Austin B. Bigley

The effects of age and latent cytomegalovirus infection on the redeployment of CD8+ T cell subsets in response to acute exercise in humans.

Dynamic exercise evokes a rapid redeployment of cytotoxic T cell subsets with high expression of β2 adrenergic receptors, presumably to enhance immunosurveillance during acute stress. As this response is affected by age and infection history, this study examined latent CMV infection as a potential confounder to age-related differences in blood CD8+ T-cell responses to exercise. Healthy young (n=16) and older (n=16) humans counterbalanced by CMV IgG serostatus (positive or negative) exercised for 30-min at ∼80% peak cycling power. Those with CMV redeployed ∼2-times more CD8+ T-cells and ∼6-times more KLRG1+/CD28- and CD45RA+/CCR7- CD8+ subsets than non-infected exercisers. Seronegative older exercisers had an impaired redeployment of total CD8+ T-cells, CD45RA+/CCR7+ and KLRG1-/CD28+ CD8+ subsets compared to young. Redeployed CD8+ T-cell numbers were similar between infected young and old. CMVpp65 specific CD8+ cells in HLA/A2(∗) subjects increased ∼2.7-fold after exercise, a response that was driven by the KLRG1+/CD28-/CD8+ subset. Stimulating PBMCs before and after exercise with CMVpp65 and CMV IE-1 antigens and overlapping peptide pools revealed a 2.1 and 4.4-fold increases in CMVpp65 and CMV IE-1 IFN-γ secreting cells respectively. The breadth of the T cell response was maintained after exercise with the magnitude of the response being amplified across the entire epitope repertoire. To conclude, latent CMV infection overrides age-related impairments in CD8+ T-cell redeployment with exercise. We also show for the first time that many T-cells redeployed with exercise are specific to CMVpp65 and CMV IE-1 antigens, have broad epitope specificity, and are mostly of a high-differentiated effector memory phenotype.

Latent cytomegalovirus infection enhances anti-tumour cytotoxicity through accumulation of NKG2C+ NK cells in healthy humans

Cytomegalovirus (CMV) infection markedly expands NKG2C+/NKG2A− NK cells, which are potent killers of infected cells expressing human leucocyte antigen (HLA)‐E. As HLA‐E is also over‐expressed in several haematological malignancies and CMV has been linked to a reduced risk of leukaemic relapse, we determined the impact of latent CMV infection on NK cell cytotoxicity against four tumour target cell lines with varying levels of HLA‐E expression. NK cell cytotoxicity against K562 (leukaemia origin) and U266 (multiple myeloma origin) target cells was strikingly greater in healthy CMV‐seropositive donors than seronegative donors and was associated strongly with target cell HLA‐E and NK cell NKG2C expression. NK cell cytotoxicity against HLA‐E transfected lymphoma target cells (221.AEH) was ∼threefold higher with CMV, while NK cell cytotoxicity against non‐transfected 721.221 cells was identical between the CMV groups. NK cell degranulation (CD107a+) and interferon (IFN)‐γ production to 221.AEH cells was localized almost exclusively to the NKG2C subset, and antibody blocking of NKG2C completely eliminated the effect of CMV on NK cell cytotoxicity against 221.AEH cells. Moreover, 221.AEH feeder cells and interleukin (IL)−15 were found to expand NKG2C+/NKG2A– NK cells preferentially from CMV‐seronegative donors and increase NK cell cytotoxicity against HLA‐E+ tumour cell lines. We conclude that latent CMV infection enhances NK cell cytotoxicity through accumulation of NKG2C+ NK cells, which may be beneficial in preventing the initiation and progression of haematological malignancies characterized by high HLA‐E expression.

Latent CMV infection enhances anti‐tumor cytotoxicity through accumulation of NKG2C+ NK‐cells in healthy humans

Cytomegalovirus (CMV) infection markedly expands NKG2C+/NKG2A− NK cells, which are potent killers of infected cells expressing human leucocyte antigen (HLA)‐E. As HLA‐E is also over‐expressed in several haematological malignancies and CMV has been linked to a reduced risk of leukaemic relapse, we determined the impact of latent CMV infection on NK cell cytotoxicity against four tumour target cell lines with varying levels of HLA‐E expression. NK cell cytotoxicity against K562 (leukaemia origin) and U266 (multiple myeloma origin) target cells was strikingly greater in healthy CMV‐seropositive donors than seronegative donors and was associated strongly with target cell HLA‐E and NK cell NKG2C expression. NK cell cytotoxicity against HLA‐E transfected lymphoma target cells (221.AEH) was ∼threefold higher with CMV, while NK cell cytotoxicity against non‐transfected 721.221 cells was identical between the CMV groups. NK cell degranulation (CD107a+) and interferon (IFN)‐γ production to 221.AEH cells was localized almost exclusively to the NKG2C subset, and antibody blocking of NKG2C completely eliminated the effect of CMV on NK cell cytotoxicity against 221.AEH cells. Moreover, 221.AEH feeder cells and interleukin (IL)−15 were found to expand NKG2C+/NKG2A– NK cells preferentially from CMV‐seronegative donors and increase NK cell cytotoxicity against HLA‐E+ tumour cell lines. We conclude that latent CMV infection enhances NK cell cytotoxicity through accumulation of NKG2C+ NK cells, which may be beneficial in preventing the initiation and progression of haematological malignancies characterized by high HLA‐E expression.

Training status and sex influence on senescent T-lymphocyte redistribution in response to acute maximal exercise

PURPOSE Investigate training status and sex effects on the redistribution of senescent and naïve T-lymphocytes following acute exercise. METHODS Sixteen (8 male, 8 female) trained (18.3±1.7yr) soccer players (Tr) and sixteen (8 male, 8 female) untrained (19.3±2.0yr) controls (UTr) performed a treadmill running test to volitional exhaustion. Blood lymphocytes were isolated before (Pre), immediately post, and 1-h post-exercise for assessment of cell surface expression of CD28 and CD57 on CD4(+) and CD8(+) T-lymphocyte subsets. Plasma was used to determine cytomegalovirus (CMV) serostatus. RESULTS Exercise elicited a redistribution of T-lymphocyte subsets. Senescent CD4(+) and CD8(+) T-lymphocytes increased by 42.4% and 45.9% respectively, while naïve CD4(+) and CD8(+) T-lymphocytes decreased by 8.7% and 22.5% respectively in response to exercise. A main effect (P<0.05) of training status was observed for senescent CD4(+), CD8(+) and naïve CD8(+) T-lymphocytes: UTr had a higher proportion of senescent and a lower proportion of naïve CD8(+) T-lymphocytes than Tr. A main effect (P<0.05) of sex was observed in senescent CD4(+), CD8(+) and naïve CD4(+), CD8(+) T-lymphocytes. Males had a higher proportion of senescent and lower proportion of naïve T-lymphocytes than females. A sex-by-training status interaction (P<0.05) was observed for the senescent and naïve CD4(+) T-lymphocytes (but not CD8(+)) with the highest percentage of senescent and lowest percentage of naïve T-lymphocytes observed in UTr males. CMV exerted a significant main covariate effect (P<0.05) in the senescent and naïve (P<0.05) CD8(+) T-lymphocytes but not in the senescent and naïve CD4(+) T-lymphocytes. CONCLUSION This study highlights important sex and training status differences in the senescent and naïve T-lymphocyte redistribution in response to exercise that warrants further investigation.

Dichotomous effects of latent CMV infection on the phenotype and functional properties of CD8+ T-cells and NK-cells

CMV markedly alters the phenotype and function of NK-cells and T-cells and has been linked to immunosenescence. We show here that subjects with effective CMV control (evidenced by low CMV IgG titers) have functional responses to CMV that are driven by either NKG2C+ NK-cells or CMV-specific T-cells (15 of 24 subjects), but not both. These data indicate that people with effective CMV control are either NK-cell or T-cell responders, and corroborates the idea that NK-cells have rheostat-like properties that regulate anti-viral T-cell responses. Whether or not lifelong CMV control through either NK-cell or T-cell responses have implications for immunosenescence remains to be determined.

The Effects of Age and Latent Cytomegalovirus Infection on NK-Cell Phenotype and Exercise Responsiveness in Man

The redeployment of NK-cells in response to an acute bout of exercise is thought to be an integral component of the “fight-or-flight” response, preparing the body for potential injury or infection. We showed previously that CMV seropositivity impairs the redeployment of NK-cells with exercise in the young. In the current study, we examined the effect of aging on the redeployment of NK-cells with exercise in the context of CMV. We show here that CMV blunts the exercise-induced redeployment of NK-cells in both younger (23–39 yrs) and older (50–64 yrs) subjects with older CMVneg subjects showing the largest postexercise mobilization and 1 h postexercise egress of NK-cells. The blunted exercise response in CMVpos individuals was associated with a decreased relative redeployment of the CD158a+ and CD57+ NK-cell subsets in younger and older individuals. In addition, we show that aging is associated with a CMV-independent increase in the proportion of NK-cells expressing the terminal differentiation marker CD57, while CMV is associated with an age-dependent decrease in the proportion of NK-cells expressing the inhibitory receptors KLRG1 (in the younger group) and CD158a (in the older group). Collectively, these data suggest that CMV may decrease NK-cell mediated immunosurveillance after exercise in both younger and older individuals.

The effects of age and viral serology on γδ T-cell numbers and exercise responsiveness in humans

γδ T-cells are cytotoxic effector cells that preferentially migrate to peripheral tissues and recognize many types of antigen. We examined the effects of age and viral serology on the exercise responsiveness of γδ T-cells. Blood was collected from 17 younger (age: 23-35yrs) and 17 older (50-64yrs) healthy males matched for cytomegalovirus (CMV), Epstein-Barr virus, herpes simplex virus-1 and Parvovirus B19 serologic status before and after a single bout of cycling exercise. Older had lower numbers and proportions of γδ T-cells than younger, while CMV was associated with increased numbers and proportions of γδ T-cells in younger but not older. Exercise evoked a ∼2-fold increase in circulating γδ T-cell numbers. The magnitude of this response was 3-times greater in younger compared to older, and 1.6-times greater in younger CMV-infected compared to younger non CMV-infected. To conclude, γδ T-cell numbers and exercise responsiveness decreases with age and may contribute to impaired immunosurveillance after acute acute physical stress.

Mobilizing Immune Cells With Exercise for Cancer Immunotherapy

Hematopoietic stem cell (HSC) transplantation and adoptive transfer immunotherapy are effective in treating blood cancers and posttransplant infections, but low-circulating cell numbers in patients and donors are oftentimes a limiting factor. We postulate that a single exercise bout will increase the yield of patient- and donor-derived HSCs and cytotoxic lymphocytes to improve this form of treatment for cancer patients.

T-lymphocyte populations following a period of high volume training in female soccer players.

PURPOSE To investigate the T-lymphocyte response to a period of increased training volume in trained females compared to habitual activity in female controls. METHODS Thirteen trained female (19.8 ± 1.9 yrs) soccer players were monitored during a two-week long high volume training period (increased by 39%) and thirteen female untrained (20.5 ± 2.2 yrs) controls were monitored during two-weeks of habitual activity. Blood lymphocytes, collected at rest, were isolated before and after the two-week period. Isolated lymphocytes were assessed for the cell surface expression of the co-receptor CD28, a marker of T-lymphocyte naivety, and CD57 a marker used to identify highly-differentiated T-lymphocytes. Co-expression of these markers was identified on helper CD4(+) and cytotoxic CD8(+) T-lymphocytes. In addition a further population of γδ(+) T-lymphocytes were identified. Plasma was used to determine Cytomegalovirus (CMV) serostatus. RESULTS No difference was observed in the T-lymphocyte populations following the two-week period of increased volume training. At baseline the number of total CD3(+), cytotoxic CD8(+), naïve (CD8(+) CD28(+) CD57(-)), intermediate (CD8(+) CD28(+) CD57(+)) T-lymphocytes and the number and proportion of γδ(+) T-lymphocytes were greater in the trained compared to the untrained females (p<0.05). The proportion of CD4(+)T-lymphocytes was greater in the untrained compared to the trained (p<0.05), in turn the CD4(+):CD8(+) ratio was also greater in the untrained females (p<0.05). Inclusion of percentage body fat as a covariate removed the main effect of training status in all T-lymphocyte sub-populations, with the exception of the γδ(+) T-lymphocyte population. 8% of the untrained group was defined as positive for CMV whereas 23% of the trained group was positive for CMV. However, CMV was not a significant covariate in the analysis of T-lymphocyte proportions. CONCLUSION The period of high volume training had no effect on T-lymphocyte populations in trained females. However, baseline training status differences were evident between groups. This indicates that long-term exercise training, as opposed to short-term changes in exercise volume, appears to elicit discernible changes in the composition of the blood T-lymphocyte pool.

CMV amplifies T-cell redeployment to acute exercise independently of HSV-1 serostatus.

PURPOSE Latent cytomegalovirus (CMV) infection has been shown to alter the lymphocyte response to acute aerobic exercise, likely due to the corresponding increase in exercise-responsive memory CD8(+) T cells. It is unknown if latent infection with another herpesvirus, herpes simplex virus 1 (HSV-1), also plays a role in shaping the lymphocyte response to exercise. METHODS Thirty-two men (ages 39.3 ± 14.7 yr) counterbalanced by CMV and HSV-1 serostatus (positive/negative) cycled for 30 min at ∼80% peak power. Blood sampled before, immediately after, and 1 h after exercise was analyzed by flow cytometry for T-cell subset enumeration. RESULTS In resting blood, HSV-1(+) had fewer lymphocytes, CD4(+) T cells, KLRG1(-) CD28(+) CD4(+) T cells, and CD45RA(-)CCR7(+)CD4(+) T cells than HSV-1(-), whereas CMV(+) had increased numbers of lymphocytes, CD8(+) T cells, KLRG1(+)CD28(-)CD4(+) and CD8(+) T cells, and CD45RA(+)CCR7(-)CD8(+) T cells and a lower CD4:CD8 T-cell ratio than CMV(-). After exercise, CMV(+) had a greater mobilization of CD8(+) T cells, KLRG1+CD28(-)CD4+ and CD8(+) T cells, and CD45RA+CCR7(-)CD8+ T cells independently of HSV-1 serostatus, as well as a greater egress of these subsets 1 h after exercise. HSV serostatus did not influence total CD8(+) T-cell response to exercise. CONCLUSIONS The impact of latent CMV infection on the redeployment of T-cell subsets with exercise is independent of HSV-1 infection. This is most likely due to the unique ability of CMV to alter the composition of the memory T-cell pool in favor of exercise-responsive T-cell subsets.