Limits...
Priming of Salmonella enterica serovar typhi-specific CD8(+) T cells by suicide dendritic cell cross-presentation in humans.

Salerno-Goncalves R, Sztein MB - PLoS ONE (2009)

Bottom Line: Typhi-infected human cells and release high levels of IFN-gamma and IL-12p70, leading to the subsequent presentation of bacterial antigens and triggering the induction of memory T cells, mostly CD3(+)CD8(+)CD45RA(-)CD62L(-) effector/memory T cells.This study is the first to demonstrate the effect of S.Typhi.

View Article: PubMed Central - PubMed

Affiliation: Center for Vaccine Development, University of Maryland School of Medicine, Baltimore, MD, USA. rmezghan@medicine.umaryland.edu

ABSTRACT

Background: The emergence of antibiotic-resistant strains of Salmonella enterica serovar Typhi (S. Typhi), the etiologic agent of typhoid fever, has aggravated an already important public health problem and added new urgency to the development of more effective typhoid vaccines. To this end it is critical to better understand the induction of immunity to S. Typhi. CD8(+) T cells are likely to play an important role in host defense against S. Typhi by several effector mechanisms, including killing of infected cells and IFN-gamma secretion. However, how S. Typhi regulates the development of specific CD8(+) responses in humans remains unclear. Recent studies in mice have shown that dendritic cells (DC) can either directly (upon uptake and processing of Salmonella) or indirectly (by bystander mechanisms) elicit Salmonella-specific CD8(+) T cells.

Methodology/principal findings: We report here that upon infection with live S. Typhi, human DC produced high levels of pro-inflammatory cytokines IL-6, IL-8 and TNF-alpha, but low levels of IL-12 p70 and IFN-gamma. In contrast, DC co-cultured with S. Typhi-infected cells, through suicide cross-presentation, uptake S. Typhi-infected human cells and release high levels of IFN-gamma and IL-12p70, leading to the subsequent presentation of bacterial antigens and triggering the induction of memory T cells, mostly CD3(+)CD8(+)CD45RA(-)CD62L(-) effector/memory T cells.

Conclusions/significance: This study is the first to demonstrate the effect of S. Typhi on human DC maturation and on their ability to prime CD8(+) cells and highlights the significance of these phenomena in eliciting adaptive immunity to S. Typhi.

Show MeSH

Related in: MedlinePlus

Induction of IFN-γ- and TNF-α-secreting CD8+ T cell subpopulations upon DC priming.PBMC from volunteer CVD4000#65 were co-cultured with DC alone (media), or pre-mixed with live or heat-killed S. Typhi at a MOI of 10∶1, or uninfected or S. Typhi-infected cells at a 1∶4 blast∶DC ratio. After 20 hours of incubation, cells were surface stained with a combination of mAb to CD3, CD4, CD8, CD14, CD19, CD45RA, and CD62L as well as ViViD. After fixation and permeabilization, cells were intracellularly stained for IFN-γ and TNF-α and analyzed by multichromatic flow cytometry. Lymphocytes gated as described in Figure 5 were followed by additional gating on CD3, CD4 and CD8, as well as CDR45RA versus CD62L to analyze TEM and TEMRA cell subsets. Numbers correspond to the percentage of positive cells in the indicated regions in each histogram. These results are representative of 1 of 4 volunteers with similar results.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2691582&req=5

pone-0005879-g009: Induction of IFN-γ- and TNF-α-secreting CD8+ T cell subpopulations upon DC priming.PBMC from volunteer CVD4000#65 were co-cultured with DC alone (media), or pre-mixed with live or heat-killed S. Typhi at a MOI of 10∶1, or uninfected or S. Typhi-infected cells at a 1∶4 blast∶DC ratio. After 20 hours of incubation, cells were surface stained with a combination of mAb to CD3, CD4, CD8, CD14, CD19, CD45RA, and CD62L as well as ViViD. After fixation and permeabilization, cells were intracellularly stained for IFN-γ and TNF-α and analyzed by multichromatic flow cytometry. Lymphocytes gated as described in Figure 5 were followed by additional gating on CD3, CD4 and CD8, as well as CDR45RA versus CD62L to analyze TEM and TEMRA cell subsets. Numbers correspond to the percentage of positive cells in the indicated regions in each histogram. These results are representative of 1 of 4 volunteers with similar results.

Mentions: We next investigated which CD8 cell populations within PBMC preferentially produced IFN-γ and TNF-α cytokines under the different stimulatory conditions described above. We evaluated the induction of cytokine production in the various memory CD8+ T cell subsets, defined as follows: central memory T cells (TCM, CD45RA-CD62L+), effector memory T cells (TEM, CD45RA−CD62L−) and naïve T cells (Tn, CD45RA+CD62L+) [14]. We observed that the majority of CD8+ T cells elicited were composed of classical TEM (Figs. 8 and 9). Minor increases were also observed in TEMRA. No significant increases were observed in TCM and T naïve subsets (data not shown).


Priming of Salmonella enterica serovar typhi-specific CD8(+) T cells by suicide dendritic cell cross-presentation in humans.

Salerno-Goncalves R, Sztein MB - PLoS ONE (2009)

Induction of IFN-γ- and TNF-α-secreting CD8+ T cell subpopulations upon DC priming.PBMC from volunteer CVD4000#65 were co-cultured with DC alone (media), or pre-mixed with live or heat-killed S. Typhi at a MOI of 10∶1, or uninfected or S. Typhi-infected cells at a 1∶4 blast∶DC ratio. After 20 hours of incubation, cells were surface stained with a combination of mAb to CD3, CD4, CD8, CD14, CD19, CD45RA, and CD62L as well as ViViD. After fixation and permeabilization, cells were intracellularly stained for IFN-γ and TNF-α and analyzed by multichromatic flow cytometry. Lymphocytes gated as described in Figure 5 were followed by additional gating on CD3, CD4 and CD8, as well as CDR45RA versus CD62L to analyze TEM and TEMRA cell subsets. Numbers correspond to the percentage of positive cells in the indicated regions in each histogram. These results are representative of 1 of 4 volunteers with similar results.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2691582&req=5

pone-0005879-g009: Induction of IFN-γ- and TNF-α-secreting CD8+ T cell subpopulations upon DC priming.PBMC from volunteer CVD4000#65 were co-cultured with DC alone (media), or pre-mixed with live or heat-killed S. Typhi at a MOI of 10∶1, or uninfected or S. Typhi-infected cells at a 1∶4 blast∶DC ratio. After 20 hours of incubation, cells were surface stained with a combination of mAb to CD3, CD4, CD8, CD14, CD19, CD45RA, and CD62L as well as ViViD. After fixation and permeabilization, cells were intracellularly stained for IFN-γ and TNF-α and analyzed by multichromatic flow cytometry. Lymphocytes gated as described in Figure 5 were followed by additional gating on CD3, CD4 and CD8, as well as CDR45RA versus CD62L to analyze TEM and TEMRA cell subsets. Numbers correspond to the percentage of positive cells in the indicated regions in each histogram. These results are representative of 1 of 4 volunteers with similar results.
Mentions: We next investigated which CD8 cell populations within PBMC preferentially produced IFN-γ and TNF-α cytokines under the different stimulatory conditions described above. We evaluated the induction of cytokine production in the various memory CD8+ T cell subsets, defined as follows: central memory T cells (TCM, CD45RA-CD62L+), effector memory T cells (TEM, CD45RA−CD62L−) and naïve T cells (Tn, CD45RA+CD62L+) [14]. We observed that the majority of CD8+ T cells elicited were composed of classical TEM (Figs. 8 and 9). Minor increases were also observed in TEMRA. No significant increases were observed in TCM and T naïve subsets (data not shown).

Bottom Line: Typhi-infected human cells and release high levels of IFN-gamma and IL-12p70, leading to the subsequent presentation of bacterial antigens and triggering the induction of memory T cells, mostly CD3(+)CD8(+)CD45RA(-)CD62L(-) effector/memory T cells.This study is the first to demonstrate the effect of S.Typhi.

View Article: PubMed Central - PubMed

Affiliation: Center for Vaccine Development, University of Maryland School of Medicine, Baltimore, MD, USA. rmezghan@medicine.umaryland.edu

ABSTRACT

Background: The emergence of antibiotic-resistant strains of Salmonella enterica serovar Typhi (S. Typhi), the etiologic agent of typhoid fever, has aggravated an already important public health problem and added new urgency to the development of more effective typhoid vaccines. To this end it is critical to better understand the induction of immunity to S. Typhi. CD8(+) T cells are likely to play an important role in host defense against S. Typhi by several effector mechanisms, including killing of infected cells and IFN-gamma secretion. However, how S. Typhi regulates the development of specific CD8(+) responses in humans remains unclear. Recent studies in mice have shown that dendritic cells (DC) can either directly (upon uptake and processing of Salmonella) or indirectly (by bystander mechanisms) elicit Salmonella-specific CD8(+) T cells.

Methodology/principal findings: We report here that upon infection with live S. Typhi, human DC produced high levels of pro-inflammatory cytokines IL-6, IL-8 and TNF-alpha, but low levels of IL-12 p70 and IFN-gamma. In contrast, DC co-cultured with S. Typhi-infected cells, through suicide cross-presentation, uptake S. Typhi-infected human cells and release high levels of IFN-gamma and IL-12p70, leading to the subsequent presentation of bacterial antigens and triggering the induction of memory T cells, mostly CD3(+)CD8(+)CD45RA(-)CD62L(-) effector/memory T cells.

Conclusions/significance: This study is the first to demonstrate the effect of S. Typhi on human DC maturation and on their ability to prime CD8(+) cells and highlights the significance of these phenomena in eliciting adaptive immunity to S. Typhi.

Show MeSH
Related in: MedlinePlus