Acute exercise mobilises CD8+ T lymphocytes exhibiting an effector-memory phenotype

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Abstract

An acute bout of exercise evokes mobilisation of lymphocytes into the bloodstream, which can be largely attributed to increases in CD8+ T lymphocytes (CD8TLs) and natural killer (NK) cells. Evidence further suggests that, even within these lymphocyte subsets, there is preferential mobilisation of cells that share certain functional and phenotypic characteristics, such as high cytotoxicity, low proliferative ability, and high tissue-migrating potential. These features are characteristic of effector-memory CD8TL subsets. The current study therefore investigated the effect of exercise on these newly-identified subsets.

Thirteen healthy and physically active males (mean ± SD: age 20.9 ± 1.5 yr) attended three sessions: a control session (no exercise); cycling at 35% Wattmax (low intensity exercise); and 85% Wattmax (high intensity exercise). Each bout lasted 20 min. Blood samples were obtained before exercise, during the final min of exercise, and +15, and +60 min post-exercise. CD8TLs were classified into naïve, central memory (CM), effector-memory (EM), and CD45RA+ effector-memory (RAEM) using combinations of the cell surface markers CCR7, CD27, CD62L, CD57, and CD45RA. In parallel, the phenotypically distinct CD56bright ‘regulatory’ and CD56dim ‘cytotoxic’ NK subsets were quantified.

The results show a strong differential mobilisation of CD8TL subsets (RAEM > EM > CM > naïve); during high intensity exercise the greatest increase was observed for RAEM CD8Tls (+450%) and the smallest for naïve cells (+84%). Similarly, CD56dim NK cells (+995%) were mobilised to a greater extent than CD56bright (+153%) NK cells. In conclusion, memory CD8TL that exhibit a high effector and tissue-migrating potential are preferentially mobilised during exercise. This finding unifies a range of independent observations regarding exercise-induced phenotypic and functional changes in circulating lymphocytes. The selective mobilisation of cytotoxic tissue-migrating subsets, both within the NK and CD8TL population, may enhance immune-surveillance during exercise.

Introduction

The rapid increase in peripheral blood lymphocytes during exercise is one of the most replicated findings in exercise immunology (Gleeson, 2007, Kruger and Mooren, 2007). This transient lymphocytosis is largely comprised of CD8+ T lymphocytes (CD8TLs) and natural killer (NK) cells and is thought to be driven by two physiological processes. First, increases in cardiac output and concomitant haemodynamic shear forces flush the marginal pools (Shephard, 2003). In addition to these non-specific forces, a preferential mobilisation of cytotoxic lymphocytes is likely to result from stimulation of the β2-adrenergic receptors that are highly expressed on these cells (Landmann, 1992, Maisel et al., 1990, Murray et al., 1992). Adrenergic stimulation mediates endothelial detachment and subsequent recirculation (Benschop et al., 1994). Indeed, the exercise-induced mobilisation of CD8TLs and NK cells is replicated during psychological stress (Atanackovic et al., 2006, Bosch et al., 2005) and administration of β2-adrenergic agonists (Mills et al., 2000, van Tits et al., 1990). Moreover, exercise lymphocytosis is partially inhibited by administering β-adrenergic antagonists (Mills et al., 1999, Murray et al., 1992).

Later studies have also shown that the exercise-induced mobilisation of CD8TL and NK cells is accompanied by specific phenotypic changes. For example, mobilised CD8TLs appear to have lower levels of the adhesion molecule CD62L and higher levels of CD11a (for further examples, see Table 1, left panel). Although these changes have typically been discussed as if they represent independent observations, it is conceivable that these findings reflect a single underlying response. This hypothesis would be supported if the mobilised CD8TLs are a distinct subset that shares all of the characteristics outlined in Table 1. Such a CD8TL subset may indeed exist. CD8TLs have traditionally been divided into naïve and memory subsets, according to the expression or loss of CD45RA, respectively (Mackay, 1999). In recent years, however, it has become clear that at least four broad subsets of CD8TLs can be discerned: naïve; central memory (CM); effector-memory (EM); and, finally, effector-memory CD8TLs that have re-expressed the ‘naïve’ marker CD45RA (RAEM) (Campbell et al., 2001, Champagne et al., 2001, Hamann et al., 1997, Sallusto et al., 1999). These subsets reflect different maturation stages ranging from naïve, which have not been stimulated by antigen, to RAEM, which have an extensive history of antigen exposure. Naïve and CMs typically recirculate between the blood and secondary lymphoid tissues, where they, upon antigen exposure, develop into cytotoxic effector cells. In contrast, EMs and RAEMs are already cytotoxic and preferentially recirculate between the blood and the peripheral tissues (Hamann et al., 1997, Joshi and Kaech, 2008, Lanzavecchia and Sallusto, 2005). These two effector-memory subsets, in particular the RAEM subset, are characterised by certain phenotypic (i.e., cell marker) and functional characteristics that closely mirror those of exercise-mobilised CD8TLs (see Table 1 for a comparison). Hence, the selective mobilisation of effector-memory CD8TLs could provide a coherent explanation for the array of changes during acute exercise.

In order to analyse these various CD8TL subsets, research groups have used different identification methods which are all based on the observation that effector-memory T cells down-regulate lymphoid homing markers (e.g., CD62L, CCR7) and co-stimulatory receptor ligands (e.g., CD27) (Campbell et al., 2001, Faint et al., 2001, Hamann et al., 1997, Sallusto et al., 1999). These markers can be used in combination with CD45RA to divide human T cells into the aforementioned distinct subsets (Sallusto et al., 1999). Each of these marker combinations (e.g., CCR7, CD62L, CD27) identify largely overlapping, although not fully identical, CD8TL subsets. Therefore, the current study compared these distinct identification methods.

The predicted preferential mobilisation of effector-memory CD8TLs would be consistent with previous observations in NK cells. NK cells too can be divided into functionally distinct subsets that exhibit specific migratory patterns and different cytotoxic potential (Bosch et al., 2005, Cooper et al., 2001a). The majority of circulating NK cells (∼90%) express relatively low levels of CD56 (CD56dim) and function as cytotoxic cells; functionally and phenotypically, these cells show a resemblance to the effector-memory CD8TL subsets (Cooper et al., 2001a, Cooper et al., 2001b). It has been found that these cytotoxic CD56dim NK cells are preferentially mobilised during exercise (Timmons and Cieslak, 2008) and acute psychological stress (Bosch et al., 2005). In contrast, CD56bright NK cells, which have a predominantly immunomodulatory role and recirculate between the blood and the secondary lymphoid tissues (Cooper et al., 2001b), are relatively unaffected by exercise. Hence, any preferential mobilisation of effector-memory CD8TLs may reflect a more general principle; exercise may recruit cytotoxic lymphocytes with a tissue-migrating and immediate effector capability, while leaving slower responding cells bound for the lymphoid tissues, largely unaffected.

This study investigated the effects of different intensities of brief exercise on the mobilisation of CD8TL and NK subsets. As indicated, a selective mobilisation of effector-memory CD8TLs and CD56dim NK cells would provide a unifying explanation for a range of previous observations. As there are several classification methods to identify CD8TL subsets, a secondary aim of the current study was to compare the results obtained using these distinct approaches.

Section snippets

Participants

Thirteen healthy male students (mean ± SD: age 20.9 ± 1.5 yr; body mass index 22.9 ± 2.4 kg/m2) participated in this study. Participants were screened prior to inclusion and were ineligible if sedentary, medicated or reporting health problems indicative of cardiovascular, inflammatory, or infectious disease. All participants provided written informed consent and the study was approved by the School of Sport and Exercise Sciences Ethics Committee, University of Birmingham.

Experimental design

Participants visited the

Results

All participants completed each of the three conditions. Table 2 provides the summary data and results for oxygen consumption (ml kg−1 min−1), heart rate (beats/min), workload (W), and RPE (Borg Scale) during these sessions. As expected, mean oxygen consumption and heart rate increased in both exercise conditions (F > 196.1; p < .001), with greater increases seen during higher intensity exercise (F > 96.0; p < .001). These results confirmed that the three conditions were clearly distinct in terms of

Discussion

This study analysed the effect of brief exercise on the mobilisation of peripheral blood lymphocyte subsets. There was a robust increase in lymphocyte number during low intensity exercise, and an even larger elevation during high intensity exercise. As expected, this mobilisation response was largely comprised of CD8TLs and NK cells. The primary focus of this study was to determine whether specific subsets of CD8TLs and NK cells account for this increase, applying recent advances in cell

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