Immune Response in Young Thoroughbred Racehorses under Training

SIMPLE SUMMARY: Stressful stimuli, both infectious and non-infectious, can modify and trigger an innate immune response and inflammation, via an attempt to restore a homeostatic state. Coping with stressors can be measured by different procedures, including the evaluation of immunological parameters. These are also modulated by exercise, which can be considered stress prototypic in the Thoroughbred racehorse. To evaluate the complex of physiological regulations during the training period, twenty-nine clinically healthy, two-year-old Thoroughbred racehorses were followed during their first 3 months of sprint training. Blood collection was performed at rest, three times until 90 days of training, for testing immunological parameters during incremental sprint training to evaluate its effect on the immunological status of the animals. During the training period, we observed the following: (A) an increase in red blood cell parameters that are crucial for exercise performance adaptation, improving O(2) transport and muscle cell respiration; (B) variations of blood granulocytes; and (C) changes in inflammatory cytokine gene expression. On the basis of clinical and laboratory findings, training exercise probably played a major role in the modulation of the above parameters. These latter changes could be seen as a preparation of the innate immune system to respond quickly and adequately to environmental conditions. ABSTRACT: Training has a great impact on the physiology of an athlete and, like all stressful stimuli, can trigger an innate immune response and inflammation, which is part of a wider coping strategy of the host to restore homeostasis. The Thoroughbred racehorse is a valid animal model to investigate these changes thanks to its homogeneous training and highly selected genetic background. The aim of this study was to investigate modifications of the innate immune response and inflammation in young untrained Thoroughbred racehorses during the first training season through haematological and molecular investigations. Twenty-nine Thoroughbred racehorses were followed during their incremental 3-month sprint exercise schedule. Blood collection was performed at time 0 (T0; before starting the intense training period), 30 days after T0 (T30), and 90 days after T0 (T90). Haematological parameters (red and white blood cells, haemoglobin, and platelets) were evaluated and haematocrit (HCT), mean corpuscular haemoglobin concentration (MCHC), and red cells width distribution + standard deviation (RDW-SD) were calculated. Moreover, via RT-qPCR, we investigated the expression of, Interleukin 1β (IL-1β), Interleukin 4 (IL-4) Interleukin 6 (IL-6), Interleukin 2 (IL-2), Interleukin 3 (IL-3), Interleukin 5 (IL-5) Interleukin 8 (IL-8), Trasformig Growth Factor β and α (TGF-β), Tumor necrosis factor α (TNF-α), and Interferon γ (IFN-γ)genes. Main corpuscular volume (MCV) showed a significant (p = 0.008) increase at T90. Main corpuscular haemoglobin (MCH) and haemoglobin concentration (MCHC) values were significantly augmented at both T30 (p < 0.001) and T90 (p < 0.001). Basophils were significant increased at T30 (p = 0.02) and eosinophils were significantly increased at T90 (p = 0.03). Significant differences in gene expression were found for all the genes under study, with the exception of IFN-γ and TNF-α. In particular, IL-2 (T30, p = 0.011; T90, p = 0.015), IL-4 (T30, p = 0.009; T90, p < 0.001), and IL-8 (T30, p < 0.001; T90, p < 0.001) genes were significantly upregulated at both T30 and T90 with respect to T0, TGF-β was intensely downregulated at T30 (p < 0.001), IL-5 gene expression was significantly decreased at T90 (p = 0.001), while IL-1β (p = 0.005) and IL-3 (p = 0.001) expression was strongly augmented at the same time. This study highlighted long-term adjustments of O(2) transport capability that can be reasonably traced back to exercise adaptation. Moreover, the observed changes of granulocyte numbers and functions and inflammatory cytokine gene expression confirm a major role of the innate immune system in the response to the complex of stressful stimuli experienced during the training period.