Assessment of exercise-induced alterations in neutrophil function in horses

Douglas C. Donovan Department of Population Health and Large Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, GA 30602

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Christie A. Jackson Department of Large Animal Clinical Sciences, College of Veterinary Medicine, Gainesville, FL 32610

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Patrick T. Colahan Department of Large Animal Clinical Sciences, College of Veterinary Medicine, Gainesville, FL 32610

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Natalie N. Norton Department of Large Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, GA 30602

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Joan L. Clapper Department of Population Health, College of Veterinary Medicine, University of Georgia, Athens, GA 30602

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James N. Moore Department of Large Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, GA 30602

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David J. Hurley Department of Population Health and Large Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, GA 30602

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Abstract

Objective—To evaluate the effects of a standardized exercise test to exhaustion in horses on leukocyte function ex vivo.

Animals—6 Thoroughbred geldings.

Procedures—Blood samples were obtained from each horse before exercise; at exhaustion (termed failure); and at 2, 6, 24, 48, and 72 hours after exercise to evaluate hematologic changes, rate of leukocyte apoptosis, and leukocyte production of reactive oxygen species (ROS) ex vivo. To assess leukocyte function, leukocyte ROS production in response to stimulation with lipopolysaccharide, peptidoglycan, zymosan, and phorbol myristate acetate was evaluated. Apoptosis was evaluated via assessment of caspase activity in leukocyte lysates.

Results—In response to lipopolysaccharide, production of ROS by leukocytes was significantly increased at 2 hours and remained increased (albeit not significantly) at 6 hours after exercise, compared with the preexercise value. In the absence of any stimulus, leukocyte ROS production was significantly increased at 6 and 24 hours after exercise. In contrast, ROS production in response to phorbol myristate acetate was significantly decreased at 6, 24, and 72 hours after exercise. Leukocyte ROS production induced by zymosan or peptidoglycan was not altered by exercise. Leukocytosis was evident for 24 hours after exercise, and neutrophilia was detected during the first 6 hours. A significant increase in the rate of leukocyte apoptosis was detected at failure and 72 hours after exercise.

Conclusions and Clinical Relevance—Results indicated that strenuous exercise undertaken by horses causes alterations in innate immune system functions, some of which persist for as long as 72 hours after exercise.

Abstract

Objective—To evaluate the effects of a standardized exercise test to exhaustion in horses on leukocyte function ex vivo.

Animals—6 Thoroughbred geldings.

Procedures—Blood samples were obtained from each horse before exercise; at exhaustion (termed failure); and at 2, 6, 24, 48, and 72 hours after exercise to evaluate hematologic changes, rate of leukocyte apoptosis, and leukocyte production of reactive oxygen species (ROS) ex vivo. To assess leukocyte function, leukocyte ROS production in response to stimulation with lipopolysaccharide, peptidoglycan, zymosan, and phorbol myristate acetate was evaluated. Apoptosis was evaluated via assessment of caspase activity in leukocyte lysates.

Results—In response to lipopolysaccharide, production of ROS by leukocytes was significantly increased at 2 hours and remained increased (albeit not significantly) at 6 hours after exercise, compared with the preexercise value. In the absence of any stimulus, leukocyte ROS production was significantly increased at 6 and 24 hours after exercise. In contrast, ROS production in response to phorbol myristate acetate was significantly decreased at 6, 24, and 72 hours after exercise. Leukocyte ROS production induced by zymosan or peptidoglycan was not altered by exercise. Leukocytosis was evident for 24 hours after exercise, and neutrophilia was detected during the first 6 hours. A significant increase in the rate of leukocyte apoptosis was detected at failure and 72 hours after exercise.

Conclusions and Clinical Relevance—Results indicated that strenuous exercise undertaken by horses causes alterations in innate immune system functions, some of which persist for as long as 72 hours after exercise.

Contributor Notes

Supported by the White Fox Farm Research Fund.

Address correspondence to Dr. Hurley.
  • 1.

    Parks CM, Manohar M. Blood-gas tensions and acid-base status in ponies during treadmill exercise. Am J Vet Res 1984;45:1519.

  • 2.

    Nieman DC. Current perspectives on exercise immunology. Curr Sports Med Rep 2003;2:539542.

  • 3.

    Wong CW, Smith SE, Thong YH, et al. Effects of exercise stress on various immune functions in horses. Am J Vet Res 1992;53:14141417.

  • 4.

    Buschmann H, Baumann M. Alterations of cellular immune response during intensive training of event horses. Zentralbl Veterinarmed [B] 1991;38:9094.

    • Search Google Scholar
    • Export Citation
  • 5.

    Raidal SL, Love DN, Bailey GD, et al. Effect of single bouts of moderate and high intensity exercise and training on equine peripheral blood neutrophil function. Res Vet Sci 2000;68:141146.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 6.

    Folsom RW, Littlefield-Chabaud MA, French DD, et al. Exercise alters the immune response to equine influenza virus and increases susceptibility to infection. Equine Vet J 2001;33:664669.

    • Search Google Scholar
    • Export Citation
  • 7.

    Horohov DW, Dimock A, Guirnalda P, et al. Effect of exercise on the immune response of young and old horses. Am J Vet Res 1999;60:643647.

    • Search Google Scholar
    • Export Citation
  • 8.

    Peters EM, Van Eden M, Tyler N, et al. Prolonged exercise does not cause lymphocyte DNA damage or increased apoptosis in well-trained endurance athletes. Eur J Appl Physiol 2006;98:124131.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 9.

    Raidal SL, Love DN, Bailey GD, et al. The effect of high intensity exercise on the functional capacity of equine pulmonary alveolar macrophages and BAL-derived lymphocytes. Res Vet Sci 2000;68:249253.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 10.

    Raidal SL, Love DN, Bailey GD, et al. Effect of a single bout of high intensity exercise on lower respiratory tract contamination in the horse. Aust Vet J 1997;75:293295.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 11.

    Kothakota S, Azuma T, Reinhard C, et al. Caspase-3-generated fragment of gelosin: effector of morphological change in apoptosis. Science 1997;278:294298.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 12.

    Hurley DJ, Parks RJ, Reber AJ, et al. Dynamic changes in circulating leukocytes during the induction of equine laminitis with black walnut extract. Vet Immunol Immunopathol 2006;110:195206.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 13.

    Skoog DA, West DM. Rejection of data. In: Fundamentals of analytical chemistry. 2nd ed. New York, NY: Holt, Rinehart, and Winston Inc, 1969;45.

    • Search Google Scholar
    • Export Citation
  • 14.

    Goihman-Yahr M, Villalba-Pimentel L, Rodriguez-Ochoa G, et al. Studies on the effect of serum and proteins on in vitro-induced neutrophil activation. J Reticuloendothel Soc 1978;23:435446.

    • Search Google Scholar
    • Export Citation
  • 15.

    Okano S, Hurley DJ, Bergh MS, et al. Optimization of condition for measurement of tissue factor expression and radical oxygen species induction by canine mononuclear cell and granulocytes in vitro. Vet Immunol Immunopathol 2006;112:234242.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 16.

    Baker B, Gaffin SL, Wells M, et al. Endotoxaemia in racehorses following exertion. J S Afr Vet Assoc 1988;59:6366.

  • 17.

    Barton MH, Williamson L, Jacks S, et al. Effects on plasma endotoxin and eicosanoid concentrations and serum cytokine activities in horses competing in a 48-, 83-, or 159-km endurance ride under similar terrain and weather conditions. Am J Vet Res 2003;64:754761.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 18.

    Donovan DC, Jackson CA, Colahan PT, et al. Exercise-induced alterations in pro-inflammatory cytokines and prostaglandin F2 A. Vet Immunol Immunopathol 2007;118:263269.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 19.

    Weiss DJ, Evanson OA. Evaluation of lipopolysacharide-induced activation of equine neutrophils. Am J Vet Res 2002;63:811815.

  • 20.

    Shimazu R, Akashi S, Ogata H, et al. MD-2, a molecule that confers lipopolysaccharide responsiveness on Toll-like receptor 4. J Exp Med 1999;189:17771782.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 21.

    Akira S. TLR Signaling. Curr Top Microbiol Immunol 2006;311:116.

  • 22.

    Takeuchi O, Hoshino K, Kawai T, et al. Differential roles of TLR2 and TLR4 in recognition of gram-negative and gram-positive bacterial cell wall components. Immunity 1999;11:443451.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 23.

    Windheim M, Lang C, Peggie M, et al. Molecular mechanisms involved in the regulation of cytokine production by muramyl peptide. Biochem J 2007;404:179190.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 24.

    Brown GD, Taylor PR, Reid DM, et al. Dectin-1 is a major betaglucan receptor on macrophages. J Exp Med 2002;196:407412.

  • 25.

    Willment JA, Lin HH, Reid DM, et al. Dectin-1 expression and function are enhanced on alternatively activated and GM-CSF-treated macrophages and are negatively regulated by IL-10, dexamethasone, and lipopolysaccharide. J Immunol 2003;171:45694573.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 26.

    Hancock JT, Desikan R, Neill SJ. Role of reactive oxygen species in cell signaling pathways. Biochem Soc Trans 2001;29:345350.

  • 27.

    Gujral JS, Hinson JA, Farhood A, et al. NADPH oxidase-derived oxidative stress is critical for neutrophil cytotoxicity during endotoxemia. Am J Physiol Gastrointest Liver Physiol 2004;287:G243G252.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 28.

    Navalta JW, Sedlock DA, Park K-S. Effect of exercise intensity on exercise-induced lymphocyte apoptosis. Int J Sports Med 2007;28:539542.

  • 29.

    Rath PC, Aggarwai BB. TNF-induced signaling in apoptosis. J Clin Immunol 1999;19:350364.

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