Neutrophil activation, antioxidant supplements and exercise-induced oxidative stress

Neutrophils produce free radicals known as reactive oxygen species (ROS), which assist in the clearance of damaged host tissue. Tissue damage may occur during exercise due to muscle damage, thermal stress and ischaemia/reperfusion. When produced in excess, neutrophil-derived ROS may overwhelm the body's endogenous antioxidant defence mechanisms, and this can lead to oxidative stress. There is increasing evidence for links between oxidative stress and a variety of pathological disorders such as cardiovascular diseases, cancer, chronic inflammatory diseases and post-ischaemic organ injury. A small number of studies have investigated whether there is a link between neutrophil activation and oxidative stress during exercise. In this review, we have summarised the findings of these studies. Exercise promotes the release of neutrophils into the circulation, and some evidence suggests that neutrophils mobilised after exercise have an enhanced capacity to generate some forms of ROS when stimulated in vitro. Neutrophil activation during exercise may challenge endogenous antioxidant defence mechanisms, but does not appear to increase lipid markers of oxidative stress to any significant degree, at least in the circulation. Antioxidant supplements such as N-acetylcysteine are effective at attenuating increases in the capacity of neutrophils to generate ROS when stimulated in vitro, whereas vitamin E reduces tissue infiltration of neutrophils during exercise. Free radicals generated during intense exercise may lead to DNA damage in leukocytes, but it is unknown if this damage is the result of neutrophil activation. Exercise enhances the expression of inducible haem (heme)-oxygenase (HO-1) in neutrophils after exercise, however, it is uncertain whether oxidative stress is the stimulus for this response.

Antioxidants in Athlete's Basic Nutrition: Considerations towards a Guideline for the Intake of Vitamin C and Vitamin E

Antioxidants in acute physical exercise and exercise training remain a hot topic in sport nutrition, exercise physiology and biology, in general (Jackson, 2008; Margaritis and Rousseau, 2008; Gomez-Cabrera et al., 2012; Nikolaidis et al., 2012). During the past few decades, antioxidants have received attention predominantly as a nutritional strategy for preventing or minimising detrimental effects of reactive oxygen and nitrogen species (RONS), which are generated during and after strenuous exercise (Jackson, 2008, 2009; Powers and Jackson, 2008). Antioxidant supplementation has become a common practice among athletes as a means to (theoretically) reduce oxidative stress, promote recovery and enhance performance (Peternelj and Coombes, 2011). However, until now, requirements of antioxidant micronutrients and antioxidant compounds for athletes training for and competing in different sport events, including marathon running, triathlon races or team sport events involving repeated sprinting, have not been determined sufficiently (Williams et al., 2006; Margaritis and Rousseau, 2008). Crucially, evidence has been emerging that higher dosages of antioxidants may not necessarily be beneficial in this context, but can also elicit detrimental effects by interfering with performance-enhancing (Gomez-Cabrera et al., 2008) and health-promoting training adaptations (Ristow et al., 2009). As originally postulated in a pioneering study on exercise-induced production of RONS by Davies et al. (1982) in the early 1980s, evidence has been increasing in recent years that RONS are not only damaging agents, but also act as signalling molecules for regulating muscle function (Reid, 2001; Jackson, 2008) and for initiating adaptive responses to exercise (Jackson, 2009; Powers et al., 2010). The recognition that antioxidants could, vice versa, interact with the signalling pathways underlying the responses to acute (and repeated) bouts of exercise has contributed important novel aspects to the continued discussion on antioxidant requirements for athletes. In view of the recent advances in this field, it is the aim of this report to examine the current knowledge of antioxidants, in particular of vitamins C and E, in the basic nutrition of athletes. While overviews on related topics including basic mechanisms of exercise-induced oxidative stress, redox biology, antioxidant defence systems and a summary of studies on antioxidant supplementation during exercise training are provided, this does not mean that this report is comprehensive. Several issues of the expanding and multidisciplinary field of antioxidants and exercise are covered elsewhere in this book and/or in the literature. Exemplarily, the reader is referred to reviews on oxidative stress (Konig et al., 2001; Vollaard et al., 2005; Knez et al., 2006; Powers and Jackson, 2008; Nikolaidis et al., 2012), redox-sensitive signalling and muscle function (Reid, 2001; Vollaard et al., 2005; Jackson, 2008; Ji, 2008; Powers and Jackson, 2008; Powers et al., 2010; Radak et al., 2013) and antioxidant supplementation (Williams et al., 2006; Peake et al., 2007; Peternelj and Coombes, 2011) in the context with exercise. Within the scope of the report, we rather aim to address the question regarding requirements of antioxidants, specifically vitamins C and E, during exercise training, draw conclusions and provide practical implications from the recent research.

Well-trained, healthy triathletes experience no adverse health risks regarding oxidative stress and DNA damage by participating in an ultra-endurance event

Also physical exercise in general is accepted to be protective, acute and strenuous exercise has been shown to induce oxidative stress. Enhanced formation of free radicals leads to oxidation of macromolecules and to DNA damage. On the other hand ultra-endurance events which require strenuous exercise are very popular and the number of participants is continuously increasing worldwide. Since only few data exists on Ironman triathletes, who are prototypes of ultra-endurance athletes, this study was aimed at assessing the risk of oxidative stress and DNA damage after finishing a triathlon and to predict a possible health risk. Blood samples of 42 male athletes were taken 2 days before, within 20 min after the race, 1, 5 and 19 days post-race. Oxidative stress marker increased only moderately after the race and returned to baseline after 5 days. Marker of DNA damage measured by the SCGE assay with and without restriction enzymes as well as by the sister chromatid exchange assay did either show no change or deceased within the first day after the race. Due to intake during the race and the release by the cells plasma concentrations of vitamin C and α-tocopherol increased after the event and returned to baseline 1 day after. This study indicates that despite a temporary increase in some oxidative stress markers, there is no persistent oxidative stress and no DNA damage in response to an Ironman triathlon in trained athletes, mainly due to an appropriate antioxidant intake and general protective alterations in the antioxidant defence system.

Conifer defence against insects: microarray gene expression profiling of Sitka spruce (Picea sitchensis) induced by mechanical wounding or feeding by spruce budworms (Choristoneura occidentalis) or white pine weevils (Pissodes strobi) reveals large

Conifers are resistant to attack from a large number of potential herbivores or pathogens. Previous molecular and biochemical characterization of selected conifer defence systems support a model of multigenic, constitutive and induced defences that act on invading insects via physical, chemical, biochemical or ecological (multitrophic) mechanisms. However, the genomic foundation of the complex defence and resistance mechanisms of conifers is largely unknown. As part of a genomics strategy to characterize inducible defences and possible resistance mechanisms of conifers against insect herbivory, we developed a cDNA microarray building upon a new spruce (Picea spp.) expressed sequence tag resource. This first-generation spruce cDNA microarray contains 9720 cDNA elements representing c. 5500 unique genes. We used this array to monitor gene expression in Sitka spruce (Picea sitchensis) bark in response to herbivory by white pine weevils (Pissodes strobi, Curculionidae) or wounding, and in young shoot tips in response to western spruce budworm (Choristoneura occidentalis, Lepidopterae) feeding. Weevils are stem-boring insects that feed on phloem, while budworms are foliage feeding larvae that consume needles and young shoot tips. Both insect species and wounding treatment caused substantial changes of the host plant transcriptome detected in each case by differential gene expression of several thousand array elements at 1 or 2 d after the onset of treatment. Overall, there was considerable overlap among differentially expressed gene sets from these three stress treatments. Functional classification of the induced transcripts revealed genes with roles in general plant defence, octadecanoid and ethylene signalling, transport, secondary metabolism, and transcriptional regulation. Several genes involved in primary metabolic processes such as photosynthesis were down-regulated upon insect feeding or wounding, fitting with the concept of dynamic resource allocation in plant defence. Refined expression analysis using gene-specific primers and real-time PCR for selected transcripts was in agreement with microarray results for most genes tested. This study provides the first large-scale survey of insect-induced defence transcripts in a gymnosperm and provides a platform for functional investigation of plant-insect interactions in spruce. Induction of spruce genes of octadecanoid and ethylene signalling, terpenoid biosynthesis, and phenolic secondary metabolism are discussed in more detail.

Cystine-mediated oxidative defence in Lactobacillus reuteri BR11

Lactobacillus reuteri BR11 possesses an abundant cystine uptake (Cyu) ABC-transporter that was previously found to be involved in a novel mechanism of oxidative defence mediated by cystine. The current study aimed to elucidate this mechanism with a focus on the role of the co-transcribed cystathionine ã-lyase (Cgl). Growth studies of wild-type L. reuteri BR11 and mutants inactivated in cgl and the cystine-binding protein encoding gene cyuC showed that in contrast to the Cyu transporter, whose inactivation led to growth arrest in aerated cultures, Cgl is not crucial for oxidative defence. However, the role of Cgl in oxidative defence became apparent in the presence of severe oxidative damage and cysteine deprivation. Cysteine was found to be protective against oxidative stress, and the action of Cgl in both cysteine biosynthesis and degradation poses a seemingly futile pathway that deprives the intracellular cysteine pool. To further characterise the relationship between Cgl activity and cysteine and their roles in oxidative defence, enzymatic assays were performed on purified Cgl, and intracellular concentrations of cysteine, cystathionine and methionine were determined. Cgl was highly active towards cystine and cystathionine and less active towards cysteine in vitro, suggesting the main function of Cgl to be cysteine biosynthesis. Cysteine was found at high concentrations in the cell, but the levels were not significantly affected by inactivation of cgl or growth under aerobic conditions. It was concluded that both anabolic and catabolic activities of Cgl towards cysteine contribute to oxidative defence, the former by maintaining an intracellular reservoir of thiol analogous to glutathione, and the latter by producing H2S which is readily secreted, thus creating a reducing extracellular environment. The significance of the Cyu transporter to the physiology of L. reuteri BR11 prompted a phylogenetic study to determine its presence in bacteria. Orthologs of the Cyu transporter that are closest matches to the Cyu transporter are only limited to several species of Lactobacillus and Leuconostoc. Outside the Lactobacillales order, the closest matching orthologs belong to Proteobacteria, and there are more orthologs in Proteobacteria than non-Lactobacillales Firmicutes, suggesting that the Cyu transporter locus was present in the ancestor of the Proteobacteria and Firmicutes, and over evolutionary time has been lost or diverged in many Firmicutes. The clustering of the Cyu transporter locus with a gene encoding a Cgl family protein is even rarer. It was only found in L. reuteri, Lactobacillus vaginalis, Weissella paramesenteroides, the Lactobacillus casei group, and several Campylobacter sp. An accompanying phylogenetic study of L. reuteri BR11 using multi-locus sequence analysis showed that L. reuteri BR11 had diverged from more than 100 strains of L. reuteri isolated from various hosts and geographical locations. However, comparison with other Lactobacillus species supported the current classification of BR11 as L. reuteri. The most closely related species to L. reuteri is L. vaginalis or Lactobacillus antri, depending on the housekeeping gene used for analysis. The close evolutionary relationship of L. vaginalis to L. reuteri and the high degree of sequence identity between the cgl-cyuABC loci in both species suggest that the Cyu system is highly likely to perform similar functions in L. vaginalis. In search of other genes that function in oxidative defence, a number of mutants which were inactivated in genes that confer increased resistance to oxidative stress in other bacteria were constructed. The genes targeted were ahpC (peroxidase component of the alkyl hydroperoxide reductase system), tpx (thiol peroxidase), osmC (osmotically induced protein C), mntH (Mn2+/Fe2+ transporter), gshA (ã-glutamylcysteine synthetase) and msrA (methionine sulfoxide reductase). The ahpC and mntH mutants had slightly lower minimum inhibitory concentrations of organic peroxides, suggesting these genes might be involved in resistance to organic peroxides in L. reuteri. However, none of the mutants exhibited growth defects in aerated cultures, in stark contrast to the cyuC mutant. This may be due to compensatory functions of other genes, a hypothesis which cannot be tested until a robust protocol for constructing markerless multiple gene deletion mutants in L. reuteri is developed. These results highlight the importance of the Cyu transporter in oxidative defence and provide a foundation for extending the research of this system in other bacteria.

Self-defence against terrorism in the post 9/11 world

In 1986 the then United States Secretary of State George Shultz asserted that: It is absurd to argue that international law prohibits us from capturing terrorists in international waters or airspace; from attacking them on the soil of other nations, even for the purpose of rescuing hostages; or from using force against states that support, train and harbor terrorists or guerrillas. At that time the United States’ claim of a right to use military force in self-defence against terrorism2 received little support from other states.3 The predominant view then was that terrorist attacks committed by private or non-state actors were a form of criminal activity to be combated through domestic and international criminal justice mechanisms. The notion that such terrorist acts should be treated as ‘armed attacks’ triggering a victim state’s right of self-defence was not accepted by the majority of states. To suggest, as Shultz had done, that a state not directly responsible for terrorist acts could have its territorial integrity violated by military action targeting terrorists located within that state, was a controversial proposition in 1986. However, some fifteen years later, when the United States and a coalition of allies launched a military campaign in Afghanistan following the 11 September 2001 (hereafter ‘9/11’) terrorist attacks, there was virtually unanimous international support for the use of force.

Israel's airstrike on Syria's Al-Kibar facility : a test case for the doctrine of pre-emptive self-defence?

This article analyses the legality of Israel’s 2007 airstrike on an alleged Syrian nuclear facility at Al-Kibar—an incident that has been largely overlooked by international lawyers to date. The absence of a threat of imminent attack from Syria means Israel’s military action was not a lawful exercise of anticipatory self-defence. Yet, despite Israel’s clear violation of the prohibition on the use of force there was remarkably little condemnation from other states, suggesting the possibility of growing international support for the doctrine of pre-emptive self-defence. This article argues that the muted international reaction to Israel’s pre-emptive action was the result of political factors, and should not be seen as endorsement of the legality of the airstrike. As such, a lack of opinio juris means the Al-Kibar episode cannot be viewed as extending the scope of the customary international law right of self-defence so as to permit the use of force against non-imminent threats. However, two features of this incident—namely, Israel’s failure to offer any legal justification for its airstrike, and the international community’s apparent lack of concern over legality—are also evident in other recent uses of force in the ‘war on terror’ context. These developments may indicate a shift in state practice involving a downgrading of the role of international law in discussions of the use of force. This may signal a declining perception of the legitimacy of the jus ad bellum, at least in cases involving minor uses of force.

A forced titration study of the antioxidant and immunomodulatory effects of Ambrotose AO supplement

Background Oxidative stress plays a role in acute and chronic inflammatory disease and antioxidant supplementation has demonstrated beneficial effects in the treatment of these conditions. This study was designed to determine the optimal dose of an antioxidant supplement in healthy volunteers to inform a Phase 3 clinical trial. Methods The study was designed as a combined Phase 1 and 2 open label, forced titration dose response study in healthy volunteers (n = 21) to determine both acute safety and efficacy. Participants received a dietary supplement in a forced titration over five weeks commencing with a no treatment baseline through 1, 2, 4 and 8 capsules. The primary outcome measurement was ex vivo changes in serum oxygen radical absorbance capacity (ORAC). The secondary outcome measures were undertaken as an exploratory investigation of immune function. Results A significant increase in antioxidant activity (serum ORAC) was observed between baseline (no capsules) and the highest dose of 8 capsules per day (p = 0.040) representing a change of 36.6%. A quadratic function for dose levels was fitted in order to estimate a dose response curve for estimating the optimal dose. The quadratic component of the curve was significant (p = 0.047), with predicted serum ORAC scores increasing from the zero dose to a maximum at a predicted dose of 4.7 capsules per day and decreasing for higher doses. Among the secondary outcome measures, a significant dose effect was observed on phagocytosis of granulocytes, and a significant increase was also observed on Cox 2 expression. Conclusion This study suggests that Ambrotose AO® capsules appear to be safe and most effective at a dosage of 4 capsules/day. It is important that this study is not over interpreted; it aimed to find an optimal dose to assess the dietary supplement using a more rigorous clinical trial design. The study achieved this aim and demonstrated that the dietary supplement has the potential to increase antioxidant activity. The most significant limitation of this study was that it was open label Phase 1/Phase 2 trial and is subject to potential bias that is reduced with the use of randomization and blinding. To confirm the benefits of this dietary supplement these effects now need to be demonstrated in a Phase 3 randomised controlled trial (RCT).

The defence of joint illegal activity must be looked at in context

In Miller v Miller (2011) 85 ALJR 480; [2011] HCA 9 the High Court examined the complex issue of joint illegal activity. The issue before the court was whether a plaintiff who had engaged in an illegal activity with the defendant may claim damages in negligence. In its decision the court analysed the cases of Henwood v Municipal Tramways Trust (SA) (1938) 60 CLR 438, Smith v Jenkins (1970) 119 CLR 397, Jackson v Harrison (1978) 138 CLR 438 and Gala v Preston (1991) 172 CLR 243.