Elsevier

Immunobiology

Volume 216, Issue 4, April 2011, Pages 477-484
Immunobiology

The immune responses of central and effector memory BCG-specific CD4+ T cells in BCG-vaccinated PPD+ donors were modulated by Treg cells

https://doi.org/10.1016/j.imbio.2010.09.003Get rights and content

Abstract

Most of the studies evaluating the function of tuberculosis (TB)-specific T cells were only based on the ability to produce cytokines, which may not fully reflect the function of T cells. In the present study, we confirmed that Bacille Calmette Guerin (BCG) could significantly induce cytokine production by CD4+ T cells from BCG-vaccinated PPD+ donors. In addition, CD4+ T cells were activated, divided and proliferated in response to BCG stimulation. Phenotypic analysis showed that IFN-γ+CD4+ T cells displayed CD45RACCR7+/−CD62L, indicating that these CD4+ T cells were central and effector memory cells. The analysis of cytokine profiles demonstrated that most of BCG-specific BrdU+CD4+ T cells produced Th1 cytokines in response to polyclonal stimulation. In addition, we found that regulatory T cells (Treg) suppressed BCG-induced proliferation and IFN-γ production by memory CD4+ T cells. The suppressive effects of Treg on BCG-specific responses of CD4+ T cells could be partially reversed by blocking the production of IL-10. Taken together, our results demonstrated that functional central and effector memory BCG-specific CD4+ T cells could be detected based on the activation, proliferation and division of these cells, and modulated by Treg in PBMCs from BCG-vaccinated PPD+ donors.

Introduction

Studies in humans and animal models demonstrated that CD4+ T cells played an important role in protective immunity to Mycobacterium tuberculosis (Boom et al., 2003, Kaufmann, 2006, Kaufmann and Winau, 2005). The HIV pandemic provided direct evidence that loss of CD4+ T cell numbers and function resulted in progressive primary infection, reactivation of endogenous M. tuberculosis and enhanced susceptibility to re-infection (Selwyn et al. 1989). Early studies in mice demonstrated that protection against M. tuberculosis could be adoptively transferred by CD4+ T cells (Orme and Collins, 1984, Orme and Collins, 1983). In vivo depletion of CD4+ T cells with anti-CD4 mAb enhanced susceptibility to the infection of M. tuberculosis (Muller et al., 1987, Tascon et al., 1998, Caruso et al., 1999). Human CD4+ T cells activated by M. tuberculosis antigens secreted IFN-γ, TNF-α and IL-2. IFN-γ and TNF-α contributed to the recruitment of monocytes and granulocytes and activated antimicrobial activity of macrophages, while IL-2 could induce proliferation and amplified the effector function of activated CD4+ and CD8+ T cells (Rook et al., 1986, Flesch and Kaufmann, 1987, Flesch and Kaufmann, 1990).

BCG is the only available vaccine preventing tuberculosis used clinically. It has been demonstrated that BCG protects against disseminated forms of TB infection in childhood but not in adults (Kaufmann, 2005, Rook et al., 2005). In correlation with the protective immune responses, recent studies demonstrated that BCG vaccination of human newborns induced specific and functional CD4+ and CD8+ T cell responses (Hanekom, 2005, Soares et al., 2008, Murray et al., 2006). Several studies have demonstrated that Treg are increased both at disease sites and in peripheral blood in patients with TB (Hougardy et al., 2007, Li et al., 2007, Guyot-Revol et al., 2006). Naturally occurring CD4+CD25+ T cells (nTreg), which constitutively express Foxp3, have been suggested to inhibit the effector functions of CD4+ and CD8+ T cells (Fletcher, 2007, Belkaid and Rouse, 2005, Hilchey and Bernstein, 2007).

In the current study, we found that BCG significantly induced cytokine production, activation, proliferation and division of CD4+ T cells. After polyclonal stimulation, BCG-specific CD4+ T cells produced Th1-like cytokines. Importantly, we found that the proliferation and cytokine production of CD4+ T cells were inhibited by Treg and partially reversed by blocking of IL-10 production. Taken together, these data demonstrate that BCG-specific CD4 T cells are persistent in BCG-vaccinated adults and the specific responses of CD4+ T cells are modulated by Treg cells, implying the possibility of enhancing immune responses of TB infection by down-regulating the function of Treg cells.

Section snippets

Study participants

Twenty-four healthy volunteers comprised of 13 females and 11 males aged from 19 to 53 years were recruited from Zhongshan School of Medicine, Sun Yat-sen University. Heparinized blood samples (10 IU mL−1 final) were collected from each donor under protocols approved by the Medical School Review Board at Sun Yat-sen University, China. The participants were found to be skin test positive for PPD. Individuals that had been diagnosed with HIV, hepatitis B virus (HBV), hepatitis C virus (HCV), or

BCG induces IFN-γ production by PBMCs

To determine whether BCG could induce IFN-γ production, PBMCs were stimulated with or without BCG plus anti-CD28 mAb. After incubation for 72 h, the culture supernatants were collected and assessed for IFN-γ production by ELISA. The results from 24 individuals revealed that BCG significantly induced higher levels of IFN-γ production compared to medium alone (p < 0.05, Fig. 1A). These results were further confirmed by the detection of frequency of IFN-γ producing cells by ELISPOT assay after PBMCs

Discussion

BCG, a viable attenuated vaccine, with more than 3 billion doses, is one of the most widely used vaccines worldwide. However, it had been considered that BCG could only prevent severe childhood tuberculosis whereas insufficient for control of adult tuberculosis, especially in developing countries (Kaufmann, 2005, Kaufmann, 2006Kaufmann and Winau, 2005, Rook et al., 2005, Colditz et al., 1995, Colditz et al., 1994). One explanation assumes that BCG is a potent inducer of CD4+ T cells, which are

Acknowledgments

This work was supported by the 115 grant (No. 2008ZX10003011), National Nature Science Foundation of China (No. 30872300) and the National Key Basic Research Program of China (973; No. 2007CB512404).

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