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Distinct differences in the expansion and phenotype of TB10.4 specific CD8 and CD4 T cells after infection with Mycobacterium tuberculosis.

Hoang TT, Nansen A, Roy S, Billeskov R, Aagaard C, Elvang T, Dietrich J, Andersen P - PLoS ONE (2009)

Bottom Line: Recently we and others have identified CD8 and CD4 T cell epitopes within the highly expressed M. tuberculosis protein TB10.4.We focused on T cells directed to two epitopes in TB10.4; the MHC class I restricted epitope TB10.4 (3-11) (CD8/10.4 T cells) and the MHC class II restricted epitope TB10.4 (74-88) (CD4/10.4 T cells).In addition, the observed strong expansion of CD8 T cells in the late stages of infection could have implications for the development of post exposure vaccines against latent TB.

View Article: PubMed Central - PubMed

Affiliation: Department of Infectious Disease Immunology, Statens Serum Institut, Copenhagen, Denmark.

ABSTRACT

Background: Recently we and others have identified CD8 and CD4 T cell epitopes within the highly expressed M. tuberculosis protein TB10.4. This has enabled, for the first time, a comparative study of the dynamics and function of CD4 and CD8 T cells specific for epitopes within the same protein in various stages of TB infection.

Methods and findings: We focused on T cells directed to two epitopes in TB10.4; the MHC class I restricted epitope TB10.4 (3-11) (CD8/10.4 T cells) and the MHC class II restricted epitope TB10.4 (74-88) (CD4/10.4 T cells). CD4/10.4 and CD8/10.4 T cells displayed marked differences in terms of expansion and contraction in a mouse TB model. CD4/10.4 T cells dominated in the early phase of infection whereas CD8/10.4 T cells were expanded after week 16 and reached 5-8 fold higher numbers in the late phase of infection. In the early phase of infection both CD4/10.4 and CD8/10.4 T cells were characterized by 20-25% polyfunctional cells (IL-2(+), IFN-gamma(+), TNF-alpha(+)), but whereas the majority of CD4/10.4 T cells were maintained as polyfunctional T cells throughout infection, CD8/10.4 T cells differentiated almost exclusively into effector cells (IFN-gamma(+), TNF-alpha(+)). Both CD4/10.4 and CD8/10.4 T cells exhibited cytotoxicity in vivo in the early phase of infection, but whereas CD4/10.4 cell mediated cytotoxicity waned during the infection, CD8/10.4 T cells exhibited increasing cytotoxic potential throughout the infection.

Conclusions/significance: Our results show that CD4 and CD8 T cells directed to epitopes in the same antigen differ both in their kinetics and functional characteristics throughout an infection with M. tuberculosis. In addition, the observed strong expansion of CD8 T cells in the late stages of infection could have implications for the development of post exposure vaccines against latent TB.

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The bacterial load in the lungs throughout infection shown as Log10 CFU.Mice were challenged by the aerosol route with virulent M. tuberculosis. At the indicated timepoints 3–6 mice were killed and the bacterial burden (CFU) was measured in the lung.
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pone-0005928-g002: The bacterial load in the lungs throughout infection shown as Log10 CFU.Mice were challenged by the aerosol route with virulent M. tuberculosis. At the indicated timepoints 3–6 mice were killed and the bacterial burden (CFU) was measured in the lung.

Mentions: Anti-TB10.4 3–11 CD8 T cells and anti-TB10.4 74–88 CD4 T cells (hereafter called CD8/10.4 and CD4/10.4 cells) represent a significant proportion of the T cells induced by infection with M.tb [3]. To study and compare the dynamic development of these T cells, CB6F1 (BALB/c×C57BL/6) mice were infected by the aerosol route with M.tb Erdman and the T cell immune response against TB10.4 74–88 and TB10.4 3–11 was analyzed at different time points following infection. Epitope recognition was assessed using TB10.4 74–88 and TB10.4 3–11 peptides for in vitro stimulation of lymphocytes from infected mice and the frequency of CD4/10.4 or CD8/10.4 T cells out of all T cells following 6 hour stimulation with antigen was analyzed by flow cytometry (calculation shown in materials and methods) (Fig. 1). The CFU levels in the lung showed an increase up to week 4–5 post infection where after the CFU levels did not change significantly throughout the course of infection (Fig. 2). In terms of the CD4/10.4 and CD8/10.4 cells the dynamic development of priming, expansion and contraction of these cells followed different patterns (Fig. 3). In the spleen and blood, both CD4/10.4 and CD8/10.4 cells could be detected, but the frequencies out of all lymphocytes, were around 1%, and not as high as seen in the lung. In the lungs, CD4/10.4 cells expanded rapidly and reached around 3% of all T cells (5.5% of all CD4 T cells, data not shown) 4–6 weeks after infection, which represented a significant increase in CD4/10.4 cell numbers compared to week 0 post infection (student's T test, p<0.05). The percentage then declined to approximately 1.5% of all T cells and it stayed at that level for the rest of the experiment (until week 43) (Fig. 3A). In contrast, CD8/10.4 displayed a delayed kinetic and this T cell population reached its initial peak as late as week 15 post-infection where 6.5% of all T cells (and 18% of all CD8 T cells, data not shown) recognized this epitope. This was followed by a contraction from week 22–27 (to around 2%) after which CD8/10.4 cells again stabilized at a level between 5–6% of all T cells (and 18% of all CD8 T cells, data not shown) (Fig. 3B). The accelerated expansion of CD4/10.4 was confirmed by ELISPOT at week 6 where TB10.4 74–88 stimulation induced up to 7 fold more spot forming (CD4/10.4) cells than stimulation with the CD8 epitope TB10.4 3–11 (data not shown). Staining the cells with the H-2Kb/TB10.4 pentamer that specifically identified CD8/10.4 cells, confirmed the kinetic pattern observed after peptide stimulation (Fig. 3C) and the similar percentages obtained by pentamer and IFN-γ staining indicated that this CD8 T cell population expressed IFN-γ throughout the observation period. Thus, following infection the frequency of CD4 and CD8 T cells specific for epitopes both encoded within the TB10.4 molecule, followed distinct patterns.


Distinct differences in the expansion and phenotype of TB10.4 specific CD8 and CD4 T cells after infection with Mycobacterium tuberculosis.

Hoang TT, Nansen A, Roy S, Billeskov R, Aagaard C, Elvang T, Dietrich J, Andersen P - PLoS ONE (2009)

The bacterial load in the lungs throughout infection shown as Log10 CFU.Mice were challenged by the aerosol route with virulent M. tuberculosis. At the indicated timepoints 3–6 mice were killed and the bacterial burden (CFU) was measured in the lung.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2691482&req=5

pone-0005928-g002: The bacterial load in the lungs throughout infection shown as Log10 CFU.Mice were challenged by the aerosol route with virulent M. tuberculosis. At the indicated timepoints 3–6 mice were killed and the bacterial burden (CFU) was measured in the lung.
Mentions: Anti-TB10.4 3–11 CD8 T cells and anti-TB10.4 74–88 CD4 T cells (hereafter called CD8/10.4 and CD4/10.4 cells) represent a significant proportion of the T cells induced by infection with M.tb [3]. To study and compare the dynamic development of these T cells, CB6F1 (BALB/c×C57BL/6) mice were infected by the aerosol route with M.tb Erdman and the T cell immune response against TB10.4 74–88 and TB10.4 3–11 was analyzed at different time points following infection. Epitope recognition was assessed using TB10.4 74–88 and TB10.4 3–11 peptides for in vitro stimulation of lymphocytes from infected mice and the frequency of CD4/10.4 or CD8/10.4 T cells out of all T cells following 6 hour stimulation with antigen was analyzed by flow cytometry (calculation shown in materials and methods) (Fig. 1). The CFU levels in the lung showed an increase up to week 4–5 post infection where after the CFU levels did not change significantly throughout the course of infection (Fig. 2). In terms of the CD4/10.4 and CD8/10.4 cells the dynamic development of priming, expansion and contraction of these cells followed different patterns (Fig. 3). In the spleen and blood, both CD4/10.4 and CD8/10.4 cells could be detected, but the frequencies out of all lymphocytes, were around 1%, and not as high as seen in the lung. In the lungs, CD4/10.4 cells expanded rapidly and reached around 3% of all T cells (5.5% of all CD4 T cells, data not shown) 4–6 weeks after infection, which represented a significant increase in CD4/10.4 cell numbers compared to week 0 post infection (student's T test, p<0.05). The percentage then declined to approximately 1.5% of all T cells and it stayed at that level for the rest of the experiment (until week 43) (Fig. 3A). In contrast, CD8/10.4 displayed a delayed kinetic and this T cell population reached its initial peak as late as week 15 post-infection where 6.5% of all T cells (and 18% of all CD8 T cells, data not shown) recognized this epitope. This was followed by a contraction from week 22–27 (to around 2%) after which CD8/10.4 cells again stabilized at a level between 5–6% of all T cells (and 18% of all CD8 T cells, data not shown) (Fig. 3B). The accelerated expansion of CD4/10.4 was confirmed by ELISPOT at week 6 where TB10.4 74–88 stimulation induced up to 7 fold more spot forming (CD4/10.4) cells than stimulation with the CD8 epitope TB10.4 3–11 (data not shown). Staining the cells with the H-2Kb/TB10.4 pentamer that specifically identified CD8/10.4 cells, confirmed the kinetic pattern observed after peptide stimulation (Fig. 3C) and the similar percentages obtained by pentamer and IFN-γ staining indicated that this CD8 T cell population expressed IFN-γ throughout the observation period. Thus, following infection the frequency of CD4 and CD8 T cells specific for epitopes both encoded within the TB10.4 molecule, followed distinct patterns.

Bottom Line: Recently we and others have identified CD8 and CD4 T cell epitopes within the highly expressed M. tuberculosis protein TB10.4.We focused on T cells directed to two epitopes in TB10.4; the MHC class I restricted epitope TB10.4 (3-11) (CD8/10.4 T cells) and the MHC class II restricted epitope TB10.4 (74-88) (CD4/10.4 T cells).In addition, the observed strong expansion of CD8 T cells in the late stages of infection could have implications for the development of post exposure vaccines against latent TB.

View Article: PubMed Central - PubMed

Affiliation: Department of Infectious Disease Immunology, Statens Serum Institut, Copenhagen, Denmark.

ABSTRACT

Background: Recently we and others have identified CD8 and CD4 T cell epitopes within the highly expressed M. tuberculosis protein TB10.4. This has enabled, for the first time, a comparative study of the dynamics and function of CD4 and CD8 T cells specific for epitopes within the same protein in various stages of TB infection.

Methods and findings: We focused on T cells directed to two epitopes in TB10.4; the MHC class I restricted epitope TB10.4 (3-11) (CD8/10.4 T cells) and the MHC class II restricted epitope TB10.4 (74-88) (CD4/10.4 T cells). CD4/10.4 and CD8/10.4 T cells displayed marked differences in terms of expansion and contraction in a mouse TB model. CD4/10.4 T cells dominated in the early phase of infection whereas CD8/10.4 T cells were expanded after week 16 and reached 5-8 fold higher numbers in the late phase of infection. In the early phase of infection both CD4/10.4 and CD8/10.4 T cells were characterized by 20-25% polyfunctional cells (IL-2(+), IFN-gamma(+), TNF-alpha(+)), but whereas the majority of CD4/10.4 T cells were maintained as polyfunctional T cells throughout infection, CD8/10.4 T cells differentiated almost exclusively into effector cells (IFN-gamma(+), TNF-alpha(+)). Both CD4/10.4 and CD8/10.4 T cells exhibited cytotoxicity in vivo in the early phase of infection, but whereas CD4/10.4 cell mediated cytotoxicity waned during the infection, CD8/10.4 T cells exhibited increasing cytotoxic potential throughout the infection.

Conclusions/significance: Our results show that CD4 and CD8 T cells directed to epitopes in the same antigen differ both in their kinetics and functional characteristics throughout an infection with M. tuberculosis. In addition, the observed strong expansion of CD8 T cells in the late stages of infection could have implications for the development of post exposure vaccines against latent TB.

Show MeSH
Related in: MedlinePlus