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Provides methods for a range of common sperm function tests that are structured as standard operating procedures (SOPs) for easy use at the bench. Methods are focussed on objectivity, robustness, standardized reporting, controlling the risk of errors, and minimizing measurement uncertainty. Includes sperm hyperactivation, acrosome reaction testing, and sperm-zone pellucida binding tests (hemi-zona and competititve binding formats). A protocol for using the sperm survival test in also provided. Limitations of the hyaluronan bibding assay, and of sperm fertilizing ability testing using zona pellucida-free hanster oocytes, are summarized.
Patients with diabetes continue to suffer from impaired visual performance before the appearance of overt damage to the retinal microvasculature and later sight-threatening complications. This diabetic retinopathy (DR) has long been thought to start with endothelial cell oxidative stress. Yet newer data surprisingly finds that the avascular outer retina is the primary site of oxidative stress before microvascular histopathology in experimental DR. Importantly, correcting this early oxidative stress is sufficient to restore vision and mitigate the histopathology in diabetic models. However, translating these promising results into the clinic has been stymied by an absence of methods that can measure and optimize anti-oxidant treatment efficacy in vivo. Here, we review imaging approaches that address this problem. In particular, diabetes-induced oxidative stress impairs dark–light regulation of subretinal space hydration, which regulates the distribution of interphotoreceptor binding protein (IRBP). IRBP is a vision-critical, anti-oxidant, lipid transporter, and pro-survival factor. We show how optical coherence tomography can measure subretinal space oxidative stress thus setting the stage for personalizing anti-oxidant treatment and prevention of impactful declines and loss of vision in patients with diabetes.
Magnetic resonance is a field that has expanded to a range of disciplines and applications, both in basic research and in its applications, and polarized targets have played an important role in this growth. This volume covers the range of disciplines required for understanding polarized targets, focusing in particular on the theoretical and technical developments made in dynamic nuclear polarization (DNP), NMR polarization measurement, high-power refrigeration, and magnet technology. Beyond particle and nuclear physics experiments, dynamically polarized nuclei have been used for experiments involving structural studies of biomolecules by neutron scattering and by NMR spectroscopy. Emerging applications in MRI are also benefiting from the sensitivity and contrast enhancements made possible by DNP or other hyperpolarization techniques. Topics are introduced theoretically using language and terminology suitable for scientists and advanced students from a range of disciplines, making this an accessible resource to this interdisciplinary field.
Paraquat is known to affect all green plants and other eukaryotic organisms including mammalian cells. The aim of this study was to improve the understanding of paraquat toxicity in nonphotosynthetic plant cells using dark-grown kidney bean cells in tissue culture. It is shown that uptake of paraquat is an active process, and that paraquat inhibits cell growth, reduces DNA synthesis, and inhibits the activity of hydroxypyruvate reductase while enhancing the activity of glutathione reductase which is involved in cellular defense against oxidant stress. Additionally, it is demonstrated that iron ions are involved in paraquat toxicity. We conclude that uptake of paraquat into cells is via polyamine channels and that the deleterious effects of paraquat on these nonphotosynthetic cells are mediated by iron.
Early physiological mechanisms that occur in crop plants in response to neighboring weeds are not well understood. In this experiment, it was hypothesized that, in the absence of direct competition for resources, low red to far red ratio (R:FR) reflected from neighboring weeds will modulate the phenylpropanoid pathway, increase hydrogen peroxide (H2O2), and up-regulate the expression of ethylene biosynthesis and auxin transport genes. Laboratory experiments were conducted under conditions of nonlimiting resources using perennial ryegrass as a model weed species. We discovered that the detection by phytochrome (Phy) of low R:FR signals reflected from both biological and nonbiological sources triggered an up-regulation of ethylene biosynthesis genes and stimulated an auxin transport gene. The low R:FR also modulated the phenylpropanoid pathway resulting in a reduction in anthocyanin content and an enhancement of lignin synthesis. The presence of neighboring weeds also caused an accumulation of H2O2 in the first leaf and crown root tissues of the maize seedling. Stomata were observed to be closed as H2O2 accumulated in leaf tissue. This is the first study to report the modulation of phenylpropanoid pathway and the accumulation of H2O2 attributed to low R:FR. We further suggest that these physiological changes that occur in response to early weed competition result in a physiological cost to the crop plant, which contributes to the rapid loss in yield observed in weed competition studies conducted under field conditions.
Introduction. Elaeagnus latifolia Linn. is a type of wild
edible fruit found in northeast India, Thailand and also in Vietnam. Although the fruit is
reported to be a source of vitamins, minerals, essential fatty acids and other bioactive
compounds, only a few studies have been concerned with the antioxidant activity of this
plant. Materials and methods. Our study revealed in vitro
antioxidant and free radical scavenging activity of 70% methanolic extract of
Elaeagnus latifolia Linn. (ELME). Various tests including
identification and quantification of phytochemicals, total antioxidant activity, hydroxyl
radical, superoxide radical, singlet oxygen, hypochlorous acid scavenging, reducing power
and DNA protection assays were performed. Results and discussion. Among the
tests, ELME scavenged superoxide radical [IC50 = (150.78 ± 4.2) μg×mL–1], hydroxyl radical
[IC50 = (238.09 ± 11.63) μg×mL–1] and protected pUC18 DNA [P50 = (695.91
± 15.84) μg×mL–1]; P50 signifies the concentration for 50% protection . The fruit is found
to be a source of minute amounts of carbohydrates, ascorbic acid, tannins, phenolics and
flavonoids. HPLC data showed that purpurin, tannic acid, quercetin, catechin, reserpine
and rutin are present in ELME. Conclusion. Our results provide evidence that
70% methanol extract of E. latifolia Linn. acts as a promising
antioxidant as well as DNA protector, which is partly due to the phenolic and flavonoid
compounds present in it.
Cisplatin is an effective chemotherapeutic agent commonly used in the treatment of malignant tumours, but ototoxicity is a significant side effect.
Objectives:
To discuss the mechanisms of cisplatin ototoxicity and subsequent cell death, and to present the results of experimental studies.
Material and methods:
We conducted a systematic search for data published in national and international journals and books, using the Medline, SciELO, Bireme, LILACS and PubMed databases.
Results:
The nicotinamide adenine dinucleotide phosphate oxidase 3 isoform (also termed NOX3) seems to be the main source of reactive oxygen species in the cochlea. These reactive oxygen species react with other molecules and trigger processes such as lipid peroxidation of the plasma membrane and increases in expression of the transient vanilloid receptor potential 1 ion channel.
Conclusion:
Cisplatin ototoxicity proceeds via the formation of reactive oxygen species in cochlear tissue, with apoptotic cell death as a consequence.
There are many well-known aetiological mechanisms of presbyacusis, and free radicals have been shown to play an important role. This study aimed to evaluate the effect of antioxidant agents on the hearing threshold of patients with presbyacusis.
Methods:
One hundred and twenty individuals were divided into four groups and received one of the following treatment schemes: ginkgo biloba dry extract, α-lipoic acid plus vitamin C, papaverine chlorhydrate plus vitamin E, or placebo. All participants were evaluated at recruitment and after six months, using pure tone audiometry (at isolated and average frequencies), speech recognition threshold and percentage index of speech recognition.
Results:
The various treatments had no effect on any of the evaluated measures of hearing, either between groups or over time.
Conclusion:
There was no statistically significant change in the hearing threshold after treatment with any of the tested drugs, during the study period.
Epidemiological studies have reported a greater reduction in cardiovascular risk and metabolic disorders associated with diets rich in polyphenols. The antioxidant effects of polyphenols are attributed to the regulation of redox enzymes by reducing reactive oxygen species production from mitochondria, NADPH oxidases and uncoupled endothelial NO synthase in addition to also up-regulating multiple antioxidant enzymes. Although data supporting the effects of polyphenols in reducing oxidative stress are promising, several studies have suggested additional mechanisms in the health benefits of polyphenols. Polyphenols from red wine increase endothelial NO production leading to endothelium-dependent relaxation in conditions such as hypertension, stroke or the metabolic syndrome. Numerous molecules contained in fruits and vegetables can activate sirtuins to increase lifespan and silence metabolic and physiological disturbances associated with endothelial NO dysfunction. Although intracellular pathways involved in the endothelial effects of polyphenols are partially described, the molecular targets of these polyphenols are not completely elucidated. We review the novel aspects of polyphenols on several targets that could trigger the health benefits of polyphenols in conditions such as metabolic and cardiovascular disturbances.
According to the free radical theory, aging is caused by the damaging effects of oxygen radicals. These agents are produced continuously and they become toxic when their intra-cellular levels become elevated. Although well-known for the damage they cause, recent studies indicate that free radicals may also have a beneficial side regulating some of the processes within a cell. With aging, however, it seems that the ability to control endogenous concentrations declines, and rising levels become progressively more harmful. In addition to the primary changes, the effects of free radical damage should also be examined at the secondary level. Because of its importance in regulating homeostatic mechanisms, it is possible that in this regard the hypothalamus may play a crucial role in the aging process. Progressive loss of function in this centre may lead to systemic changes which cause widespread disruption throughout the organism.
The free radical theory of aging is critically discussed. This review deals with the biological effects of free radicals during the aging process, the relationship between aging and diseases and how they both may be affected by free radicals, and the effects of free radicals on the human lifespan. The paper emphasizes aspects which require additional investigation.
Changes in plasma endogenous and dietary antioxidants and oxidative stress markers were studied following two 90 min elite female soccer games separated by 72 h of either active or passive recovery. The active recovery group (n 8) trained for 1 h at 22 and 46 h after the first game (low-intensity cycling and resistance training), while the passive group rested (n 8). Blood samples were taken before the games; immediately after the games; 21, 45 and 69 h after the first game; and immediately after the second game. The oxidative stress markers and antioxidants were not affected by active recovery. The oxidative stress marker GSSG increased by the same extent after both the games, while the lipid peroxidation marker diacron-reactive oxygen metabolite remained unchanged. The endogenous antioxidants total glutathione and uric acid and ferric reducing/antioxidant power increased immediately after both the games with the same amplitude, while increases in cysteine, cysteine–glycine and total thiols reached significant levels only after the second game. The changes in dietary antioxidants after the first game were either rapid and persistent (tocopherols and ascorbic acid (AA) increased; polyphenols decreased) or delayed (carotenoids). This resulted in high pre-second game levels of tocopherols, AA and carotenoids. Polyphenols returned to baseline at 69 h, and were not affected by the second game. In conclusion, the soccer-associated dietary antioxidant defence, but not the endogenous antioxidant defence, is persistent. Similar acute oxidative stress and endogenous antioxidant responses and dissimilar dietary antioxidant reactions occur during two repeated female soccer games. Finally, the complex antioxidant response to soccer is not affected by active recovery training.
The reaction of free radicals with polyunsaturated fatty acids (PUFAs) initiates a chain-reaction process known as lipid peroxidation in living systems and oxidative rancidity in foods. PUFAs are more susceptible to oxidation, resulting in the formation of toxic products. As embryonic tissues are characterised by high concentration of PUFAs, the use of antioxidants in broiler breeder diets improves chick oxidative status. Eggs ‘in shell’ are considered resistant to oxidative rancidity, however studies showed that yolk lipids within table eggs oxidise during storage and this oxidation is influenced by time and temperature of storage and the degree of unsaturation of yolk fatty acids. Thus, experiments into supplementing laying hen diet with antioxidants have been conducted to preserve the table eggs nutritional value. The literature shows that vitamin E has antioxidant activity in the egg yolk, protecting embryonic tissues efficiently during incubation and in the first days of a chick's life. However, these studies have not considered the effect of this vitamin on incubation yield. Without this economic variable, the positive results observed in the embryo and for chick oxidative status have little value for application in the field, mainly because the concentrations recommended in studies are much higher than those used in practice. Other antioxidants, e.g. canthaxanthin, act in synergy with vitamin E, however, the high cost of these ingredients could prevent its use for broiler breeders. This review discusses the effect of vitamins (especially vitamin E) in broiler breeder's and laying hens nutrition on oxidative stability of embryonic tissues and table egg yolk lipids, respectively.
Objective: Numerous studies have revealed the presence of oxidative stress in parasitic infections. However, such studies were lacking in the Malaysian population. Previously, we have provided evidence that oxidative stress is elevated in Malaysians infected with intestinal parasites. Stool examinations revealed that about 47·5% of them were infected with the polymorphic protozoa, Blastocystis hominis. However, they were found to have mixed infection with other intestinal parasites. Methodology: Therefore, in order to investigate the role of B. hominis alone in affecting oxidative stress status, here we compared the levels of oxidative stress biomarkers in urine and blood samples between uninfected and B. hominis-infected rats. Results: Infected rats exhibited elevated levels of oxidative indices namely advanced oxidative protein products (AOPP), hydrogen peroxide (H2O2) and lipid hydroperoxide (LHP) indicating that their overall oxidative damage level was higher. Ferric reducing antioxidant power (FRAP) was elevated at the initial stage of infection but decreased significantly during the last week of study duration suggesting that the antioxidant status of the host may be overwhelmed by oxidative damage. Conclusion: To date, this is the first comprehensive in vivo study to provide evidence for Blastocystis infection to correlate with significant oxidative burst leading to oxidative stress.
High dietary intake of fats has been thought to be one of the major risk factors for the development of CVD. Less is known about the possible influence of fats from various sources on haemorheological abnormalities, which are considered an important factor in the pathogenesis of these diseases. The goal of the present study was to investigate effects of high-fat diets enriched in unsaturated fatty acids (USFA), SFA or trans-fatty acids (TFA), respectively, on haemorheological parameters in rats. Wistar female rats were divided into four groups and fed diets based on the AIN-93M formulation containing approximately 10 % energy from soyabean oil (control group) or 40 % energy from soyabean oil (USFA), palm oil (SFA) and vegetable shortening (TFA) for 8 weeks. The results showed that rats fed high-fat diets exhibited significant increases in serum TAG levels (P < 0·01), plasma viscosity (P < 0·01), whole blood viscosity (P < 0·01) and internal viscosity (P < 0·01) compared to the controls. The TFA group showed a significant decrease in erythrocyte deformability (P < 0·01) and increase in internal viscosity (P < 0·01) compared with the other groups. In addition, a significant increase in blood levels of free radicals (P < 0·01) was found in the TFA group, suggesting that the attack of oxygen-free radicals could be responsible for the impaired erythrocyte deformability. These impairments could be partly responsible for the development of various circulatory disorders. The present haemorheological study provides additional insights into the potential adverse effects of trans-fat and high-fat diets on haemorheological parameters.
A 3-week-feeding study (1–21 d post-hatch) was conducted to evaluate the efficacy of total curcuminoids (TCMN), as an antioxidant, to ameliorate the adverse effects of aflatoxin B1 (AFB1) in broiler chickens. Turmeric powder (Curcuma longa L.) that contained 2·55 % TCMN was used as a source of TCMN. Six cage replicates of five chicks each were assigned to each of six dietary treatments, which included: basal diet; basal diet supplemented with 444 mg/kg TCMN; basal diet supplemented with 1·0 mg/kg AFB1; basal diet supplemented with 74 mg/kg TCMN and 1·0 mg/kg AFB1; basal diet supplemented with 222 mg/kg TCMN and 1·0 mg/kg AFB1; basal diet supplemented with 444 mg/kg TCMN and 1·0 mg/kg AFB1. The addition of 74 and 222 mg/kg TCMN to the AFB1 diet significantly (P < 0·05) improved weight gain and feed efficiency. Increase (P < 0·05) in relative liver weight in birds fed AFB1 was significantly reduced (P < 0·05) with the addition of 74, 222 and 444 mg/kg TCMN to the AFB1 diet. The inclusion of 222 mg/kg TCMN ameliorated the adverse effects of AFB1 on serum chemistry in terms of total protein, albumin and γ-glutamyl transferase activity. The decreased antioxidant functions due to AFB1 were also alleviated by the inclusion of 222 mg/kg TCMN. It is concluded that the addition of 222 mg/kg TCMN to the 1·0 mg/kg AFB1 diet demonstrated maximum antioxidant activity against AFB1.
The role of free radical-induced damage as a cause of loss of vigour in seeds is by no means resolved. In this contribution, the effects of environmental treatments known to reduce viability rapidly were compared with the effects of long-term, low-temperature storage on germination, hypocotyl growth and free radical accumulation and lipid peroxidation in soybean seeds. Accelerated aging was achieved by incubating seeds at 35°C and 1% relative humidity over H2SO4 for up to 69 days in the light and in darkness. In contrast, seeds under long-term storage were maintained at 5°C and 6% moisture content in darkness for up to 6 years. At 35°C there were rapid and significant reductions in rates of seed germination and hypocotyl extension. Loss of viability and declining vigour were associated with increases in lipid peroxidation and free radical build-up but the latter, surprisingly, was largely confined to the testa rather than the cotyledon. Exposure to light greatly enhanced lipid peroxidation and increased organic free radical accumulation in the translucent testas of seeds, but not in the cotyledons. Similar responses to light were recorded in testas detached from seeds. These results show that in soybean the testa is a significant locus of free radical degenerative events induced by high temperature combined with low moisture.
The desiccation tolerance of Zizania palustris seeds has been debatable, but recently it has emerged that survival to low moisture contents is a function of the temperature during dehydration. Survival of dehydration is greatest at 25–30°C and viability declines as the dehydration temperature is reduced. On the other hand, extended hydrated chilling (stratification) is required to break dormancy. The present contribution examines the ultrastructural condition of embryonic axes of Zizania palustris after dehydration at various temperatures, and following reimbibition and stratification. Axis cells sustained least damage when dehydration was carried out at 25°C and ultrastructural deterioration was more severe with lower temperatures during water loss. Damage sustained as a result of unfavourably low dehydration temperatures was visible when seeds were fixed from the dry state and was generally exacerbated during fully imbibed stratification. However, 25°C represented the optimum dehydration temperature; seeds that had been dried at 30°C also showed considerable ultrastructural disturbance when fixed from the dry state. This was largely reversed during fully imbibed stratification, although signs of the damage that had been sustained still persisted. These observations are in keeping with the germination behaviour of Z. palustris seeds dehydrated at temperatures above and below 25°C. The results are discussed in terms of the predominance of one of two mechanisms of membrane deterioration, depending on the dehydration temperature, which are probably not mutually exclusive. It is hypothesized that, at temperatures above 25°C, damage by free-radical-mediated events may predominate, whereas, at temperatures below the optimum, irreversible lipid-phase transitions may be the major factor resulting in membrane damage.
Electron paramagnetic resonance studies of high vigour (99% viable) and low vigour (38% viable) dry embryos of rice seeds (Oryza sativa L.) stored in a natural (warm and humid) environment were carried out. Loss in viability due to hot and humid conditions was found to be correlated with a decrease in free radical levels. The free radicals could be carbon-based and derived from quinones. Presence of the active oxygen scavenging enzyme, superoxide dismutase (EC 1.15.1.1) in embryos isolated from dry seeds showed a positive corrrelation with the state of vigour or viability. Anodic peroxidase (EC 1.11.1.7) activity in imbibed seeds also declined with the decline in vigour and viability. It is concluded that the deterioration of cells in the embryonic axis depends on the balance between free radical accumulation and the activity of active oxygen-scavenging enzymes which constitutes the active oxygen scavenging system (AOSS) during early imbibition. During prolonged storage under hot and humid conditions, cumulative effects of macromolecular damage due to oxidative chain products, compounded with the loss of enzyme activity, result in the final catastrophe, the death of the embryo.
The origin and significance of free radicals is described in the broad setting of animal and plant biology and in the specific context of seeds and seed viability. Evidence is given that free radicals play a central if not causal role in promoting molecular damage under the widest range of environmental stresses and induced ageing in mature plant tissuesIn ungerminated seeds, the evidence is less certain. The reason, we argue, is that many attempts to measure free radical processes in seeds are set not against gradients of damage or ageing (as they are in most other biological tissues) but against one of only two options, either seed germinability or mortality. Because free radical reactions differ quantitatively and qualitatively in living and dead tissues attempts to correlate radicalmediated damage with the viability of a population of seeds may be unreliable particularly when measurements are made at uncertain intervals after death. Despite this, the evidence that seeds, uniquely, are exempt from the ravages of oxygen assault is weak. Instead it ismore probable that oxygen plays a central role in seed mortality and may have significance in the evolution of seed persistence.