Aging and human CD4+ regulatory T cells

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Abstract

Alterations in immunity that occur with aging likely contribute to the development of infection, malignancy and inflammatory diseases. Naturally occurring CD4+ regulatory T cells (Treg) expressing high levels of CD25 and forkhead box P3 (FOXP3) are essential for regulating immune responses. Here we investigated the effect of aging on the number, phenotypes and function of CD4+ Treg in humans. The frequency and phenotypic characteristics of CD4+, FOXP3+ T cells as well as their capacity to suppress inflammatory cytokine production and proliferation of CD4+, CD25 T cells (target cells) were comparable in young (age ≤40) and elderly (age ≥65) individuals. However, when CD4+, FOXP3+ Treg and CD4+, CD25 T cells were co-cultured at a ratio of 1:1, the production of anti-inflammatory cytokine IL-10 from CD4+, CD25 T cells was more potently suppressed in the elderly than in the young. This finding was not due to changes in CTLA-4 expression or apoptosis of CD4+, FOXP3+ Treg and CD4+, CD25 T cells. Taken together, our observations suggest that aging may affect the capacity of CD4+, FOXP3+ T cells in regulating IL-10 production from target CD4+ T cells in humans although their other cellular characteristics remain unchanged.

Introduction

Alterations in the immune system that occur with aging (Effros et al., 2003, Frasca et al., 2005, Goronzy and Weyand, 2005, Linton and Dorshkind, 2004, Miller, 1999) likely contribute to increased risk of infection and malignancy in the elderly (Effros et al., 2003, Goronzy and Weyand, 2005, Linton and Dorshkind, 2004, Miller, 1999). Although the exact cause for such findings is yet to be determined, it is conceivable that aging may cause diminished immunity against microorganisms and malignant cells via altering the mechanisms involved in generating immune responses. The latter response is inevitably accompanied by some degrees of inflammation, which should be kept in balance. The increased frequency of infection and tumors in the elderly suggests the possibility of reduced inflammatory responses with aging (Effros et al., 2003, Goronzy and Weyand, 2005, Linton and Dorshkind, 2004, Miller, 1999). However, aging appears to be associated with dysregulated inflammation as suggested by increased plasma levels of inflammatory cytokines IL-6 and TNF-α that predict an increased mortality risk independently of other risk factors in the elderly (Huang et al., 2005, Krabbe et al., 2004). Furthermore, some chronic inflammatory diseases such as polymyalgia rheumatica and giant cell arteritis are more commonly found in the elderly (Hasler and Zouali, 2005, Hunder, 2000), and metabolic disorders like atherosclerosis and type II diabetes that develop with aging are now viewed as chronic inflammatory disorders driven in part by an imbalance between pro- and anti-inflammatory cytokines (Huang et al., 2005, Vasto et al., 2007). However, it is largely unknown about the mechanisms underlying dysregulated inflammation with aging in humans.

A population of naturally occurring CD4+, CD25+ T cells with immune regulatory function exists in humans and mice (Sakaguchi et al., 1995, von Boehmer, 2005, Wing et al., 2006). However, CD25 as a single marker for CD4+ Treg has been challenged since T cells stimulated by T cell receptor (TCR) triggering can up-regulate the expression of this molecule. Recent studies demonstrate that the forkhead family transcriptional factor FOXP3 is expressed in CD4+, CD25+ Treg and that transfection of the same molecule to CD4+, CD25 T cells, which do not have regulatory function, confers the immune regulatory property (reviewed in (Ziegler, 2006)) (Fontenot et al., 2003, Hori et al., 2003, Khattri et al., 2003). Furthermore, mutations in the Foxp3 gene have been found in scurfy mice with X-linked lymphoproliferative disease as well as in humans with immune dysregulation, polyendocrinopathy, enteropathy, and X-linked syndrome (IPEX) (Bennett et al., 2001, Brunkow et al., 2001). Although the mechanism(s) of immune regulation by CD4+, CD25+ Treg is not fully understood, such regulation is dependent on cell contact rather than on soluble factors like cytokines (reviewed in (von Boehmer, 2005, Wing et al., 2006)). The target cells for suppression appear to be both T cells and antigen presenting cells (APC) (von Boehmer, 2005, Wing et al., 2006).

In the current study, we investigated whether aging affects the number, phenotype and function of human CD4+ Treg, defined by their expression of CD25 and FOXP3. In particular, we focused on the direct inhibitory effect of Treg on conventional CD4+ T cells in the absence of APC since the latter cells can affect T cell function. The results of our study showed that the frequency, phenotypic characteristics and anti-proliferative function of CD4+, FOXP3+ Treg were comparable in the young (age ≤40) and the elderly (age ≥65). However, when CD4+, CD25 T cells were stimulated in the presence of the same number of CD4+, FOXP3+ Treg, the production of anti-inflammatory cytokine IL-10 from the former cells was more potently suppressed in the elderly than in the young. These findings suggest that aging may affect the capacity of CD4+, FOXP3+ Treg in regulating IL-10 production from CD4+, CD25 T cells in humans although other cellular characteristics of CD4+, FOXP3+ T cells remain unchanged with aging.

Section snippets

Human subjects

Healthy elderly (age ≥65, n = 32) and young subjects (age ≤40, n = 29) were recruited for this study (mean age ± SD, 77.1 ± 7.8 and 30.5 ± 5.9). There was no gender difference between the two groups (P = 0.427 and 0.576 by Fisher's exact tests for phenotypic and functional studies, respectively). Individuals who were taking immunosuppressive drugs or who had any disease potentially affecting the immune system including autoimmune diseases, infectious diseases, malignancy, diabetes, and asthma were excluded

The relationships of CD25, FOXP3 and IL-7 receptor α chain (IL-7Rα) expression by CD4+ T cells are similar between young and elderly individuals

CD25 has been used as a marker for identifying naturally occurring CD4+ Treg with immune regulatory function (Sakaguchi and Sakaguchi, 2005, Wang et al., 2006, Yamaguchi and Sakaguchi, 2006). However, the expression of CD25, which can be up-regulated upon T cell activation, on CD4+ T cells is heterogeneous, and there is no established limit for the minimal level of CD25 expression that defines a homogeneous Treg population. Now FOXP3 is considered the best marker for CD4+ Treg although cell

Discussion

Alterations in the immune system that occurs with aging likely contribute to the development of infection, malignancy and inflammatory diseases in the elderly (Effros et al., 2003, Goronzy and Weyand, 2005, Hasler and Zouali, 2005, Huang et al., 2005, Krabbe et al., 2004, Linton and Dorshkind, 2004, Miller, 1999). Although the exact cause for such alterations is yet to be determined, changes in regulating inflammatory immune responses may account in part for developing such conditions with

Conflict of interest

The authors have no conflicting financial interests in this work.

Acknowledgements

We thank Dr. Alexia Belperron for critical review of this manuscript and Lynne Iannone, Barbara Foster and Yale Center for Clinical Investigation for assisting the recruitment of human subjects. This work was supported in part by grants from the National Institute of Health (K08 AR49444, R01 AG028069, R21 AG030834), the Arthritis Foundation, the American Foundation for Aging Research and Claude D. Pepper Older Americans Independence Center (P30AG21342 NIH/NIA). Insoo Kang is a recipient of the

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