Exercise, healthy ageing, and the potential role of small extracellular vesicles.

Extracellular vesicles (EVs) can be released from most cells in the body and act as intercellular messengers transferring information in their cargo to affect cellular function. A growing body of evidence suggests that a subset of EVs, referred to here as 'small extracellular vesicles' (sEVs), can accelerate or slow the processes of ageing and age-related diseases dependent on their molecular cargo and cellular origin. Continued exploration of the vast complexity of the sEV cargo aims to further characterise these systemic vehicles that may be targeted to ameliorate age-related pathologies. Marked progress in the development of mass spectrometry-based technologies means that it is now possible to characterise a significant proportion of the proteome of sEVs (surface and cargo) via unbiased proteomics. This information is vital for identifying biomarkers and the development of sEV-based therapeutics in the context of ageing. Although exercise and physical activity are prominent features in maintaining health in advancing years, the mechanisms responsible are unclear. A potential mechanism by which plasma sEVs released during exercise could influence ageing and senescence is via the increased delivery of cargo proteins that function as antioxidant enzymes or inhibitors of senescence. These have been observed to increase in sEVs following acute and chronic exercise, as identified via independent interrogation of high coverage, publicly available proteomic datasets. Establishing tropism and exchange of functionally active proteins by these processes represents a promising line of enquiry in implicating sEVs as biologically relevant mediators of the ageing process.

Single vesicle analysis reveals the release of tetraspanin positive extracellular vesicles into circulation with high intensity intermittent exercise.

Small extracellular vesicles (sEVs) are released from all cell types and participate in the intercellular exchange of proteins, lipids, metabolites and nucleic acids. Proteomic, flow cytometry and nanoparticle tracking analyses suggest sEVs are released into circulation with exercise. However, interpretation of these data may be influenced by sources of bias introduced by different analytical approaches. Seven healthy participants carried out a high intensity intermittent training (HIIT) cycle protocol consisting of 4 × 30 s at a work-rate corresponding to 200% of individual max power (watts) interspersed by 4.5 min of active recovery. EDTA-treated blood was collected before and immediately after the final effort. Platelet-poor (PPP) and platelet-free (PFP) plasma was derived by one or two centrifugal spins at 2500 g, respectively (15 min, room temperature). Platelets were counted on an automated haemocytometer. Plasma samples were assessed with the Exoview R100 platform, which immobilises sEVs expressing common tetraspanin markers CD9, CD63, CD81 and CD41a on microfluidic chips and with the aid of fluorescence imaging, counts their abundance at a single sEV resolution, importantly, without a pre-isolation step. There was a lower number of platelets in the PFP than PPP, which was associated with a lower number of CD9, CD63 and CD41a positive sEVs. HIIT induced an increase in fluorescence counts in CD9, CD63 and CD81 positive sEVs in both PPP and PFP. These data support the concept that sEVs are released into circulation with exercise. Furthermore, platelet-free plasma is the preferred, representative analyte to study sEV dynamics and phenotype during exercise. KEY POINTS: Small extracellular vesicles (sEV) are nano-sized particles containing protein, metabolites, lipid and RNA that can be transferred from cell to cell. Previous findings implicate that sEVs are released into circulation with exhaustive, aerobic exercise, but since there is no gold standard method to isolate sEVs, these findings may be subject to bias introduced by different approaches. Here, we use a novel method to immobilise and image sEVs, at single-vesicle resolution, to show sEVs are released into circulation with high intensity intermittent exercise. Since platelet depletion of plasma results in a reduction in sEVs, platelet-free plasma is the preferred analyte to examine sEV dynamics and phenotype in the context of exercise.

Intravascular Follistatin gene delivery improves glycemic control in a mouse model of type 2 diabetes.

Type 2 diabetes (T2D) manifests from inadequate glucose control due to insulin resistance, hypoinsulinemia, and deteriorating pancreatic ß-cell function. The pro-inflammatory factor Activin has been implicated as a positive correlate of severity in T2D patients, and as a negative regulator of glucose uptake by skeletal muscle, and of pancreatic ß-cell phenotype in mice. Accordingly, we sought to determine whether intervention with the Activin antagonist Follistatin can ameliorate the diabetic pathology. Here, we report that an intravenous Follistatin gene delivery intervention with tropism for striated muscle reduced the serum concentrations of Activin B and improved glycemic control in the db/db mouse model of T2D. Treatment reversed the hyperglycemic progression with a corresponding reduction in the percentage of glycated-hemoglobin to levels similar to lean, healthy mice. Follistatin gene delivery promoted insulinemia and abundance of insulin-positive pancreatic ß-cells, even when treatment was administered to mice with advanced diabetes, supporting a mechanism for improved glycemic control associated with maintenance of functional ß-cells. Our data demonstrate that single-dose intravascular Follistatin gene delivery can ameliorate the diabetic progression and improve prognostic markers of disease. These findings are consistent with other observations of Activin-mediated mechanisms exerting deleterious effects in models of obesity and diabetes, and suggest that interventions that attenuate Activin signaling could help further understanding of T2D and the development of novel T2D therapeutics.

Exercise dose affects the circulating microRNA profile in response to acute endurance exercise in male amateur runners.

The systemic response to exercise is dose-dependent and involves a complex gene expression regulation and cross-talk between tissues. This context ARISES the need for analyzing the influence of exercise dose on the profile of circulating microRNAs (c-miRNAs), as emerging posttranscriptional regulators and intercellular communicators. Thus, we hypothesized that different exercise doses will determine specific c-miRNA signatures that will highlight its potential as exercise dose biomarker. Nine active middle-aged males completed a 10-km race (10K), a half-marathon (HM), and a marathon (M). Blood samples were collected immediately before and after races. Plasma RNA was extracted, and a global screening of 752 microRNAs was analyzed using RT-qPCR. Three different c-miRNA profiles were defined according to the three doses. In 10K, 14 c-miRNAs were found to be differentially expressed between pre- and post-exercise, 13 upregulated and 1 downregulated. Regarding HM, 13 c-miRNAs were found to be differentially modulated, in all the cases upregulated. A total of 28 c-miRNAs were found to be differentially expressed in M, 21 overexpressed and 7 repressed after this race. We had also found 3 common c-miRNAs between 10K and M and 2 common c-miRNAs between 10K and HM. In silico analysis supported a close association between exercise dose c-miRNA profiles and cellular pathways linked to energy metabolism and cell cycle. In conclusion, we have observed that different exercise doses induced specific c-miRNA profiles. So, our results point to c-miRNAs as emerging exercise dose biomarkers and as one of regulatory mechanisms modulating the response to endurance exercise.

Redefining Tissue Crosstalk via Shotgun Proteomic Analyses of Plasma Extracellular Vesicles.

Protein signaling between tissues, or tissue cross-talk is becoming recognized as a fundamental biological process that is incompletely understood. Shotgun proteomic analyses of tissues and plasma to explore this concept are regularly challenged by high dynamic range of protein abundance, which limits the identification of lower abundance proteins. In this viewpoint article, it is highlighted how a focus on proteins contained within extracellular vesicles (EVs) not only partially addresses this issue, but can also reveal an underappreciated complexity of the circulating proteome in various physiological and pathological contexts. Furthermore, how quantitative proteomics can inform EV mediated crosstalk is highlighted and the importance of high coverage, sensitive proteomic analyses of EVs to identify both the optimal methods to isolate EV subtypes of interest and proteins that characterize them is stressed.

Role of exercise-induced hepatokines in metabolic disorders.

The health-promoting effects of physical activity to prevent and treat metabolic disorders are numerous. However, the underlying molecular mechanisms are not yet completely deciphered. In recent years, studies have referred to the liver as an endocrine organ, since it releases specific proteins called hepatokines. Some of these hepatokines are involved in whole body metabolic homeostasis and are theorized to participate in the development of metabolic disease. In this regard, the present review describes the role of Fibroblast Growth Factor 21, Fetuin-A, Angiopoietin-like protein 4, and Follistatin in metabolic disease and their production in response to acute exercise. Also, we discuss the potential role of hepatokines in mediating the beneficial effects of regular exercise and the future challenges to the discovery of new exercise-induced hepatokines.

Adipocyte-specific deletion of IL-6 does not attenuate obesity-induced weight gain or glucose intolerance in mice.

It has been suggested that interleukin-6 (IL-6) produced by adipocytes in obesity leads to liver insulin resistance, although this hypothesis has never been definitively tested. Accordingly, we did so by generating adipocyte-specific IL-6-deficient (AdipoIL-6-/-) mice and studying them in the context of diet-induced and genetic obesity. Mice carrying two floxed alleles of IL-6 (C57Bl/6J) were crossed with Cre recombinase-overexpressing mice driven by the adiponectin promoter to generate AdipoIL-6-/- mice. AdipoIL-6-/- and floxed littermate controls were fed a standard chow or high-fat diet (HFD) for 16 wk and comprehensively metabolically phenotyped. In addition to a diet-induced obesity model, we also examined the role of adipocyte-derived IL-6 in a genetic model of obesity and insulin resistance by crossing the AdipoIL-6-/- mice with leptin-deficient (ob/ob) mice. As expected, mice on HFD and ob/ob mice displayed marked weight gain and increased fat mass compared with chow-fed and ob/+ (littermate control) animals, respectively. However, deletion of IL-6 from adipocytes in either model had no effect on glucose tolerance or fasting hyperinsulinemia. We concluded that adipocyte-specific IL-6 does not contribute to whole body glucose intolerance in obese mice.

GeneXX: an online tool for the exploration of transcript changes in skeletal muscle associated with exercise.

Exercise stimulates a wide array of biological processes, but the mechanisms involved are incompletely understood. Many previous studies have adopted transcriptomic analyses of skeletal muscle to address particular research questions, a process that ultimately results in the collection of large amounts of publicly available data that has not been fully integrated or interrogated. To maximize the use of these available transcriptomic exercise data sets, we have downloaded and reanalyzed them and formulated the data into a searchable online tool, geneXX. GeneXX is highly intuitive and free and provides immediate information regarding the response of a transcript of interest to exercise in skeletal muscle. To demonstrate its utility, we carried out a meta-analysis on the included data sets and show transcript changes in skeletal muscle that persist regardless of sex, exercise mode, and duration, some of which have had minimal attention in the context of exercise. We also demonstrate how geneXX can be used to formulate novel hypotheses on the complex effects of exercise, using preliminary data already generated. This resource represents a valuable tool for researchers with interests in human skeletal muscle adaptation to exercise.

From cytokine to myokine: the emerging role of interleukin-6 in metabolic regulation.

The lack of physical activity and overnutrition in our modern lifestyle culminates in what we now experience as the current obesity and diabetes pandemic. Medical research performed over the past 20 years identified chronic low-grade inflammation as a mediator of these metabolic disorders. Hence, finding therapeutic strategies against this underlying inflammation and identifying molecules implicated in this process is of significant importance. Following the observation of an increased plasma concentration of interleukin-6 (IL-6) in obese patients, this protein, known predominantly as a pro-inflammatory cytokine, came into focus. In an attempt to clarify its importance, several studies implicated IL-6 as a co-inducer of the development of obesity-associated insulin resistance, which precedes the development of type 2 diabetes. However, the identification of IL-6 as a myokine, a protein produced and secreted by skeletal muscle to fulfil paracrine or endocrine roles in the insulin-sensitizing effects following exercise, provides a contrasting and hence paradoxical identity of this protein in the context of metabolism. We review here the literature considering the complex, pleiotropic role of IL-6 in the context of metabolism in health and disease.

Next-generation sequencing reveals that miR-16-5p, miR-19a-3p, miR-451a, and miR-25-3p cargo in plasma extracellular vesicles differentiates sedentary young males from athletes.

A sedentary lifestyle and Olympic participation are contrary risk factors for global mortality and incidence of cancer and cardiovascular disease. Extracellular vesicle miRNAs have been described to respond to exercise. No molecular characterization of young male sedentary people versus athletes is available; so, our aim was to identify the extracellular vesicle miRNA profile of chronically trained young endurance and resistance male athletes compared to their sedentary counterparts. A descriptive case-control design was used with 16 sedentary young men, 16 Olympic male endurance athletes, and 16 Olympic male resistance athletes. Next-generation sequencing and RT-qPCR and external and internal validation were performed in order to analyze extracellular vesicle miRNA profiles. Endurance and resistance athletes had significant lower levels of miR-16-5p, miR-19a-3p, and miR-451a compared to sedentary people. Taking all together, exercise-trained miRNA profile in extracellular vesicles provides a differential signature of athletes irrespective of the type of exercise compared to sedentary people. Besides, miR-25-3p levels were specifically lower in endurance athletes which defines its role as a specific responder in this type of athletes. In silico analysis of this profile suggests a role in adaptive energy metabolism in this context that needs to be experimentally validated. Therefore, this study provides for the first time basal levels of circulating miRNA in extracellular vesicles emerge as relevant players in intertissue communication in response to chronic exercise exposure in young elite male athletes.

Contraction-induced interleukin-6 gene transcription in skeletal muscle is regulated by c-Jun terminal kinase/activator protein-1.

Exercise increases the expression of the prototypical myokine IL-6, but the precise mechanism by which this occurs has yet to be identified. To mimic exercise conditions, C2C12 myotubes were mechanically stimulated via electrical pulse stimulation (EPS). We compared the responses of EPS with the pharmacological Ca(2+) carrier calcimycin (A23187) because contraction induces marked increases in cytosolic Ca(2+) levels or the classical IκB kinase/NFκB inflammatory response elicited by H(2)O(2). We demonstrate that, unlike H(2)O(2)-stimulated increases in IL-6 mRNA, neither calcimycin- nor EPS-induced IL-6 mRNA expression is under the transcriptional control of NFκB. Rather, we show that EPS increased the phosphorylation of JNK and the reporter activity of the downstream transcription factor AP-1. Furthermore, JNK inhibition abolished the EPS-induced increase in IL-6 mRNA and protein expression. Finally, we observed an exercise-induced increase in both JNK phosphorylation and IL-6 mRNA expression in the skeletal muscles of mice after 30 min of treadmill running. Importantly, exercise did not increase IL-6 mRNA expression in skeletal muscle-specific JNK-deficient mice. These data identify a novel contraction-mediated transcriptional regulatory pathway for IL-6 in skeletal muscle.

The Protective Effect of Exercise in Neurodegenerative Diseases: The Potential Role of Extracellular Vesicles.

Physical activity has systemic effects on the body, affecting almost every organ. It is important not only for general health and wellbeing, but also in the prevention of diseases. The mechanisms behind the therapeutic effects of physical activity are not completely understood; however, studies indicate these benefits are not confined to simply managing energy balance and body weight. They also include systemic factors which are released into the circulation during exercise and which appear to underlie the myriad of benefits exercise can elicit. It was shown that along with a number of classical cytokines, active tissues also engage in inter-tissue communication via extracellular vesicles (EVs), specifically exosomes and other small EVs, which are able to deliver biomolecules to cells and alter their metabolism. Thus, EVs may play a role in the acute and systemic adaptations that take place during and after physical activity, and may be therapeutically useful in the treatment of a range of diseases, including metabolic disorders such as type 2 diabetes and obesity; and the focus of this review, neurological disorders such as Alzheimer's disease.

No endogenous circadian rhythm in resting plasma Hsp72 concentration in humans.

Extra-cellular (e) heat shock protein (Hsp)72 has been shown to be elevated in a number of clinical conditions and has been proposed as a potential diagnostic marker. From a methodological and diagnostic perspective, it is important to investigate if concentrations of eHsp72 fluctuate throughout the day; hence, the purpose of the study was to measure resting concentrations of plasma eHsp72 throughout a 24-h period. Blood samples were taken every hour from 1200-2100 hours and from 0700-1200 hours the following day from seven healthy recreationally active males. Participants remained in the laboratory throughout the trial, performed light sedentary activities and were provided with standardised meals and fluids. Physical activity was quantified throughout by the use of an accelerometer. Ethylenediaminetetraacetic acid blood samples were analysed for eHsp72 concentration using a commercially available high-sensitivity enzyme-linked immunosorbent assay (intra-assay coefficient of variation = 1.4%). One-way repeated measures analysis of variance revealed that measures of physiological stress such as heart rate, systolic and diastolic blood pressure remained stable throughout the trial and subjects remained sedentary throughout (mean activity energy expenditure above resting metabolic rate-35.7 +/- 10.0 kcalh(-1)). Plasma Hsp72 concentration did not fluctuate significantly throughout the day and showed no apparent endogenous circadian rhythm in absolute (P = 0.367) or plasma volume change corrected data (P = 0.380). Individual coefficients of variation ranged from 3.8-7.7% (mean 5.4%). Mean Hsp72 concentration across all subjects and time points was 1.49 +/- 0.08 ngml(-1). These data show that in a rested state, plasma eHsp72 concentration shows no apparent endogenous circadian rhythm.

Heat shock protein 72: release and biological significance during exercise.

The cumulative stressors of exercise manifest themselves at a cellular level by threatening the protein homeostasis of the cell. In these conditions, Heat Shock Proteins (HSP) are synthesised to chaperone mis-folded and denatured proteins. As such, the intracellular HSP response is thought to aid cell survival in the face of otherwise lethal cellular stress. Recently, the inducible isoform of the 70 Kda heat shock protein family, Hsp72 has been detected in the extracellular environment. Furthermore, the release of this protein into the circulation has been shown to occur in response to a range of exercise bouts. The present review summarises the current research on the exercise Hsp72 response, the possible mediators and mechanisms of extracellular (e)Hsp72 release, and the possible biological significance of this systemic response. In particular, the possible role of eHsp72 in the modulation of immunity during exercise is discussed.

Human blood neutrophil responses to prolonged exercise with and without a thermal clamp.

The purpose of this study was to investigate the effects of prolonged exercise with and without a thermal clamp on neutrophil trafficking, bacterial-stimulated neutrophil degranulation, stress hormones, and cytokine responses. Thirteen healthy male volunteers (means +/- SE: age 21 +/- 1 yr; mass 74.9 +/- 2.1 kg; maximal oxygen uptake 58 +/- 1 ml x kg(-1) x min(-1)) completed four randomly assigned, 2-h water-immersion trials separated by 7 days. Trials were exercise-induced heating (EX-H: water temperature 36 degrees C), exercise with a thermal clamp (EX-C: 24 degrees C), passive heating (PA-H: 38.5 degrees C), and control (CON: 35 degrees C). EX-H and EX-C was comprised of 2 h of deep water running at 58% maximal oxygen uptake. Blood samples were collected at pre-, post-, and 1 h postimmersion. Core body temperature was unaltered on CON, clamped on EX-C (-0.02 degrees C), and rose by 2.23 degrees C and 2.31 degrees C on EX-H and PA-H, respectively. Exercising with a thermal clamp did not blunt the neutrophilia postexercise (EX-C postexercise: 9.6 +/- 1.1 and EX-H postexercise: 9.8 +/- 1.0 x 10(9)/liter). Neutrophil degranulation decreased (P < 0.01) similarly immediately after PA-H (-21%), EX-C, and EX-H (-28%). EX-C blunted the circulating norepinephrine, cortisol, granulocyte-colony stimulating factor, and IL-6 response (P < 0.01) but not the plasma epinephrine and serum growth hormone response. These results show a similar neutrophilia and decrease in neutrophil degranulation after prolonged exercise with and without a thermal clamp. As such, the rise in core body temperature does not appear to mediate neutrophil trafficking and degranulation responses to prolonged exercise. In addition, these results suggest a limited role for cortisol, granulocyte-colony stimulating factor, and IL-6 in the observed neutrophil responses to prolonged exercise.

Extracellular Vesicles Provide a Means for Tissue Crosstalk during Exercise.

Exercise stimulates the release of molecules into the circulation, supporting the concept that inter-tissue signaling proteins are important mediators of adaptations to exercise. Recognizing that many circulating proteins are packaged in extracellular vesicles (EVs), we employed quantitative proteomic techniques to characterize the exercise-induced secretion of EV-contained proteins. Following a 1-hr bout of cycling exercise in healthy humans, we observed an increase in the circulation of over 300 proteins, with a notable enrichment of several classes of proteins that compose exosomes and small vesicles. Pulse-chase and intravital imaging experiments suggested EVs liberated by exercise have a propensity to localize in the liver and can transfer their protein cargo. Moreover, by employing arteriovenous balance studies across the contracting human limb, we identified several novel candidate myokines, released into circulation independently of classical secretion. These data identify a new paradigm by which tissue crosstalk during exercise can exert systemic biological effects.

Effect of exercise with and without a thermal clamp on the plasma heat shock protein 72 response.

The contribution of heat and exercise related stress to the release of heat shock protein 72 (HSP72) is currently unknown. The purpose of the present study was to determine the combined and independent effects of heat and exercise on the extracellular (e)HSP72 response. Eleven moderately trained male volunteers [means +/- SD: age 21 +/- 4 yr; body mass 75.7 +/- 7.7 kg; maximal oxygen uptake ((.)Vo(2 max)) 57.8 +/- 3.3 ml.kg(-1).min(-1)] completed four 2-h, heat-manipulated, water-immersion trials. Trials were exercise-induced heat (EIH; rectal temperature change +2.2 degrees C), clamped exercise (CEx; 0 degrees C), passive heating (PHT; +2.3 degrees C), and control (Con; 0 degrees C). Exercise trials (EIH and CEx) comprised deep-water running at 58.5 +/- 2.4 and 59.1 +/- 1.7% (.)vo(2)max. eHSP72 and catecholamine concentrations were determined by ELISA and HPLC, respectively, pre- and postimmersion. All trials induced an eHSP72 response (P < 0.05) with postimmersion values significantly greater on EIH compared with other trials (6.0 +/- 3.4; CEx 3.8 +/- 2.6; PHT 2.7 +/- 2.1; Con 2.2 +/- 1.9 ng/ml). Exercising with a thermal clamp blunted the eHSP72 response, but postimmersion values were also greater than Con. PHT induced a large catecholamine response, but postimmersion eHSP72 values did not reach significance vs. Con. Given that exercising with a thermal clamp evoked a significant increase in plasma eHSP72 concentration, exercise-related stressors other than heat appeared influential in stimulating HSP72 release. Moreover, the catecholamine data from PHT suggest neither epinephrine nor norepinephrine was solely responsible for eHSP72 release.

Effect of caffeine supplementation on the extracellular heat shock protein 72 response to exercise.

The stimulus for the release of 72-kDa heat shock protein (HSP72) during exercise in humans is currently unclear. Recent evidence in an animal model is suggestive of an involvement of catecholamines. The present study, therefore, investigated the effect of caffeine supplementation, a known stimulator of sympathetic activity, on the extracellular (e)HSP72 response to prolonged exercise. Ten healthy male endurance-trained cyclists were recruited (age: 21 +/- 1 yr, maximum O(2) uptake 61.1 +/- 1.7 ml x kg(-1) x min(-1), mean +/- SE). Each subject was randomly assigned to ingest either 6 mg/kg body mass of caffeine (Caff) or placebo (Pla) 60 min before one of two 90-min bouts of cycling at 74 +/- 1% maximum O(2) uptake. Trials were performed at least 7 days apart in a counterbalanced design. Venous blood samples were collected by venepuncture at pretreatment, preexercise, postexercise, and 1 h postexercise. Serum caffeine and plasma catecholamines were determined using a spectrophotometric assay and high-performance liquid chromatography, respectively. Plasma HSP72 and cortisol were determined by ELISA. Serum caffeine concentrations were significantly increased throughout Caff, while no increases were detected in Pla. Caffeine supplementation and exercise was associated with a greater eHSP72 response than exercise alone (postexercise Caff 8.6 +/- 1.3 ng/ml; Pla 5.9 +/- 0.9 ng/ml). This greater eHSP72 response was associated with a greater epinephrine response to exercise in Caff. There was a significant increase in norepinephrine and cortisol, with no intertrial differences. The present data suggest that, in humans, catecholamines may be an important mediator of the exercise-induced increase in eHSP72 concentration.

Exercising in environmental extremes : a greater threat to immune function?

Athletes, military personnel, fire fighters, mountaineers and astronauts may be required to perform in environmental extremes (e.g. heat, cold, high altitude and microgravity). Exercising in hot versus thermoneutral conditions (where core temperature is > or = 1 degrees C higher in hot conditions) augments circulating stress hormones, catecholamines and cytokines with associated increases in circulating leukocytes. Studies that have clamped the rise in core temperature during exercise (by exercising in cool water) demonstrate a large contribution of the rise in core temperature in the leukocytosis and cytokinaemia of exercise. However, with the exception of lowered stimulated lymphocyte responses after exercise in the heat, and in exertional heat illness patients (core temperature > 40 degrees C), recent laboratory studies show a limited effect of exercise in the heat on neutrophil function, monocyte function, natural killer cell activity and mucosal immunity. Therefore, most of the available evidence does not support the contention that exercising in the heat poses a greater threat to immune function (vs thermoneutral conditions). From a critical standpoint, due to ethical committee restrictions, most laboratory studies have evoked modest core temperature responses (< 39 degrees C). Given that core temperature during exercise in the field often exceeds levels associated with fever and hyperthermia (approximately 39.5 degrees C) field studies may provide an opportunity to determine the effects of severe heat stress on immunity. Field studies may also provide insight into the possible involvement of immune modulation in the aetiology of exertional heat stroke (core temperature > 40.6 degrees C) and identify the effects of acclimatisation on neuroendocrine and immune responses to exercise-heat stress. Laboratory studies can provide useful information by, for example, applying the thermal clamp model to examine the involvement of the rise in core temperature in the functional immune modifications associated with prolonged exercise. Studies investigating the effects of cold, high altitude and microgravity on immunity and infection incidence are often hindered by extraneous stressors (e.g. isolation). Nevertheless, the available evidence does not support the popular belief that short- or long-term cold exposure, with or without exercise, suppresses immunity and increases infection incidence. In fact, controlled laboratory studies indicate immuno-stimulatory effects of cold exposure. Although some evidence shows that ascent to high altitude increases infection incidence, clear conclusions are difficult to make because of some overlap with the symptoms of acute mountain sickness. Studies have reported suppressed cell-mediated immunity in mountaineers at high altitude and in astronauts after re-entering the normal gravity environment; however, the impact of this finding on resistance to infection remains unclear.

The influence of an arduous military training program on immune function and upper respiratory tract infection incidence.

The effects of the first 19 weeks of U.K. Parachute Regiment (PARA) training on upper respiratory tract infection (URTI) incidence and immune function (circulating leukocyte counts, lymphocyte subsets, lipopolysaccharide-stimulated neutrophil degranulation, and salivary immunoglobulin A concentrations) were investigated for 14 PARA recruits and 12 control subjects. No significant differences were reported between groups for the number or duration of URTIs, lymphocyte subsets, or salivary immunoglobulin A concentrations during training. URTI incidence was greater in the PARA group at weeks 2 and 3 (p < 0.05), coinciding with a decrease in circulating leukocyte and lymphocyte counts (p < 0.05). Neutrophil degranulation was similar in the PARA and control groups at weeks 0 and 19. Decreases in saliva flow rate occurred in the PARA group at week 15 and weeks 18 to 20 (p < 0.05). These results show a limited effect of PARA training on URTI incidence and immune function. The progressive decrease in saliva flow rate during PARA training may indicate an ensuing state of hypohydration.