Effects of aging on human leukocytes (part II): immunophenotyping of adaptive immune B and T cell subsets

Abstract

Immunosenescence results from a continuous deterioration of immune responses resulting in a decreased response to vaccines. A well-described age-related alteration of the immune system is the decrease of de novo generation of T and B cells. In addition, the accumulation of memory cells and loss of diversity in antigen specificities resulting from a lifetime of exposure to pathogens has also been described. However, the effect of aging on subsets of γδTCR(+) T cells and Tregs has been poorly described, and the efficacy of the recall response to common persistent infections in the elderly remains obscure. Here, we investigated alterations in the subpopulations of the B and T cells among 24 healthy young (aged 19-30) and 26 healthy elderly (aged 53-67) individuals. The analysis was performed by flow cytometry using freshly collected peripheral blood. γδTCR(+) T cells were overall decreased, while CD4(+)CD8(-) cells among γδTCR(+) T cells were increased in the elderly. Helios(+)Foxp3(+) and Helios(-)Foxp3(+) Treg cells were unaffected with age. Recent thymic emigrants, based on CD31 expression, were decreased among the Helios(+)Foxp3(+), but not the Helios(-)Foxp3(+) cell populations. We observed a decrease in Adenovirus-specific CD4(+) and CD8(+) T cells and an increase in CMV-specific CD4(+) T cells in the elderly. Similarly, INFγ(+)TNFα(+) double-positive cells were decreased among activated T cells after Adenovirus stimulation but increased after CMV stimulation. The data presented here indicate that γδTCR(+) T cells might stabilize B cells, and functional senescence might dominate at higher ages than those studied here.

Fig. 1

Global view of all analyzed populations. Each circle represents a population, and the arrow indicates the relationship between populations. Diamond shape indicates live cells. a Cell fraction (left half circle) or cell count (right half circle). White indicates no difference; the fill color indicates significant decrease or increase in the elderly compared to the young. Difference tested by Wilcoxon rank-sum test, p values <0.05 were considered significant. b Fraction of cells normalized per cell population. c Estimated cells/μl normalized per cell population. Bold circle borders indicate population with significant difference

Fig. 2

Responsiveness in CD4⁺ and CD8⁺ T cells. Cells were stimulated with indicated antigen for 6 h. Shown are individual measurements after subtraction of values of unstimulated cells and boxplots of indicated populations and total cell number/μl whole blood for a CD40L⁺ among CD4⁺ cells and b INFγ⁺ among CD8⁺ cells. c Distribution of number of co-expressed cytokines within the activated cells. d Distribution of individual cytokine combinations among the activated cells. The numbers above the plot-sections indicate the number of co-expressed cytokines. The box indicates the 25th, 50th, and 75th percentiles, the whiskers the minimum and maximum values excluding outliers. Gating was performed as described in Supplementary Fig. 1b. Wilcoxon rank-sum test was used to test for differences and p values <0.05 are reported

Fig. 3

CD4⁺Foxp3⁺ Tregs in the elderly and in the young. Shown are individual measurements and boxplots of indicated populations and total cell number/μl whole blood for a Foxp3⁺Helios⁺ among CD4⁺, b Foxp3⁺Helios⁻ among CD4⁺, c Foxp3⁺Helios⁺CD31⁺ among CD4⁺, and d Foxp3⁺Helios⁻CD31⁺ among CD4⁺. The box indicates the 25th, 50th, and 75th percentiles, the whiskers the minimum and maximum values excluding outliers. Gating was performed as described in Supplementary Fig. 2B. Wilcoxon rank-sum test was used to test for differences and p values <0.05 are reported