The aim of this thesis was to investigate the interactions between stress responses, muscle damage, and neutrophil activation after exercise. Intense exercise is characterised by the release of stress hormones and cytokines that influence the activity of neutrophils in vitro. Downhill running causes muscle damage that may also lead to neutrophil priming and activation. By comparing moderate versus high intensity exercise, and using downhill running, the intention of the thesis was to contribute to a more thorough understanding of the factors influencing the activity of neutrophils during exercise. In addition, it was expected that studying the effects of carbohydrate supplementation during exercise would improve our knowledge of the effects of metabolic stress on the release of Cortisol, and the efflux of myocellular proteins from muscle during exercise. The thesis is divided into six chapters. The first and last chapters provide,
respectively, a general introduction and a summary of the research programme. The second chapter is a review of the literature in this field of research. The other three chapters are a series of separate scientific studies. Each forms an essential part of the thesis.
Chapter 2 is a review of the literature in this field. The first section provides an overview of the morphology and function of neutrophils, and the role of neutrophils in host defence. The second section summarises the results of studies of exercise and neutrophil function, and addresses some of the intracellular and systemic factors known to influence neutrophil function. The last section examines findings on the relationships between carbohydrate supplementation, muscle damage and Cortisol responses to exercise.
The first experimental study, described in Chapter 3, focuses on the influence of exercise intensity on changes in neutrophil surface receptor expression,
degranulation, and respiratory burst activity. The rationale for performing this study was that a variety of systemic factors known to regulate neutrophil number and function are affected by exercise intensity. To address these issues, ten well-trained male runners completed two 60-min running sessions on a treadmill at a moderate intensity (60% V02max), and a high intensity (85% VO2max). Exercise caused an intensity-dependent increase in the number of circulating neutrophils, myeloperoxidase release, and plasma levels of stress hormones and the cytokine interleukin-6 (P < 0.01). There was also an increase in the production of reactive oxygen species by neutrophils after high intensity exercise (P < 0.01). In contrast to these changes, exercise caused a decrease in the expression of receptors CDl lb, CD16, CD35 and CD88 (P < 0.01). The decline in CD16
expression was greater after high intensity exercise (P < 0.01). Consistent with previous literature, the increase in neutrophil count was associated with the systemic release of stress hormone and cytokines. The decline in receptor expression is in contrast with previous exercise studies, and may have represented an adaptive response to exercise-induced muscle damage. The increase in plasma creatine kinase activity and myoglobin concentration is indirect evidence that the exercise protocols used in this study caused a mild degree of muscle damage. There was evidence that the release of myeloperoxidase was associated with alterations in the metabolism of reactive oxygen species after exercise. These results are novel, and suggest that some measures of neutrophil function are influenced by exercise intensity.
The second experimental study, presented in Chapter 4, investigates the effects of downhill running on these same neutrophil
parameters. The justification for this study was that neutrophils play an important role in the clearance of damaged tissue. Previous exercise studies have generally only examined changes in individual aspects of neutrophil function after downhill running. To address these issues, ten well-trained male runners ran downhill for 45 min on a treadmill at a gradient of -10%. Downhill running caused a large increase in plasma myoglobin concentration (1800% increase; P < 0.01) and creatine kinase activity (420% increase; P < 0.01). There was also an increase in plasma interleukin-6 concentration (460% increase; P < 0.01), but no change in plasma IL-8 levels (P > 0.05). The efflux of myoglobin and creatine kinase correlated with the mobilisation of neutrophils after exercise. However, contrary to previous studies, there were no significant changes in any aspect of neutrophil activation,
despite the large increases in plasma levels of myoglobin and creatine kinase. This disparity could have been due to differences in exercise protocols and the training status of the subjects involved.
The last experimental study, outlined in Chapter 5, explores die effects of carbohydrate supplementation on changes in plasma Cortisol and myoglobin levels. The basis for this study was that carbohydrate reduces the Cortisol response to prolonged submaximal exercise, but its effects during high intensity exercise are unknown. Furthermore, although carbohydrate ingestion attenuates increases in the plasma concentration of several anti-inflammatory cytokines, it is not known if carbohydrate ingestion influences changes in markers of muscle damage. To address these questions, eight well-trained runners completed two 60-min running sessions on a treadmill during which they consumed either a 10% carbohydrate drink or a placebo drink. Both trials lead to a marked
increase in plasma Cortisol levels. However, despite an increase in plasma glucose concentration in response to carbohydrate ingestion (P < 0.05), the pattern of change was not significantly different between the trials. Plasma myoglobin concentration increased significantly after exercise (P < 0.01), providing indirect evidence that exercise had induced muscle damage. The pattern of change between the trials was significantly different (P < 0.01), with carbohydrate consumption resulting in lower plasma myoglobin levels. Plasma ascorbic acid concentration increased significantly (P < 0.01). However, there was no change on plasma total antioxidant capacity. This data suggests that during high intensity exercise, Cortisol is released independently of changes in plasma glucose levels. In addition, carbohydrate could help reduce the leakage of myocellular proteins from damaged muscle
during prolonged exercise with an eccentric component (e.g., downhill running).
Chapter 6 summarises the main findings of the thesis, addresses the possible relationships between exercise stress and immune changes, and discusses the results of the thesis in the context of previous research.
In summary, the data presented in this thesis confirm previous observations that intense exercise is accompanied by the release of stress hormones and cytokines, which contribute to the mobilisation of neutrophils. The thesis's novel data indicate that, compared with moderate intensity exercise, high intensity exercise may also be associated with a greater decrease in the expression of some surface receptors on neutrophils, neutrophil degranulation and an increase in respiratory burst activity. The decrease in the expression of neutrophil surface receptors may represent an adaptive response to skeletal muscle injury. Alternatively, if these
receptors are not replaced in the hours after exercise, this could render athletes more susceptible to infection. The increase in plasma myoglobin concentration and creatine kinase activity indicated that downhill running caused injury to skeletal muscle. However, this apparent muscle damage did not lead to activation of neutrophils. Carbohydrate supplementation appeared to attenuate the release of neutrophils and myoglobin from muscle, but may not affect the Cortisol response to high intensity exercise in some individuals. From all of the present work it may be concluded that exercise causes a non-specific increase in the circulating neutrophil count that is mediated by the intensity-dependent systemic release of stress hormones and cytokines. Whereas muscle damage may act as a specific stimulus for neutrophil release, it may not always lead to changes in neutrophil priming and activation. These alterations may be more closely related to the intensity of exercise.