)-IsoP-M), the major urinary metabolite of 15-F(2t)-IsoP, in eight asthmatic adults, during and following an asthma hospitalization. F(2)-IsoP-M concentrations at admission and follow-up were significantly higher than discharge (admission median: 4.12 ng/Cr mg, range 1.89-7.8; follow-up: 2.47 ng/Cr mg (1.56-6.86); discharge: 1.42 ng/Cr mg (0.7-4.44); both p<0.01), but not significantly different between admission and follow-up. F(2)-IsoP-M concentrations at follow-up were higher than a control group with stable asthma (0.68 ng/Cr mg (0.31-1.5), p=0.0008). In conclusion, asthma exacerbations requiring hospitalization are associated with 6-fold higher urinary F(2)-IsoP-M concentrations compared to stable asthmatics. F(2)-IsoP-M concentrations decreased significantly during hospitalization, but significant elevations 3 months following hospitalization suggest ongoing oxidative stress despite clinical improvement. Urinary F(2)-IsoP-M may be a clinically useful, simple non-invasive systemic measure of oxidative stress in asthmatics, providing information not captured by spirometry or symptoms.     

Evaluation of urinary biomarkers of oxidative/nitrosative stress in adolescents and adults with Down syndrome

Urinary biomarkers of oxidative stress have been little studied in adults with Down syndrome (DS), usually no more than two biomarkers have been measured in the population studied and controversial results are reported in literature. Thus, we aimed to assess a set of oxidative and nitrosative stress biomarkers in urine samples of adolescents and adults with DS, with and without hypothyroidism, which comprise: 8-hydroxy-2′-deoxyguanosine (8-OHdG), isoprostane 15-F_2t-IsoP, thiobarbituric acid-reacting substances (TBARS), advanced glycation end products (AGEs), dityrosine (diTyr), hydrogen peroxide (H₂O₂) and nitrite/nitrate (NOx). Fluorimetric and spectrophotometric assays were performed in DS (n = 78), some of them taking levothyroxine for hypothyroidism (n = 24), and in their healthy age-matched controls (n = 65). We found that levels of AGEs, diTyr, H₂O₂ and NOx are increased in DS patients in any or in all age groups, whereas Cr levels were lower in DS than in controls in all age groups. Besides, correlations with age in DS were positive for diTyr and negative for Cr, TBARS, 15-F_2t-IsoP and NOx. We also found lower levels of Cr from 15 to 19 years, higher levels of TBARS and AGEs from 20 to 40 years and higher levels of diTyr from 15 to 40 years in DS patients receiving levothyroxine than in DS without hypothyroidism diagnosed. We conclude that AGEs, diTyr, H₂O₂ and NOx could be used as oxidative stress biomarkers in DS in contrast to 8-OHdG, 15-F_2t-IsoP and TBARS, at least with the methods used. However, renal impairment could occur in DS and Cr adjustment may bias the results, particularly in hypothyroid patients.     

Determination of isoprostaglandin F2α type III in human urine by gas chromatography–electronic impact mass spectrometry. Comparison with enzyme immunoassay

F₂-Isoprostanes are stable lipid peroxidation products of arachidonic acid, the quantification of which provides an index of oxidative stress in vivo. We describe a method for analysing isoprostaglandin F_2α type III (15-F_2t-IsoP) in biological fluids. The method involves solid-phase extraction on octadecyl endcapped and aminopropyl cartridges. After conversion to trimethylsilyl ester trimethylsilyl ether derivatives, isoprostaglandin F_2α type III is analysed by mass spectrometry, operated in electronic impact selected ion monitoring mode. We have compared enzyme immunoassay (EIA; Cayman, Ann Arbor, MI, USA) to this method with 30 human urine aliquots following the same extraction procedure in order to determine the agreement between both methods. Isoprostaglandin F_2α type III concentrations determined with gas chromatography–mass spectrometry (GC–MS) did not agree with those determined with EIA. Our results suggest that GC–MS and EIA do not measure the same compounds. As a consequence, comparison of clinical results using GC–MS and EIA should be avoided.     

An improved GC/MS-based procedure for the quantitation of the isoprostane 15-F2t-IsoP in rat plasma

This article describes a procedure for the quantitation of the isoprostane 15-F_2t-IsoP (9a,11a,15S-trihydroxy-(8b)-prosta-5Z,13E-dien-1-oic acid [CAS#27415-26-5] formerly known as 8-epi-PGF_2a or 8-iso-PGF_2a, and also as iPF_2a-III). We have combined features from several earlier methods for 15-F_2t-IsoP and prostaglandins, and identified and modified those steps that may lead to poor recoveries. The resulting protocol is precise and reliable, and was validated by a blind time-course study of plasma levels in rats treated with 120 and 1200 mg CCl₄/kg body weight. Plasma levels of 15-F_2t-IsoP, as measured according to the procedure described above, are good indicators of acute oxidative stress as induced by CCl₄. The precision of the measurements allows detection of elevated plasma 15-F_2t-IsoP levels as long as 16 h after an acute exposure of 120 mg CCl₄/kg body weight, and 2 h after an exposure of 1 mg CCl₄/kg body weight. The results of this low-dose, pilot study suggest that this method has sufficient analytical precision to allow the detection of the small changes in plasma isoprostane levels, which result from chronic and/or lower-level exposures to agents causing oxidative stress.     

F₂-isoprostane formation, measurement and interpretation: the role of exercise

The level of F₂-isoprostanes (F₂-IsoP) in blood or urine is widely regarded as the reference marker for the assessment of oxidative stress. As a result, nowadays, F₂-IsoP is the most frequently measured oxidative stress marker. Nevertheless, determining F₂-IsoP is a challenging task and the measurement is neither free of mishaps nor straightforward. This review presents for the first time the effect of acute and chronic exercise on F₂-IsoP levels in plasma, urine and skeletal muscle, placing emphasis on the origin, the methodological caveats and the interpretation of F₂-IsoP alterations. From data analysis, the following effects of exercise have emerged: (i) acute exercise clearly increases F₂-IsoP levels in plasma and this effect is generally short-lived, (ii) acute exercise and increased contractile activity markedly increase F₂-IsoP levels in skeletal muscle, (iii) chronic exercise exhibits trend for decreased F₂-IsoP levels in urine but further research is needed. Theoretically, it seems that significant amounts of F₂-IsoP can be produced not only from phospholipids but from neutral lipids as well. The origin of F₂-IsoP detected in plasma and urine (as done by almost all studies in humans) remains controversial, as a multitude of tissues (including skeletal muscle and plasma) can independently produce F₂-IsoP.     

Correlation of F4-neuroprostanes levels in cerebrospinal fluid with outcome of aneurysmal subarachnoid hemorrhage in humans

Aneurysmal subarachnoid hemorrhage (aSAH) is one type of hemorrhagic stroke in humans. F₂-isoprostanes (F₂-IsoPs) and F₄-neuroprostanes (F₄-NPs), derived from arachidonic acid and docosahexaenoic acid (DHA), respectively, are specific markers of lipid peroxidation. We previously demonstrated that F₂-IsoPs levels in cerebrospinal fluid (CSF) of aSAH patients positively correlated with poor clinical conditions. In this work, we refined F₄-NPs analysis and investigated the role of potential oxidative damage to neurons in aSAH patients by detecting F₄-NPs in CSF. [²H₄]-15-F_2t-IsoP, rather than [¹⁸O₂]-17-F_4c-NP or [²H₄]-PGF_2α, was used as the internal standard for F₄-NPs analysis. One problem of the use of [¹⁸O₂]-17-F_4c-NP was the potential interference resulting from F₂-dihomo-IsoPs in CSF. CSF specimens of 15 aSAH patients for up to 10 days and those of 12 non-aSAH controls were analyzed. First day, mean, and peak levels of F₄-NPs were all significantly higher in aSAH patients than in controls and correlated with the Fisher Scale and 3-month Glasgow Outcome Scale, but only mean levels of F₄-NPs correlated with Hunt and Hess Grade. The results first demonstrate oxidative damage to DHA in brain tissue following aSAH and suggest that F₄-NPs in CSF could be a better predictor for outcome of aSAH than F₂-IsoPs at early time points.     

Quantification of dinor, dihydro metabolites of F2-isoprostanes in urine by liquid chromatography/tandem mass spectrometry

The F2-isoprostanes (F2-IsoP) are a series of prostaglandin (PG)-F2-like compounds that are produced by free-radical-mediated oxidation of arachidonic acid. One F2-IsoP with potent biological activity is 15-F2t-IsoP and increased levels of 15-F(2t)-IsoP have been measured in several diseases. The major urinary metabolite of 15-F2t-IsoP (8-iso-PGF(2alpha)) is 2,3-dinor-5,6-dihydro-15-F2t-IsoP (15-F2t-IsoP-M). Previously, we developed a stable isotope dilution gas chromatography/negative chemical ionization/mass spectrometry (MS) assay for 15-F2t-IsoP-M, which, while highly sensitive, required time-consuming derivatization and thin-layer chromatography purification. We now report the development of a more rapid high-performance liquid chromatography method coupled to electrospray ionization-tandem mass spectrometry (LC/MS/MS) to analyze all of the dinor,dihydro metabolites of the F2-IsoP isomers (F2-IsoP-M). The precision of this assay was +/-5.0% and the accuracy 80%. The assay remained linear over a range of 1-100 ng injected onto the LC column. Levels of F2-IsoP-M determined by the LC/MS/MS assay method significantly correlated with levels of 15-F2t-IsoP-M determined by the GC/MS assay (R = 0.77y = 67.2x-0.5). The levels of F2-IsoP-M detected in spot urines from 40 normal subjects were 38.1+/-19.1 ng/mg creatinine (mean+/-SD). This method provides an accurate and rapid assay to assess oxidative status in vivo.     

Current development in non-enzymatic lipid peroxidation products,isoprostanoids and isofuranoids,in novel biological samples

Abstract

Isoprostanoids and isofuranoids are lipid mediators that can be formed from omega-3 and omega-6 polyunsaturated fatty acids (PUFAs). F₂-isoprostanes formed from arachidonic acid, especially 15-F_2t-isoprostane, are commonly measured in biological tissues for decades as the biomarker for oxidative stress and diseases. Recently, other forms of isoprostanoids derived from adrenic, eicosapentaenoic, and docosahexaenoic acids namely F₂-dihomo-isoprostanes, F₃-isoprostanes, and F₄-neuroprostanes respectively, and isofuranoids including isofurans, dihomo-isofurans, and neurofurans are reported as oxidative damage markers for different metabolisms. The most widely used samples in measuring lipid peroxidation products include but not limited to the blood and urine; other biological fluids, specialized tissues, and cells can also be determined. In this review, measurement of isoprostanoids and isofuranoids in novel biological samples by gas chromatography (GC)–mass spectrometry (MS), GC–MS/MS, liquid chromatography (LC)–MS, and LC–MS/MS will be discussed.     

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.     

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.     

Physiological responses of carbon-sequestering microalgae to elevated carbon regimes

In order to identify a high carbon-sequestering microalgal strain, the physiological effect of different concentrations of carbon sources on microalgae growth was investigated. Five indigenous strains (I-1, I-2, I-3, I-4 and I-5) and a reference strain (I-0: Coccolithus pelagicus 913/3) were subjected to CO₂ concentrations of 0.03–15% and NaHCO₃ of 0.05–2 g CO₂ l^–1. The logistic model was applied for data fitting, as well as for estimation of the maximum growth rate (μ_max) and the biomass carrying capacity (B_max). Amongst the five indigenous strains, I-3 was similar to the reference strain with regards to biomass production values. The B_max of I-3 significantly increased from 214 to 828 mg l^–1 when CO₂ concentration was increased from 0.03 to 15% (r = 0.955, P = 0.012). Additionally, the B_max of I-3 increased with increasing NaHCO₃ (r = 0.885, P = 0.046) and was recorded at 153 mg l^–1 (at 0.05 g CO₂ l^–1) and 774 mg l^–1 at (2 g CO₂ l^–1). Relative electron transport rate (rETR) and maximum quantum yield (F_v/F_m) were also applied to assess the impact of elevated carbon sources on the microalgal cells at the physiological level. Isolate I-3 displayed the highest rETR confirming its tolerance to higher quantities of carbon. Additionally, the decline in F_v/F_m with increasing carbon was similar for strains I-3 and the reference strain. Based on partial 28s ribosomal RNA gene sequencing, strain I-3 was homologous to the ribosomal genes of Chlorella sp.     

Hemoglobin attenuates the effects of inspired oxygen on plasma isofurans in humans during upper-limb surgery

Reperfusion injury is characterized by significant oxidative stress. F₂-isoprostanes (F₂-IsoP's) and isofurans (IsoF's), the latter preferentially produced during increased oxygen tension, are recognized markers of in vivo oxidative stress. We aimed to determine whether increasing oxygen tension during reperfusion modified levels of plasma total IsoF's and F₂-IsoP's. Forty-five patients undergoing upper-limb surgery were randomized to receive inspired oxygen concentrations of 30, 50, or 80% during the last 15 min of surgery. Venous blood samples were taken before the change in inspired oxygen, after 10 min (before reperfusion), and after 15 min (5 min after reperfusion). IsoF's and F₂-IsoP's were measured by gas chromatography-mass spectrometry. Venous oxygen tension and hemoglobin concentrations were also measured. Plasma IsoF and F₂-IsoP levels in the 50 and 80% O₂ groups were not significantly different from those of the 30% O₂ group. In secondary analyses, using data combining all groups, levels of IsoF's, but not F₂-IsoP's, associated with higher venous oxygen tension (P = 0.038). Hemoglobin negatively modified the influence of oxygen tension on levels of IsoF's (P = 0.014). This study has shown, for the first time, that plasma IsoF levels associate with higher oxygen tension in a human model of reperfusion, and this effect is significantly attenuated by hemoglobin.     

Enhanced zinc consumption prevents cadmium-induced alterations in lipid metabolism in male rats

It has been investigated, based on a rat model of human exposure to cadmium (Cd), whether zinc (Zn) supplementation may prevent Cd-induced alterations in lipid metabolism. For this purpose, the concentrations of free fatty acids (FFA), phospholipids (PL), triglycerides (TG), total cholesterol (TCh), and high and low density lipoprotein cholesterol (HDL and LDL, respectively) as well as the concentrations of chosen indices of lipid peroxidation such as lipid peroxides (LPO), F₂-isoprostane (F₂-IsoP) and oxidized LDL (oxLDL) were estimated in the serum of male Wistar rats administered Cd (5 or 50 mg/l) or/and Zn (30 or 60 mg/l) in drinking water for 6 months. The exposure to 5 and 50 mg Cd/l resulted in marked alterations in the lipid status reflected in increased concentrations of FFA, TCh, LDL, LPO, F₂-IsoP and oxLDL, and decreased concentrations of PL and HDL in the serum. The concentrations of LDL, LPO, F₂-IsoP and oxLDL were more markedly enhanced at the higher Cd dosage. The supplementation with Zn during the exposure to 5 and 50 mg Cd/l entirely prevented all the Cd-induced changes in the serum concentrations of the estimated lipid compounds and indices of lipid peroxidation, except for the F₂-IsoP for which Zn provided only partial protection. Based on the results it can be concluded that Zn supplementation during exposure to Cd may have a protective effect on lipid metabolism consisting in its ability to prevent hyperlipidemia, including especially hypercholesterolemia, and to protect from lipid peroxidation. The findings seem to suggest that enhanced dietary Zn intake during Cd exposure, via preventing alterations in the body status of lipids may, at least partly, protect against some effects of Cd toxicity, including oxidative damage to the cellular membranes and atherogenic action. The paper is the first report suggesting protective impact of Zn against proatherogenic Cd action on experimental model of chronic moderate and relatively high human exposure to this toxic metal.     

The yeast mitochondrial adenosine triphosphatase complex. Purification, subunit composition, and some effects of protease inhibitors.

(i) The method of preparing the oligomycin-insensitive F₁-ATPase by chloroform treatment of mitochondrial membranes (Beechey et al., 1975, Biochem. J.148, 533–537) has been modified such that a five-subunit protein is obtained from yeast with an activity of 140 μmol of ATP hydrolyzed/min/mg of protein. Repetition of this procedure in the presence of protease inhibitors (in particular, p-aminobenzamidine) allows isolation of a four-subunit protein with an activity of 243 μmol of ATP hydrolyzed/min/ mg of protein, (ii) A modified procedure is described for the preparation of the yeast oligomycin-sensitive F₁-F₀ ATPase complex, making use of protease inhibitors throughout and solubilization of the ATPase from mitochondrial membranes using Triton X-100 and sodium deoxycholate simultaneously. Two polypeptides Of 42,000 and 29,000 molecular weight are eliminated, the largest corresponding to the missing band of the F₁ sector. The complex retains oligomycin- and uncoupler-sensitive ATP-³²P_i exchange and ATP-driven proton uptake, indicating the retention of a complete coupling mechanism. (iii) F₁-ATPase is released from the F₁-F₀ complex by brief heating at 50 °C in the presence of ATP. The remaining hydrophobic polypeptides aggregate and are isolated by centrifugation. The F₁ sector can be isolated containing either four or five subunits depending on whether the starting F₁-F₀ complex contained the 42,000 and 29,000 molecular weight polypeptides. (iv) Sensitivity of the F₁-F₀ ATPase complex to oligomycin and dicyclohexylcarbodiimide varies considerably depending on the activity measured and whether the complex was first reconstituted with phospholipids. The degree of inhibitor sensitivity is considered a poor guide to intactness of the complex.     

Assessment of oxidative stress biomarkers – neuroprostanes and dihomo-isoprostanes – in the urine of elite triathletes after two weeks of moderate-altitude training

This randomized and controlled trial investigated whether the increase in elite training at different altitudes altered the oxidative stress biomarkers of the nervous system. This is the first study to investigate four F₄-neuroprostanes (F₄-NeuroPs) and four F₂-dihomo-isoprostanes (F₂-dihomo-IsoPs) quantified in 24-h urine. The quantification was carried out by ultra high pressure liquid chromatography-triple quadrupole-tandem mass spectrometry (UHPLC-QqQ-MS/MS). Sixteen elite triathletes agreed to participate in the project. They were randomized in two groups, a group submitted to altitude training (AT, n?=?8) and a group submitted to sea level training (SLT) (n?=?8), with a control group (Cg) of non-athletes (n?=?8). After the experimental period, the AT group triathletes gave significant data: 17-epi-17-F_2t-dihomo-IsoP (from 5.2?±?1.4?μg/mL 24?h^?1 to 6.6?±?0.6?μg/mL 24?h^?1), ent-7(RS)-7-F_2t-dihomo-IsoP (from 6.6?±?1.7?μg/mL 24?h^?1 to 8.6?±?0.9?μg/mL 24?h^?1), and ent-7-epi-7-F_2t-dihomo-IsoP (from 8.4?±?2.2?μg/mL 24?h^?1 to 11.3?±?1.8?μg/mL 24?h^?1) increased, while, of the neuronal degeneration-related compounds, only 10-epi-10-F_4t-NeuroP (8.4?±?1.7?μg/mL 24?h^?1) and 10-F_4t-NeuroP (5.2?±?2.9?μg/mL 24?h^?1) were detected in this group. For the Cg and SLT groups, no significant changes had occurred at the end of the two-week experimental period. Therefore, and as the main conclusion, the training at moderate altitude increased the F₄-NeuroPs- and F₂-dihomo-isoPs-related oxidative damage of the central nervous system compared to similar training at sea level.     

Isoprostanes and asthma

Isoprostanes are prostaglandin (PG)-like compounds generated in vivo following oxidative stress by non-enzymatic peroxidation of polyunsaturated fatty acids, including arachidonic acid. They are named based on their prostane ring structure and by the localization of hydroxyl groups on the carbon side chain; these structural differences result in a broad array of isoprostane molecules with varying biological properties. Generation of specific isoprostanes is also regulated by host cell redox conditions; reducing conditions favor F₂-isoprostane production while under conditions with deficient antioxidant capacity, D₂- and E₂-isoprostanes are formed. F₂-isoprostanes (F₂-isoP) are considered reliable markers of oxidative stress in pulmonary diseases including asthma. Importantly, F₂-isoP and other isoprostanes function as ligands for PG receptors, and potentially other receptors that have not yet been identified. They have been reported to have important biological properties in many organs. In the lung, isoprostanes regulate cellular processes affecting airway smooth muscle tone, neural secretion, epithelial ion flux, endothelial cell adhesion and permeability, and macrophage adhesion and function. In this review, we will summarize the evidence that F₂-isoP functions as a marker of oxidative stress in asthma, and that F₂-isoP and other isoprostanes exert biological effects that contribute to the pathogenesis of asthma. This article is part of a Special Issue entitled Biochemistry of Asthma.     

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.     

Comparison of three oxidative stress biomarkers in a sample of healthy adults

Oxidative stress is a potentially important aetiological factor for many chronic diseases, including cardiovascular disease, neurodegenerative disease and cancer, yet studies often find inconsistent results. The associations between three of the most widely used biomarkers of oxidative stress, i.e. F₂-isoprostanes for lipid peroxidation and 8-oxo-7,8-dihydro-2’-deoxyguanosine (8-oxo-dG) and the comet assay with FPG for oxidative DNA damage, were compared in a sample of 135 healthy African-American and white adults. Modest associations were observed between F₂-isoprostanes and the comet assay (r?=?0.22, p?=?0.01), but there were no significant correlations between 8-oxo-dG and the comet assay (r?=??0.09) or F₂-IsoP (r?=??0.04). These results are informative for researchers seeking to compare results pertaining to oxidative stress across studies and/or assessment methods in healthy disease-free populations. The development and use of oxidative stress biomarkers is a promising field; however, additional validation studies are necessary to establish accuracy and comparability across oxidative stress biomarkers.     

Short-time UVA exposure to human keratinocytes instigated polyunsaturated fatty acid without inducing lipid peroxidation

Short-term exposure to ultraviolet A (UVA) radiation can directly injure our skin through inflammatory response and indirectly through oxidative stress, triggering polyunsaturated fatty acid (PUFA) peroxidation in skin cell membrane and formation of DNA adduct, 8-hydroxy-2′-deoxyguanosine (8-OHdG). It is known that UVA exposure leads to photoaging, immunosuppression and skin cancer. However, the changes in PUFA and its oxidized metabolites, and cell cycle after short UVA exposure, are debatable. In this study, human keratinocytes (HaCaT) were exposed to low dose (5?J/cm²) and high dose (20 J/cm²) of UVA and assessed immediately, 8?h, 12?h, and 24?h post-treatment. Both doses showed a transient suppression in S-phase after 8?h of UVA exposure, and G₂/M phase arrest after 12-h UVA exposure in the cell cycle but subsequently returned to normal cycle. Also, no observable DNA damage took place, where 8-OHdG levels were below par after 24-h UVA exposure. A dose of 20 J/cm² UVA stimulated significant amount of arachidonic acid, n-3 docosapentaenoic acid, and docosahexaenoic acid (DHA) but lowered adrenic acid and eicospentaenoic acid after 24-h exposure. Among the 43 oxidized PUFA products determined, enzyme-dependent oxidized PUFAs, namely, 14-hydroxy-DHA (HDoHE) level reduced, and 8- and 13-HDoHE levels elevated significantly in a linear trend with post-treatment time. Out of the nonenzymatic oxidized PUFAs, a significant linear trend with post-treatment time was shown on the reduction of 5-F_2t-Isoprostane (IsoP), 15-F_2t-IsoP, Isofurans, 5-F_3t-IsoP, Neurofurans, and 20-HDoHE. Our observations indicate oxidative stress through short UVA exposure on human keratinocytes did not have detrimental consequences.     

Detrimental effects of environmental tobacco smoke in relation to asthma severity

Background

Environmental tobacco smoke (ETS) has adverse effects on the health of asthmatics, however the harmful consequences of ETS in relation to asthma severity are unknown.

Methods

In a multicenter study of severe asthma, we assessed the impact of ETS exposure on morbidity, health care utilization and lung functions; and activity of systemic superoxide dismutase (SOD), a potential oxidative target of ETS that is negatively associated with asthma severity.

Findings

From 2002–2006, 654 asthmatics (non-severe 366, severe 288) were enrolled, among whom 109 non-severe and 67 severe asthmatics were routinely exposed to ETS as ascertained by history and validated by urine cotinine levels. ETS-exposure was associated with lower quality of life scores; greater rescue inhaler use; lower lung function; greater bronchodilator responsiveness; and greater risk for emergency room visits, hospitalization and intensive care unit admission. ETS-exposure was associated with lower levels of serum SOD activity, particularly in asthmatic women of African heritage.

Interpretation

ETS-exposure of asthmatic individuals is associated with worse lung function, higher acuity of exacerbations, more health care utilization, and greater bronchial hyperreactivity. The association of diminished systemic SOD activity to ETS exposure provides for the first time a specific oxidant mechanism by which ETS may adversely affect patients with asthma.