Analytical and biological variation of biomarkers of oxidative stress during the menstrual cycle.

Little information is available on the intra-individual variability of oxidative stress biomarkers in healthy individuals and even less in the context of the menstrual cycle. The objective of this study was to characterize the analytical and biological variability of a panel of 21 markers of oxidative damage, antioxidant defence and micronutrients in nine healthy, regularly menstruating women aged 18-44 years. Analyses included measurement of lipid peroxidation, antioxidant enzymes and antioxidant vitamins. Blood specimens were collected, processed and stored using standardized procedures on days 2, 7, 12, 13, 14, 18, 22 and 28 in one cycle for each subject. Replicate analyses of markers were performed and two-way nested random effects ANOVA was used to describe analytical, intra-individual and inter-individual variability. No statistically significant differences at alpha=0.05, or temporal effects across the menstrual cycle were observed. Analytical variability was the smallest component of variance for all variables. The ICC among replicates ranged from 0.80 to 0.98. Imprecision based on quality control materials ranged from 1 to 11%. The critical differences between serial results varied greatly between assays ranging from 6 to 216% of the mean level. These results provide important initial information on the variability of biomarkers of oxidative stress, antioxidant defence and micronutrients across the menstrual cycle.     

Changes of markers of oxidative stress during menstrual cycle

The levels of urinary hydrogen peroxide and thiobarbituric acid reactive substances have been compared during the menstrual cycle of 12 regularly menstruating women. Higher level of both indices of oxidative stress (normalized with respect to creatinine content) were found in the luteal phase of the cycle. These results give further evidence for the usefulness of urinary hydrogen peroxide and thiobarbituric acid reactive substances as potential biomarkers of oxidative stress and for the antioxidant action of estrogens.     

Variability of salivary markers of oxidative stress and antioxidant status in young healthy individuals

Objectives: Salivary advanced glycation end-products (AGEs), advanced oxidation protein products (AOPP), total antioxidant capacity (TAC), and ferric reducing ability of saliva (FRAS) are increased in various diseases. Little data exist for these markers in the healthy population. The aim of this study was to assess the inter-individual and intra-individual variability of AGEs, AOPP, TAC, and FRAS in the saliva of young healthy individuals.

Methods: Unstimulated saliva samples were collected from 16 females and 18 males daily over a period of 30 days. Markers were measured using spectrophotometric and spectrofluorometric microplate-based methods.

Results: All salivary markers measured were significantly higher in men than in women (P?<?0.05 for AGEs; P?<?0.001 for AOPP, TAC, and FRAS). The inter-individual variability was approximately 60% for AGEs and AOPP and 30–40% for TAC and FRAS in both genders. The inter-individual variability of FRAS was higher in men vs. women (P?<?0.01). Intra-individual variability ranged from 20% for TAC, to 30% for AGES and FRAS and 45% for AOPP.

Discussion: Intra-individual variability of salivary AGEs, AOPP, TAC, and FRAS indicates that their use is currently limited to large cohort studies. Identifying the underlying factors related to the high inter-individual and intra-individual variability is needed. Sex differences should be considered in future studies.     

Impact of serum estradiol on postprandial lipemia,oxidative stress,and inflammation across a single menstrual cycle

Background: An abrupt rise in circulating lipids, oxidative stress, and inflammatory biomarkers is a common finding after ingestion of a high-fat meal. Estradiol, typically provided via hormone replacement therapy to postmenopausal women, has been reported to possess lipidemic, antioxidant, and antiinflammatory properties, all of which may minimize postprandial oxidative stress.Objective: The purpose of this study was to compare the postprandial triglyceride (TG), oxidative stress, and inflammatory responses after a lipid meal in menstruating women during the early follicular (days 1–3) and preovulatory (day 14) phases of the menstrual cycle.Methods: Healthy normolipidemic women (fasting blood TG, <200 mg/dL) with regular menstrual cycles reported to the Cardiorespiratory/Metabolic Laboratory at the University of Memphis, Memphis, Tennessee (October-December 2008) and consumed an identical lipid meal (heavy whipping cream and water) on 2 separate days during the menstrual cycle. Blood samples were collected premeal and 1, 2, 4, and 6 hours postmeal, then assayed for TG, malondialdehyde (MDA), hydrogen peroxide, Trolox equivalent antioxidant capacity (TEAL), nitrate/nitrite, and C-reactive protein (CRP). The AUC was calculated for each variable, and a 2 (menstrual cycle phase) × 5 (time) ANOVA with Tukey post hoc testing was also conducted. Estradiol concentration was measured in premeal samples for verification of cycle phase.Results: Ten women (mean [SD] age, 29 [11] years; 8 white, 2 black; body mass index, 22 [3] kg/m²) participated in the study. Despite a higher serum estradiol concentration on day 14 (113 [56] pg/mL) compared with the early follicular phase (61 [34] pg/mL), the TG, oxidative stress, and inflammatory AUC responses to feeding were not significantly different. TG (P = 0.03), MDA (P = 0.02), and hydrogen peroxide (P < 0.001) were significantly increased in response to feeding (time effect), whereas nitrate/nitrite was decreased (P = 0.01). TEAC and CRP were not significantly affected.Conclusions: These data indicate that estradiol, at the concentrations noted in the present study, had no significant effect on postprandial TG or biomarkers of oxidative stress or inflammation in a sample of young, healthy women. It is possible that a greater divergence in circulating estradiol may be needed for significant differences to be detected, as may be the case with chronic hormone replacement therapy in postmenopausal women.     

Salivary testosterone measurements among women: relative magnitude of circadian and menstrual cycles.

Measures of testosterone among women are potentially useful in behavioral research, but information is needed on how much error is introduced by variability across the menstrual cycle. Morning and evening salivary testosterone concentrations were measured at weekly intervals across one menstrual cycle in each of 22 women, using the luteinizing hormone surge to mark midcycle. Menstrual cycles were statistically significant but smaller than daily cycles or individual differences. Menstrual cycle effects can be ignored in most research relating psychological and behavioral variables to individual differences in testosterone.     

Age- and gender-related oxidative status determined in healthy subjects by means of OXY-SCORE, a potential new comprehensive index.

Oxidative stress has been related to various diseases, gender and ageing, and has been measured by various markers. The authors developed a procedure to compute a global oxidative stress index (OXY-SCORE), reflecting both oxidative and antioxidant markers in healthy subjects. Its performance was tested in relation to age and gender and in coronary artery disease (CAD) patients. Eighty-two healthy subjects and 20 CAD patients were enrolled. Plasma free and total malondialdehyde (F- and T-MDA), glutathione disulphide/reduced form ratio (GSSG/GSH) and urine isoprostanes (iPF2alpha-III) levels were combined as oxidative damage markers (damage score). GSH, alpha- and gamma-tocopherol (TH) levels, and individual antioxidant capacity were combined as antioxidant defence indexes (protection score). The OXY-SCORE was computed by subtracting the protection score from the damage score. Among single parameters, T-MDA and iPF2alpha-III significantly correlated with age; only GSH and both tocopherols correlated with male gender in healthy subjects. The OXY-SCORE was positively associated with age (p=0.004) and male gender (p=0.03). As expected, the OXY-SCORE was higher in CAD with a very significant p-value (<0.0001), after adjusting for age, gender and smoking. Combining different markers can potentially provide a powerful index in the evaluation of oxidative stress related to age, gender and CAD status.     

Prepubertal children with suitable fitness and physical activity present reduced risk of oxidative stress

To assess the impact of fitness status and physical activity on oxidative stress in prepubertal children, we measured selected biomarkers such as protein carbonyls (PC), lipid peroxidation products, and total nitrites, as well as the antioxidant system: total glutathione (TG), oxidized glutathione (GSSG), reduced glutathione (GSH), superoxide dismutase activity, and glutathione peroxidase. A total of 132 healthy children ages 7-12, at prepubertal stage, were classified into two groups according to their fitness level: low fitness (LF) and high fitness (HF). They were observed while engaged in an after-school exercise program, and a questionnaire was created to obtain information on their physical activity or sedentary habits. Plasma and red blood cells were obtained to analyze biomarkers. Regarding oxidative stress markers, the LF group and the sedentary group showed higher levels of TG and GSSG and a lower GSH/GSSG ratio than the HF group and the children engaged in physical activity. A negative association was found between PC and GSSG and TG and between TG and the GSH/GSSG ratio. Moreover, a negative correlation was found between GSSG and fitness, with a positive correlation with the GSH/GSSG ratio. TG, GSSG, and the GSH/GSSG ratio seem to be reliable markers of oxidative stress in healthy prepubertal children with low fitness or sedentary habits. This research contributes to the recognition that an adequate level of fitness and recreational physical activity in childhood leads to better health and oxidative status.     

Age- and gender-related oxidative status determined in healthy subjects by means of OXY-SCORE,a potential new comprehensive index

Oxidative stress has been related to various diseases, gender and ageing, and has been measured by various markers. The authors developed a procedure to compute a global oxidative stress index (OXY-SCORE), reflecting both oxidative and antioxidant markers in healthy subjects. Its performance was tested in relation to age and gender and in coronary artery disease (CAD) patients. Eighty-two healthy subjects and 20 CAD patients were enrolled. Plasma free and total malondialdehyde (F- and T-MDA), glutathione disulphide/reduced form ratio (GSSG/GSH) and urine isoprostanes (iPF_2α-III) levels were combined as oxidative damage markers (damage score). GSH, α- and γ-tocopherol (TH) levels, and individual antioxidant capacity were combined as antioxidant defence indexes (protection score). The OXY-SCORE was computed by subtracting the protection score from the damage score. Among single parameters, T-MDA and iPF_2α-III significantly correlated with age; only GSH and both tocopherols correlated with male gender in healthy subjects. The OXY-SCORE was positively associated with age (p=0.004) and male gender (p=0.03). As expected, the OXY-SCORE was higher in CAD with a very significant p-value (<0.0001), after adjusting for age, gender and smoking. Combining different markers can potentially provide a powerful index in the evaluation of oxidative stress related to age, gender and CAD status.     

Supplementation with a combination of antioxidants does not affect glycaemic control, oxidative stress or inflammation in type 2 diabetes subjects

The present clinical trial examined the influence of a supplement, containing a combination of antioxidants extracted from fruit, berries and vegetables, on levels of plasma antioxidants (tocopherols, carotenoids and ascorbate), glycaemic control (blood glucose, HbA1c, insulin), oxidative stress biomarkers (F(2)-isoprostane, malondialdehyd, nitrotyrosine, 8-oxo-7, 8-dihydro-2'-deoxyguanosine, formamidopyrimidine glycosylase sites, frequency of micronucleated erythrocytes) and inflammatory markers (interleukin-6, C-reactive protein, prostaglandin F(2α)-metabolite) in type 2 diabetes. Forty subjects were randomly assigned to control, single or double dose group and completed the study. In summary, 12 weeks of antioxidant supplementation did neither affect glycaemic control nor the levels of biomarkers of oxidative stress or inflammation, despite substantially increased plasma concentrations of antioxidants. The absence of an effect may be explained by the selected study subjects with relatively well-controlled diabetes, a high intake of fruit and vegetable and levels of plasma antioxidants, biomarkers of oxidative stress and inflammatory markers comparable to those found in healthy subjects.     

Plasma markers of oxidative stress are uncorrelated in a wild mammal

Oxidative stress, which results from an imbalance between the production of potentially damaging reactive oxygen species versus antioxidant defenses and repair mechanisms, has been proposed as an important mediator of life‐history trade‐offs. A plethora of biomarkers associated with oxidative stress exist, but few ecological studies have examined the relationships among different markers in organisms experiencing natural conditions or tested whether those relationships are stable across different environments and demographic groups. It is therefore not clear to what extent studies of different markers can be compared, or whether studies that focus on a single marker can draw general conclusions regarding oxidative stress. We measured widely used markers of oxidative damage (protein carbonyls and malondialdehyde) and antioxidant defense (superoxide dismutase and total antioxidant capacity) from 706 plasma samples collected over a 4‐year period in a wild population of Soay sheep on St Kilda. We quantified the correlation structure among these four markers across the entire sample set and also within separate years, age groups (lambs and adults), and sexes. We found some moderately strong correlations between some pairs of markers when data from all 4 years were pooled. However, these correlations were caused by considerable among‐year variation in mean marker values; correlation coefficients were small and not significantly different from zero after accounting for among‐year variation. Furthermore, within each year, age, and sex subgroup, the pairwise correlation coefficients among the four markers were weak, nonsignificant, and distributed around zero. In addition, principal component analysis confirmed that the four markers represented four independent axes of variation. Our results suggest that plasma markers of oxidative stress may vary dramatically among years, presumably due to environmental conditions, and that this variation can induce population‐level correlations among markers even in the absence of any correlations within contemporaneous subgroups. The absence of any consistent correlations within years or demographic subgroups implies that care must be taken when generalizing from observed relationships with oxidative stress markers, as each marker may reflect different and potentially uncoupled biochemical processes.     

Individual consistency and covariation of measures of oxidative status in greenfinches

Oxidative stress results from a mismatch between production of reactive oxygen species (ROS) and the organism's capacity to mitigate their damaging effects by building up sufficient antioxidant protection and/or repair mechanisms. Because ROS production is a universal consequence of cellular metabolism and immune responses, evolutionary animal ecologists have become increasingly interested in involvement of oxidative stress as a proximate mechanism responsible for the emergence of trade-offs related to the evolution of life-history and signal traits. Among the most practical problems pertinent to ecological research on oxidative stress is finding a combination of biomarkers of oxidative status that can be applied to typical wild animal models such as small birds, mammals, and reptiles. This study describes covariation and individual consistency of eight parameters of oxidative status in a small passerine bird, wild-caught captive greenfinch (Carduelis chloris). We measured two markers of plasma antioxidant potential--total antioxidant capacity (TAC) and oxygen radical absorbance (OXY)--and concentrations of one lipophilic (carotenoids) and two hydrophilic (uric acid and ascorbate) antioxidants in plasma. We also measured total glutathione (GSH) concentration and superoxide dismutase (SOD) activity in erythrocytes. Oxidative damage was assessed on the basis of plasma malondialdehyde (MDA) levels, measured by high-performance liquid chromatography. Plasma carotenoids, TAC, and erythrocyte GSH showed significant individual consistency over an 8-d period, indicating that those variables reflected more persistent differences between individuals than plasma OXY, MDA, and uric acid. We did not detect any strong or moderate correlations between the studied parameters, which suggests that all of these biomarkers contain potentially unique information. Injection of a synthetic mimetic of SOD and catalase--EUK-134--did not affect any of the parameters of oxidative status. Capability of phagocytes to produce oxidative burst was not associated with MDA, indicating that under our experimental conditions, ROS production by phagocytes was not a strong determinant of oxidative damage. Altogether these findings suggest that attempts to characterize oxidative balance should use a wide range of biomarkers, and further studies of oxidative status in wild animals may benefit from the experimental induction of oxidative stress.     

Supplementation with a combination of antioxidants does not affect glycaemic control,oxidative stress or inflammation in type 2 diabetes subjects

Abstract

The present clinical trial examined the influence of a supplement, containing a combination of antioxidants extracted from fruit, berries and vegetables, on levels of plasma antioxidants (tocopherols, carotenoids and ascorbate), glycaemic control (blood glucose, HbA1c, insulin), oxidative stress biomarkers (F₂-isoprostane, malondialdehyd, nitrotyrosine, 8-oxo-7, 8-dihydro-2′-deoxyguanosine, formamidopyrimidine glycosylase sites, frequency of micronucleated erythrocytes) and inflammatory markers (interleukin-6, C-reactive protein, prostaglandin F_2α-metabolite) in type 2 diabetes. Forty subjects were randomly assigned to control, single or double dose group and completed the study. In summary, 12 weeks of antioxidant supplementation did neither affect glycaemic control nor the levels of biomarkers of oxidative stress or inflammation, despite substantially increased plasma concentrations of antioxidants. The absence of an effect may be explained by the selected study subjects with relatively well-controlled diabetes, a high intake of fruit and vegetable and levels of plasma antioxidants, biomarkers of oxidative stress and inflammatory markers comparable to those found in healthy subjects.     

Oxidative stress and antioxidant defence in a healthy nonagenarian population

Results on oxidative markers during ageing are not consistent throughout the scientific literature; however, successful ageing may depend on better ability to cope with oxidative stress. A previous study of ours showed that successful ageing could actually be related to enhanced response to oxidatively modified proteins. In this study, a healthy nonagenarian population (OVER-90) was examined for various blood oxidative biomarkers and compared with a healthy population of blood donors (age range, 23-66 years). Blood glutathione, both total (tGSH) and oxidised (GSSG), and total plasmatic antioxidant status were maintained in the OVER-90 at a level similar to the control population. Sulphydryl (sulfhydryl) groups and glutathione peroxidase (GPx) were instead decreased. The results are discussed in a possible unifying view: the OVER-90 population could possess a globally preserved antioxidant ability, though some signs of oxidative damage are present and some structures could be 'sacrificed' in order to keep the redox equilibrium.     

Variability of oxidative stress biomarkers in hemodialysis patients

Abstract

Oxidative stress biomarkers may have a role in the future to assist clinical decisions regarding the use of antioxidant therapies and their efficacy. The aims of this study were to evaluate the within and between-individual variability of plasma oxidative stress biomarkers and investigate factors affecting their variability. Plasma F2-isoprostanes and protein carbonyls were measured in 14 hemodialysis patients every 2 weeks for 10 weeks. Within-individual coefficients of variation (CVs) were isoprostanes?=?30.4% (range?=?6.1–66.7%) and protein carbonyls?=?16.3% (8.4–29.5%). Between-individual CVs were isoprostanes?=?34.4% (28.9–40.2%) and protein carbonyls?=?19.5% (15.6–24.5%). There were no significant (p?>?0.05) relationships between the oxidative stress biomarkers and dietary antioxidant intake, medications, clinical and demographic parameters.     

Biological variability of superoxide dismutase and glutathione peroxidase in blood

The enzymatic antioxidant defences of mammalian cells include copper-zinc superoxide dismutase (SOD)(Cu Zn-SOD; EC 1.15.1.1) which catalyses the dismutation of superoxide anions (O₂^.-) to hydrogen peroxide(H², O²)and a seleno-dependent glutathione peroxidase (GSH-px) (GSH-px; EC 1.11.1.9) which catalyses the degradation of H²O² to H²O and O². The measurement of these enzyme activities is often used as a possible biological index of oxidative stress in various clinical conditions. Complete understanding of such information requires knowledge of the random biological fluctuation of the enzyme activity which occurs in each individual. In the present investigation we examined this normal variability in 12 healthy volunteers (four women and eight men) aged 23–45 years, over 6 months. The intra-individual coefficients of variation (estimated using analysis of variance techniques) were 15% (SOD) and 13% (GSH-px). The analytical goal for imprecision was achieved for both enzymes, i.e. it was less than one half of the measured intra-individual variation. Both enzymes showed marked individuality, indicating that an individual's reference values are more useful than population-based data. The critical difference required for significant changes in serial results is 45% for SOD and 40% for GSH-px.     

Evaluation of a urinary multi-parameter biomarker set for oxidative stress in children,adolescents and young adults

The involvement of reactive oxygen species (ROS) and oxidative stress in pediatric diseases is an important concern, but oxidative stress status in healthy young subjects and appropriate methods for its measurement remain unclear. This study evaluated a comprehensive set of urinary biomarkers for oxidative stress in healthy children, adolescents and young adults. Results show that urinary excretion of acrolein–lysine, 8-hydroxy-2′-deoxyguanosine (8-OHdG), nitrite/nitrate and pentosidine were highest in the youngest subjects and decreased to constant levels by early adolescence. Urinary acrolein–lysine, 8-OHdG, nitrite/nitrate and pentosidine showed significant inverse correlations with age, but pyrraline did not change significantly with age. No significant differences in biomarkers were apparent between males and females. Younger subjects grow rapidly and sustain immune activation, and are probably exposed to high concentrations of ROS and nitric oxide. Consequently, they are more vulnerable to oxidation of lipids, proteins, DNA and carbohydrates. Normal reported values in this study are a basis for future studies of disease mechanisms involving oxidative stress and for future trials using antioxidant therapies for oxidative stress-related diseases in the pediatric field.     

Molecular bases of the treatment of Alzheimer's disease with antioxidants: prevention of oxidative stress

Alzheimer's disease is associated with a systemic oxidative stress situation which can be followed in vivo by determining biomarkers such as plasma lipoperoxides and TBARS levels and the oxidation degree of glutathione in red blood cells. It has been observed that Alzheimer's patients show an increased level of plasma TBARS, which indicates a higher free radical oxidation of plasma unsaturated phospholipids, and an increased oxidation of red blood cells glutathione, which indicates oxidative stress in peripheral cells. This latter, glutathione oxidation, was found to correlate statistically with the cognitive status of the patients. Treatment with vitamin E resulted in an improved cognitive performance only of those patients in which the tocopherol acted as an antioxidant, according to blood indicative markers of oxidative stress. Indeed, the effect of vitamin E on Alzheimer's disease patients showed considerable variations both in its antioxidant function and in its capacity to improve cognitive functions. An important conclusion from the reported results is that epidemiological or clinical studies that aim to test the effect of antioxidant supplementation on given functions should include the determination of the antioxidant status of the patients by the measurement of blood markers of oxidative stress.     

Do the serum oxidative stress biomarkers provide a reasonable index of the general oxidative stress status?

The oxidant status of an individual is assessed by determining a group of markers in noninvasive samples. One limitation when measuring these biomarkers is that they do not give information about tissue localization of oxidative stress. The present study was undertaken to establish whether the serum oxidative stress biomarkers are indicative of oxidative stress in tissues of an individual. To accomplish this, we determined a few generic markers of oxidation in serum and tissues of six groups of rats treated experimentally, to modulate their oxidative stress status. The correlation between serum and tissue levels was calculated for each marker. Also, for each tissue, the correlation between the values of these oxidative stress biomarkers was analysed. Our results show that only lipid peroxides in serum could be useful to predict the oxidative stress in tissues. No correlation was found between any of the oxidative stress markers in serum.