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J Appl Physiol 98: 565-571, 2005. First published August 20, 2004; doi:10.1152/japplphysiol.00754.2004
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Effect of prolonged exercise and carbohydrate ingestion on type 1 and type 2 T lymphocyte distribution and intracellular cytokine production in humans

G. I. Lancaster,1 Q. Khan,2 P. T. Drysdale,2 F. Wallace,3 A. E. Jeukendrup,1 M. T. Drayson,2 and M. Gleeson4

1School of Sport and Exercise Sciences, 2Department of Immunology, The Medical School, University of Birmingham, Birmingham; 3GlaxoSmithKline Consumer Healthcare, Brentford; and 4School of Sport and Exercise Sciences, Loughborough University, Loughborough, United Kingdom

Submitted 19 July 2004 ; accepted in final form 18 August 2004


   ABSTRACT
TOP
 ABSTRACT
METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGMENTS
 REFERENCES
 
The present study was undertaken to examine the role of the exercise-induced stress hormone response on the regulation of type 1 and type 2 T lymphocyte intracellular cytokine production. Subjects performed 2.5 h of cycling exercise at 65% maximal O2 uptake while ingesting a 6.4% carbohydrate (CHO) solution, 12.8% CHO solution, or a placebo. Peripheral whole blood samples were stimulated and stained for T lymphocyte surface antigens (CD4 and CD8). Cells were then permeabilized, stained for intracellular cytokines, and analyzed using flow cytometry. Exercise resulted in a decrease (P < 0.05) in the number and percentage of IFN-{gamma} positive CD4+ and CD8+ T lymphocytes. These stimulated cells produced less IFN-{gamma} immediately postexercise (P < 0.05) and 2-h postexercise (P < 0.05) compared with preexercise. However, CHO ingestion, which attenuated the exercise-induced stress hormone response compared with placebo (P < 0.05), prevented both the decrease in the number and percentage of IFN-{gamma}-positive CD4+ and CD8+ T lymphocytes and the suppression of IFN-{gamma} production from stimulated CD4+ and CD8+ T lymphocytes. There was no effect of exercise on the number of, or cytokine production from, IL-4-positive CD4+ or CD8+ T lymphocytes. These data provide support for the role of exercise-induced elevations in stress hormones in the regulation of type 1 T lymphocyte cytokine production and distribution.

T lymphocytes; cytokines; cycling; immune

PRODUCTION OF T cell cytokines plays a critical role in the development of host immunity against invading pathogens. It is generally established that intracellular pathogens initiate a strongly cellular (type 1) immune response resulting in the differentiation of bipotential naive CD4+ and CD8+ T cells into T helper and T cytotoxic type 1 (TH1/TC1) phenotypic cells characterized by the production of interferon (IFN)-{gamma} and interleukin (IL)-2 (16). Extracellular pathogens, on the other hand, initiate a humoral (type 2) immune response resulting in the differentiation of naive CD4+ and CD8+ T cells into T helper and T cytotoxic type 2 (TH2/TC2) phenotypic cells characterized by the production of IL-4, IL-5, IL-10, and IL-13 (16).

Although a variety of genetic and immunologic factors is important in the regulation of TH1/TC1 and TH2/TC2 lymphocytes, the hypothalamic-pituitary-adrenal axis and the sympathetic nervous system play an important role in the modulation of TH1/TC1 and TH2/TC2 lymphocytes (4). Exercise provides a multifactorial stimulus to the central nervous system, acting via feedforward (motor center stimulation of endocrine centers within the brain) and feedback (blood-borne metabolic and peripheral neural) mechanisms, resulting in an increase in the systemic concentration of several immunomodulatory hormones (10). Indeed, it has been demonstrated that prolonged exercise causes a redistribution of circulating TH1/TC1 and TH2/TC2 cells resulting in a shift toward a type 2 dominance (8, 19), and this decline in type 1 cells has been suggested as a mechanism for the increased susceptibility to upper respiratory tract infection after prolonged exercise (19). In a recent study examining the role of catecholamines on T lymphocyte cytokine production (18), {alpha}- and {beta}-adrenergic blockade attenuated the postexercise increase in the circulating concentration of IFN-{gamma} producing CD3+ T cells (i.e., TH1 cells) compared with placebo, therefore providing support for the role of catecholamines in the exercise-induced modulation of the circulating type 1 T cell concentration. IFN-{gamma} production is critical to antiviral defense, and several studies (1, 17, 22) have demonstrated a decrease in the concentration of IFN-{gamma} in the supernatant of mitogen-stimulated whole blood after strenuous exercise. Indeed, it has been hypothesized that the suppression of stimulated IFN-{gamma} production may be an important mechanism leading to an increased risk of infection after prolonged exercise (13). However, this suppression is unlikely to be mediated via adrenergic receptor stimulation because {alpha}- and {beta}-adrenergic blockade did not affect the exercise-induced amount of IFN-{gamma} produced by stimulated CD3+ T lymphocytes.

Therefore, the present study further examined the regulation of TH1/TC1 and TH2/TC2 lymphocyte distribution and function after prolonged exercise. We hypothesized that prolonged exercise would cause a stress hormone-mediated redistribution of the circulating TH1/TC1 lymphocytes and would suppress TH1/TC1 and TH2/TC2 lymphocyte IFN-{gamma} and IL-4 production, respectively. However, carbohydrate (CHO) ingestion during exercise, by blunting the exercise-induced stress hormone response, would attenuate the redistribution of circulating TH1/TC1 and TH2/TC2 lymphocytes and ameliorate the suppressive effects of exercise on TH1/TC1 and TH2/TC2 lymphocyte intracellular cytokine production.


   METHODS
TOP
 ABSTRACT
 METHODS
RESULTS
 DISCUSSION
 ACKNOWLEDGMENTS
 REFERENCES
 
Subjects

Seven healthy, moderately to well endurance-trained men (age 25 ± 1 yr, body mass 77 ± 1 kg, maximal oxygen uptake 4.56 ± 0.06 l/min; means ± SE) volunteered to participate in the study. Subjects were nonsmokers who were not taking any medication and had remained free of respiratory infection for 4 wk before participation in the study. Subjects were fully informed as to the purposes and risks of the experiment before voluntarily giving their written, informed consent. The study was approved by the University of Birmingham Ethics Committee.

Preliminary Procedures

At least 7 days before the first experimental trial, each subject's maximal work rate (Wmax) was determined during an incremental exercise test to volitional exhaustion as described previously (9). At least 5 days before the first experimental trial, subjects performed a familiarization trial at 55% Wmax for 2.5 h. On the basis of their performance during the familiarization trial it was necessary to reduce the work rate employed for three of the subjects to ensure that each subject was able to complete the 2.5-h experimental trials without reaching exhaustion. On the day preceding the first experimental trial, subjects were instructed to consume a diet high in CHO and to record all food and fluid consumed. On the day preceding subsequent experimental trials, subjects were instructed to adhere as closely as possible to the recorded diet consumed before the first experimental trial. In addition, subjects were instructed to abstain from alcohol, tobacco, and exercise the day before the experimental trials.

Experimental Procedures

Subjects reported to the Human Performance Laboratory at the University of Birmingham in the morning after an overnight fast (~10 h). To avoid circadian variations in circulating hormones, individual subjects commenced all experimental trials at the same time, and all trials commenced between 7:30 and 8:30 AM. An indwelling 21-gauge Teflon catheter (Baxter, Norfolk, UK) was inserted into an antecubital vein of one arm, connected to a three-way stopcock (Sims Portex, Kent, UK), and a resting blood sample (preexercise) was drawn. Subjects then consumed 500 ml of a 6.4% CHO, 12.8% CHO, or placebo (Pla) solution. After a 10 min warm-up consisting of exercise at ~30% Wmax, subjects commenced the 2.5 h of cycling exercise at ~54% Wmax (~65% maximal oxygen uptake). Further blood samples were obtained on completion of the exercise (postexercise) and after 2 h of resting recovery (2 h postexercise). The catheter was kept patent by flushing with ~1.0 ml of sterile saline solution (0.9% NaCl) after sampling. Throughout the trial subjects consumed a further 200 ml of the experimental solution (Pla, 6.4% CHO, or 12.8% CHO) every 20 min, resulting in a total volume consumed of 2 liters. All trials were performed in comfortable ambient conditions (22–24°C, ~50% relative humidity), and trial orders were randomized.

Samples were analyzed for plasma cortisol, growth hormone (GH), adrenocorticotropic hormone (ACTH), IL-6, glucose, total and differential leukocyte counts, lymphocyte surface marker expression, and phorbol 12-myristate 13-acetate- and ionomycin-stimulated T lymphocyte intracellular cytokine production.

During each experimental trial, whole blood (10 ml) was divided into three sterile K3EDTA vacutainer tubes (Becton Dickinson, Oxford, UK). Two tubes were kept at room temperature until the end of the trial and analyzed for total and differential leukocyte counts and lymphocyte surface marker expression. The remaining whole blood was separated immediately by centrifugation (1,500 g) for 10 min, and aliquots of plasma were stored at –20°C until analysis for cortisol, ACTH, GH, IL-6, and glucose. A further 7 ml of whole blood were placed in sterile lithium-heparin vacutainer tubes (Becton Dickinson) and kept at room temperature until the end of the trial for analysis of intracellular cytokine production.

Leukocyte Counts

Samples were analyzed for total leukocyte, neutrophil, lymphocyte, and monocyte counts. This procedure was performed at the hematology laboratory, University Hospital Birmingham using a Technicon H-2 laser system.

Assessment of Intracellular Cytokine Production

Cell activation.   Whole blood (500 µl) was added to 500 µl RPMI-1640 medium supplemented with 2 mM L-glutamine in each of two 12 x 75-mm polystyrene tubes (Becton Dickinson Labware). To one set of sample tubes (stimulated), 10 µg/ml brefeldin A, 25 ng/ml of the phorbol ester phorbol 12-myristate 13-acetate, and 1 µg/ml of the calcium ionophore ionomycin were added. To the second set of sample tubes (unstimulated), only 10 µg/ml of brefeldin A were added. Brefeldin A is a potent inhibitor of intracellular protein transport and was added to samples to ensure that cytokines produced during stimulation would be retained within the cell. Unstimulated and stimulated samples were then incubated at 37°C for 4 h in a 5% CO2 humidified atmosphere.

Staining for intracellular cytokines.   Aliquots (100 µl) of unstimulated and stimulated whole blood were added to separate polystyrene tubes containing 4 µl of fluorescein isothiocyanate (FITC)-CD4 and peridinin chlorophyll protein (PerCP)-CD8-conjugated monoclonal antibodies (BD Biosciences, San Jose, CA) and incubated for 15 min. Erythrocytes were lysed by adding 2 ml lysing solution (BD Biosciences) and incubated for 10 min. Samples were centrifuged (1,000 rpm) for 5 min, and the supernatant was decanted. Cells were permeabilized by adding 500 µl permeabilizing solution (BD Biosciences) and incubated for a further 10 min. Samples were then washed (0.5% bovine serum albumin, phosphate-buffered saline, and 0.1% NaN3) and centrifuged (1,000 rpm) for 5 min, and the supernatant was decanted. Samples were then incubated with R-phycoerythrin (PE)-IFN-{gamma}, and PE-IL-4, or control mouse PE-IgG2b and PE-IgG1 conjugated monoclonal antibodies for 30 min. This resulted in the following combinations of antibodies: CD4+/IFN-{gamma}+/IL-4+ and CD8+/IFN-{gamma}+/IL-4+. Samples were then washed and centrifuged (1,000 rpm) for 5 min, and the supernatant was decanted before being resuspended in 200 µl of PBS (1% paraformaldehyde) and analyzed by flow cytometry. All incubations took place at room temperature in the dark.

Assessment of Lymphocyte Surface Marker Expression

Surface marker staining on unstimulated lymphocytes. Aliquots (50 µl) of whole blood were added to polystyrene tubes and incubated for a minimum of 15 min with the following combinations of conjugated monoclonal antibodies (all antibodies were obtained from BD Immunocytometry, San Jose, CA, unless otherwise stated): PerCP-CD3, FITC-CD4 and PE-CD8; PE-CD4 (DAKO), PerCP-CD8, and FITC-CD45RO (DAKO); CD4-PE and CD8-PerCP. Erythrocytes were lysed (1 ml lysing solution), and samples were washed (3 ml PBS, 0.1% NaN3) essentially as described above and then analyzed by flow cytometry. All incubations took place at room temperature in the dark.

Flow cytometric analysis.   Labeled cells were analyzed by flow cytometry using a fluorescence-activated cell sorter (Becton Dickinson FACSCalibur). Cells were selected on the basis of their forward vs. side light scatter and were subsequently analyzed (Cell Quest, Becton Dickinson) with gates set to identify positive and negative staining set on isotype controls (<1% positive). Lymphocyte subset counts were expressed as percentages of cells among total lymphocytes. Absolute numbers of lymphocyte subsets were determined by multiplying the corresponding percentages by the total lymphocyte count. Intracellular cytokine production results were expressed as the percentage of CD4+/IFN-{gamma}+/IL-4+ and CD8+/IFN-{gamma}+/IL-4+ cells among total CD4+ and CD8+ T cells. Absolute numbers of cytokine-positive CD4+ and CD8+ T cells were determined by multiplying the corresponding percentages by the total CD4+ and CD8+ T cell concentration. To quantify the amount of cytokine within positive cells the geometric mean fluorescence intensity was obtained.

Circulating Stress Hormones, Metabolites, and IL-6

Plasma cortisol (DRG Instruments), growth hormone (DRG Instruments), adrenocorticotropic hormone (Sangui BioTech), and IL-6 (Biosource) concentrations were determined by an ELISA. Plasma glucose concentration was determined enzymatically (Randox Laboratories) on a semiautomatic analyzer (COBAS MIRA plus, Roche).

Statistical Analysis

A two-way (trial x time) repeated-measures ANOVA was used to compare means. Where the repeated-measures ANOVA revealed a significant F ratio, differences were inspected with Tukey's honestly significant difference post hoc test. SPSS version 10 for Windows (SPSS, Chicago, IL) was used to calculate these statistics. The level of statistical significance accepted to reject the null hypothesis was P < 0.05. Data in the text, tables, and figures are means ± SE.


   RESULTS
TOP
 ABSTRACT
 METHODS
 RESULTS
DISCUSSION
 ACKNOWLEDGMENTS
 REFERENCES
 
During the Pla trial, there was a postexercise increase (P < 0.01) in neutrophil, lymphocyte, and monocyte concentrations, resulting in an increase in the total circulating leukocyte concentration (Table 1). The total leukocyte concentration remained elevated 2 h postexercise as a result of increased (P < 0.01) neutrophil and monocyte concentrations despite a return to normal of the lymphocyte concentration (Table 1). CHO ingestion attenuated (P < 0.01) the postexercise neutrophil, lymphocyte, and monocyte concentrations compared with the Pla trial, resulting in a reduced (P < 0.01) total leukocyte concentration (Table 1). Similarly, CHO ingestion compared with Pla reduced circulating neutrophil and monocyte concentrations at 2 h postexercise. No differences were observed between the 6.4% and 12.8% CHO trials (Table 1).


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  		Table 1. Circulating concentrations of leukocytes, neutrophils, lymphocytes and monocytes in response to exercise with and without carbohydrate ingestion

 
During the Pla trial there was a postexercise increase (P < 0.05) in CD3+CD4+ and CD3+CD8+ T cell concentrations, resulting in an increase (P < 0.01) in total circulating CD3+ T cells (Table 2). CHO ingestion attenuated (P < 0.05) the postexercise increase in CD3+CD4+ and CD3+CD8+ T cell concentrations compared with Pla, resulting in a decreased (P < 0.01) postexercise total CD3+ T cell concentration (Table 2). No differences were observed between the 6.4% and 12.8% CHO trials (Table 2). At 2 h postexercise, the CD3+CD8+ T cell concentration fell below (P < 0.05) preexercise concentrations (Table 2). This decline was accounted for primarily by a reduction in nonnaive CD8+ T cells (i.e., those CD8+ T cells expressing CD45RO) and was ameliorated during both 6.4% and 12.8% CHO trials (Table 2).


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  		Table 2. Circulating concentrations of CD3+ T cells and T cell subsets in response to exercise with and without carbohydrate ingestion

 
The circulating concentration and percentage of CD4+IFN-{gamma}+ T cells (TH1 cells), that is, CD4+ T cells expressing IFN-{gamma} after stimulation, were unchanged postexercise but fell below (P < 0.05) preexercise concentrations at 2 h postexercise (Table 3). CHO ingestion ameliorated this reduction, although no differences were observed between the 6.4% and 12.8% CHO trials (Table 3). To assess T cell function, we quantified the amount of cytokine produced after stimulation by obtaining the geometric mean fluorescence intensity of cytokine-positive cells. CD4+ T cell IFN-{gamma} production after stimulation was reduced immediately postexercise (P < 0.05) and 2 h postexercise (P < 0.05) during the Pla trial (Fig. 1A). However, CHO ingestion resulted in an attenuation of the suppression in CD4+ T cell IFN-{gamma} production at both immediately post and 2 h postexercise (Fig. 1A). Stimulated CD4+ T cell IFN-{gamma} production was particularly well maintained during the 12.8% CHO trial, although no statistically significant differences were observed between the 6.4% and 12.8% CHO trials.


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  		Table 3. Circulating cell counts and percentages of T cells positive for IFN-{gamma} and IL-4 in response to exercise with and without carbohydrate ingestion

 


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  		Fig. 1. Values are geometric mean fluorescence intensity (GMFI) expressed as the percentage of preexercise values of stimulated CD4+ T lymphocytes positive for interferon (IFN)-{gamma} (A) and interleukin (IL)-4 (B) in peripheral blood with [6.4% and 12.8% carbohydrate (CHO)] or without (Placebo) CHO ingestion. Blood samples were collected before (Preexercise), on completion of (Postexercise), and after 2 h of recovery from (2-h postexercise) 2.5 h of cycling exercise. Values are means ± SE; n = 7 subjects. *Interaction effect, significantly different from preexercise (P < 0.05). {ddagger}Interaction effect, significantly different from 12.8% CHO (P < 0.05).

 
The circulating concentration of CD4+ and CD8+ IL-4+ T cells (TH2/TC2 cells), that is, CD4+ and CD8+ T cells expressing IL-4 after stimulation, was unchanged after exercise and unaffected by CHO ingestion (Table 3). Furthermore, CD4+ and CD8+ T cell IL-4 production after stimulation was unchanged after exercise and unaffected by CHO ingestion (Figs. 1B and 2B).



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  		Fig. 2. Values are GMFI expressed as the percentage of preexercise values of stimulated CD8+ T lymphocytes positive for IFN-{gamma} (A) and IL-4 (B) in peripheral blood with (6.4% and 12.8% CHO) or without (Placebo) carbohydrate ingestion. Blood samples were collected before (Preexercise), on completion of (Postexercise), and after 2 h of recovery from (2-h postexercise) 2.5 h of cycling exercise. Values are means ± SE; n = 7 subjects. *Interaction effect, significantly different from preexercise (P < 0.01). {dagger}Interaction effect, significantly different from 6.4% CHO (P < 0.01). {ddagger}Interaction effect, significantly different from 12.8% CHO (P < 0.01).

 
There was a postexercise increase (P < 0.05) in the circulating concentration of CD8+IFN-{gamma}+ T cells (CD8+ T cytotoxic cells) during the Pla trial (Table 3). At 2 h postexercise, the concentration and percentage of CD8+IFN-{gamma}+ T cells was reduced (P < 0.05) below preexercise values. Furthermore, CHO ingestion attenuated both the postexercise increase in the circulating CD8+IFN-{gamma}+ T cells (P < 0.05) and the decline in CD8+IFN-{gamma}+ T cells at 2 h postexercise (P < 0.05) (Table 3). CD8+ T cell IFN-{gamma}+ production was suppressed postexercise (P < 0.05) and 2 h postexercise (P < 0.05) (Fig. 2A). Furthermore, CHO ingestion during both the 6.4% and 12.8% trials resulted in an attenuation of the suppression in IFN-{gamma} production (Fig. 2A).

The plasma ACTH concentration was increased (P < 0.01) postexercise and declined below (P < 0.05) preexercise values at 2 h postexercise (Table 4). A higher (P < 0.01) postexercise plasma cortisol concentration was observed after the Pla trial compared with the 6.4% and 12.8% CHO trials (Table 4). Whereas the plasma cortisol concentration at 2 h postexercise was lower (P < 0.01) than preexercise values during the 6.4% and 12.8% CHO trials, the plasma cortisol concentration remained elevated at 2 h postexercise during the Pla trial (Table 4). There was a significant (P < 0.05) increase in the postexercise plasma GH concentration during the Pla trial (Table 4). CHO ingestion resulted in an attenuation of the postexercise GH concentration (Table 4). The postexercise plasma glucose concentration fell below (P < 0.01) preexercise values and remained decreased 2 h postexercise (P < 0.01) during the Pla trial. CHO ingestion during exercise, particularly the 6.4% CHO solution, attenuated the decline in plasma glucose. The plasma IL-6 concentration was increased (P < 0.01) postexercise, and CHO ingestion attenuated this increase compared with placebo (P < 0.05).


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  		Table 4. Hormonal, metabolic, and cytokine responses to exercise with and without carbohydrate ingestion

 

   DISCUSSION
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
ACKNOWLEDGMENTS
 REFERENCES
 
The results of the present study demonstrate for the first time that CHO ingestion, which blunted the exercise-induced stress hormone response compared with Pla, attenuates the postexercise suppression of stimulated IFN-{gamma} production by CD4+ (TH1) and CD8+ (TC1) lymphocytes. Furthermore, CHO ingestion attenuated the postexercise decrease in the circulating concentration and percentage of CD4+IFN-{gamma}+ (TH1) and CD8+IFN-{gamma}+ (TC1) lymphocytes. However, exercise had no effect on the distribution or function of CD4+ and CD8+ IL-4-producing T cells.

The results of the present study are in agreement with previous work (8, 19) demonstrating a decreased concentration and percentage of CD4+IFN-{gamma}+ and CD8+IFN-{gamma}+ T lymphocytes in the circulation after prolonged exercise. In addition, no changes in the circulating concentration or percentage of CD4+IL-4+ T lymphocytes after exercise were observed. Our data extend previous observations (8, 19), demonstrating that CHO ingestion, which resulted in a blunted exercise-induced stress hormone response, resulted in an attenuation of the exercise-induced decrease in the circulating concentration of TH1 and TC1 lymphocytes.

Both catecholamines and glucocorticoids are known to be important mediators of T cell distribution (3, 18) and are likely to play a role in the exercise-induced modulation of type 1 T cell distribution. Accordingly, the differential effect of exercise on type 1 and type 2 lymphocyte distribution is most likely accounted for by the higher surface expression of {beta}2-adrenergic receptors on type 1 T cells (15) and by the fact that type 1 T cells are more sensitive to glucocorticoid-mediated effects (2, 7).

Alterations in the distribution of circulating TH1/TC1 and TH2/TC2 lymphocytes do not provide information on individual T cell function. Therefore, in the present study we quantified intracellular cytokine production, a critical aspect of TH1/TC1 and TH2/TC2 lymphocyte function, by obtaining the fluorescence intensity of IFN-{gamma} and IL-4 in both CD4+ and CD8+ T lymphocytes after stimulation. In support of our experimental hypothesis, exercise resulted in a suppression of stimulated CD4+ and CD8+ T lymphocyte IFN-{gamma} production. The primary aim of the present study was to examine the role of the exercise-induced stress hormone response on T cell cytokine production. Our results show that CHO ingestion, which resulted in a blunted exercise-induced stress hormone response, attenuated the suppression of stimulated IFN-{gamma} production by TH1 and TC1 lymphocytes after exercise. The best characterized of these hormones, with respect to a role in the regulation of T cell function, is the glucocorticoid hormone cortisol and it has been demonstrated that a synthetic glucocorticoid, dexamethasone, inhibits IFN-{gamma} production (7, 11, 12). It is therefore possible that in the present study an exercise-induced elevation in the circulating cortisol concentration may account for the suppressive effects of exercise on type 1 T cell IFN-{gamma} production. Contrary to our hypothesis, we observed no alteration in stimulated CD4+ and CD8+ T lymphocyte IL-4 production after exercise. Although several studies have observed a suppressive effect of glucocorticoids on T lymphocyte IL-4 production (7, 12, 14, 23), these studies used the synthetic glucocorticoid dexamethasone, which is considerably more potent in its immunosuppressive effects than endogenously produced cortisol. Given that stimulated IFN-{gamma} production by type 1 T cells is markedly more sensitive to the effects of glucocorticoids than IL-4 production by type 2 T cells (2, 7, 12), it is possible that the exercise-induced elevation in the circulating cortisol concentration was not of sufficient magnitude to cause a suppression in IL-4 production.

With regard to the mechanism by which CHO ingestion attenuates the exercise-induced stress hormone response, it is well established that prolonged exercise results in an increase in the plasma concentration of IL-6, the primary source of which is the contracting skeletal muscle (5). Given that CHO ingestion during exercise attenuates the exercise-induced increase in the IL-6 concentration due to the inhibition of IL-6 release from the contracting muscle (6), and that IL-6 is known to stimulate the hypothalamic-pituitary-adrenal axis (20), this may be the mechanism by which CHO ingestion attenuates the exercise-induced suppression of type 1 T cell IFN-{gamma} production. In addition, given that hypoglycemia stimulates the activation of glucose-sensitive neurons within the hypothalamus resulting in an increase in the plasma concentration of ACTH and cortisol (21) and that CHO ingestion prevented the exercise-induced fall in the plasma glucose concentration, this may be a further mechanism by which CHO ingestion attenuates the exercise-induced suppression of type 1 T cell IFN-{gamma} production.

In conclusion, 2.5 h of exercise with placebo ingestion resulted in a decrease in the number and percentage of circulating CD4+IFN-{gamma}+ (TH1) and CD8+IFN-{gamma}+ (TC1) T lymphocytes. Furthermore, stimulated IFN-{gamma} production by CD4+ and CD8+ T lymphocytes was decreased after exercise. However, CHO ingestion, which blunted the exercise-induced stress hormone response, attenuated both the decrease in the circulating number and percentage of CD4+IFN-{gamma}+ and CD8+IFN-{gamma}+ T lymphocytes and T lymphocyte intracellular cytokine production. No exercise-induced alterations either in the circulating percentage and number or in cytokine production were observed for CD4+ or CD8+ IL-4+ (TH2/TC2) T lymphocytes.


   ACKNOWLEDGMENTS
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGMENTS
REFERENCES
 
The authors gratefully acknowledge the financial support of GlaxoSmithKline consumer healthcare for providing funding for the present study.


   FOOTNOTES
 

Address for reprint requests and other correspondence: M. Gleeson, School of Sport and Exercise Sciences, Loughborough Univ., Loughborough LE11 3TU, United Kingdom (E-mail: m.gleeson{at}lboro.ac.uk)

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


   REFERENCES
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGMENTS
 REFERENCES
 

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