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Oral Wild-Type Salmonella Typhi Challenge Induces Activation of Circulating Monocytes and Dendritic Cells in Individuals Who Develop Typhoid Disease.

Toapanta FR, Bernal PJ, Fresnay S, Darton TC, Jones C, Waddington CS, Blohmke CJ, Dougan G, Angus B, Levine MM, Pollard AJ, Sztein MB - PLoS Negl Trop Dis (2015)

Bottom Line: Typhi bacteremia) endpoints.Typhi binding 48 h and 96 h post-TD, and only Erk1/2 phosphorylation.Typhi.

View Article: PubMed Central - PubMed

Affiliation: Center for Vaccine Development, University of Maryland School of Medicine, Baltimore, Maryland, United States of America; Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, United States of America.

ABSTRACT
A new human oral challenge model with wild-type Salmonella Typhi (S. Typhi) was recently developed. In this model, ingestion of 104 CFU of Salmonella resulted in 65% of subjects developing typhoid fever (referred here as typhoid diagnosis -TD-) 5-10 days post-challenge. TD criteria included meeting clinical (oral temperature ≥38°C for ≥12 h) and/or microbiological (S. Typhi bacteremia) endpoints. One of the first lines of defense against pathogens are the cells of the innate immune system (e.g., monocytes, dendritic cells -DCs-). Various changes in circulating monocytes and DCs have been described in the murine S. Typhimurium model; however, whether similar changes are present in humans remains to be explored. To address these questions, a subset of volunteers (5 TD and 3 who did not develop typhoid despite oral challenge -NoTD-) were evaluated for changes in circulating monocytes and DCs. Expression of CD38 and CD40 were upregulated in monocytes and DCs in TD volunteers during the disease days (TD-0h to TD-96h). Moreover, integrin α4β7, a gut homing molecule, was upregulated on monocytes but not DCs. CD21 upregulation was only identified in DCs. These changes were not observed among NoTD volunteers despite the same oral challenge. Moreover, monocytes and DCs from NoTD volunteers showed increased binding to S. Typhi one day after challenge. These monocytes showed phosphorylation of p38MAPK, NFkB and Erk1/2 upon stimulation with S. Typhi-LPS-QDot micelles. In contrast, monocytes from TD volunteers showed only a moderate increase in S. Typhi binding 48 h and 96 h post-TD, and only Erk1/2 phosphorylation. This is the first study to describe different activation and migration profiles, as well as differential signaling patterns, in monocytes and DCs which relate directly to the clinical outcome following oral challenge with wild type S. Typhi.

No MeSH data available.


Related in: MedlinePlus

Gating strategy.Stained PBMC were gated on cells with high FSC and SSC characteristics avoiding the lymphocyte region (A), doublets/multiplets were eliminated (B) and subsequently live cells were gated based on expression of CD14 (C). The viability channel (live/dead-L/D-) also contained CD3; therefore cells selected were also CD3- (C). CD14+ cells also expressed CD16 and HLA-DR. Therefore, these cells were classified as monocytes (D). All CD3-CD14- expressed HLA-DR and a percentage of these cells expressed either CD11c or CD123 but lacked CD19, CD66b, CD16 and CD56; therefore, these cells were classified as dendritic cells (DC) (E) (see also S4 Fig). The identity of these cells as DCs was confirmed using PBMC from healthy volunteers stained with monoclonal antibodies to BDCA-1, BDCA-2 and BDCA-3 (S4 Fig). Plots shown are from a representative volunteer. Gray histograms represent fluorescent minus one (FMO) samples.
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pntd.0003837.g001: Gating strategy.Stained PBMC were gated on cells with high FSC and SSC characteristics avoiding the lymphocyte region (A), doublets/multiplets were eliminated (B) and subsequently live cells were gated based on expression of CD14 (C). The viability channel (live/dead-L/D-) also contained CD3; therefore cells selected were also CD3- (C). CD14+ cells also expressed CD16 and HLA-DR. Therefore, these cells were classified as monocytes (D). All CD3-CD14- expressed HLA-DR and a percentage of these cells expressed either CD11c or CD123 but lacked CD19, CD66b, CD16 and CD56; therefore, these cells were classified as dendritic cells (DC) (E) (see also S4 Fig). The identity of these cells as DCs was confirmed using PBMC from healthy volunteers stained with monoclonal antibodies to BDCA-1, BDCA-2 and BDCA-3 (S4 Fig). Plots shown are from a representative volunteer. Gray histograms represent fluorescent minus one (FMO) samples.

Mentions: Monocytes (CD14+ CD16+ HLA-DR+ CD56- CD66b- CD3- CD19-) were identified in the non-lymphocytic region of the FSC/SSC bi-exponential plots (Fig 1). No significant differences were observed in the percentages of monocytes recorded before S. Typhi challenge between participants who developed TD and those who did not (NoTD) upon challenge with wt S. Typhi (Fig 2A). We next assessed changes in the percentage of monocytes after challenge. No changes were observed in TD volunteers until 2 days after typhoid diagnosis (TD-48h), when a small reduction in the percentage of these cells was noted (Fig 2B). The reductions in the percentages of monocytes appeared to persist for several days; however, the differences compared to pre-challenge were not statistically significant. The frequency of these cells began to return to baseline levels 18 days after diagnosis. In contrast, no changes in the frequency of monocytes following challenge were recorded in NoTD volunteers (Fig 2C). Interestingly, the number of circulating monocytes measured in whole blood (whole blood cell counts-WCC-) [11] did not show significant changes compared to pre-challenge levels in either TD or NoTD participants, although a small decrease was observed in TD participants starting 8 days after diagnosis (S1A and S1B Fig).


Oral Wild-Type Salmonella Typhi Challenge Induces Activation of Circulating Monocytes and Dendritic Cells in Individuals Who Develop Typhoid Disease.

Toapanta FR, Bernal PJ, Fresnay S, Darton TC, Jones C, Waddington CS, Blohmke CJ, Dougan G, Angus B, Levine MM, Pollard AJ, Sztein MB - PLoS Negl Trop Dis (2015)

Gating strategy.Stained PBMC were gated on cells with high FSC and SSC characteristics avoiding the lymphocyte region (A), doublets/multiplets were eliminated (B) and subsequently live cells were gated based on expression of CD14 (C). The viability channel (live/dead-L/D-) also contained CD3; therefore cells selected were also CD3- (C). CD14+ cells also expressed CD16 and HLA-DR. Therefore, these cells were classified as monocytes (D). All CD3-CD14- expressed HLA-DR and a percentage of these cells expressed either CD11c or CD123 but lacked CD19, CD66b, CD16 and CD56; therefore, these cells were classified as dendritic cells (DC) (E) (see also S4 Fig). The identity of these cells as DCs was confirmed using PBMC from healthy volunteers stained with monoclonal antibodies to BDCA-1, BDCA-2 and BDCA-3 (S4 Fig). Plots shown are from a representative volunteer. Gray histograms represent fluorescent minus one (FMO) samples.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4465829&req=5

pntd.0003837.g001: Gating strategy.Stained PBMC were gated on cells with high FSC and SSC characteristics avoiding the lymphocyte region (A), doublets/multiplets were eliminated (B) and subsequently live cells were gated based on expression of CD14 (C). The viability channel (live/dead-L/D-) also contained CD3; therefore cells selected were also CD3- (C). CD14+ cells also expressed CD16 and HLA-DR. Therefore, these cells were classified as monocytes (D). All CD3-CD14- expressed HLA-DR and a percentage of these cells expressed either CD11c or CD123 but lacked CD19, CD66b, CD16 and CD56; therefore, these cells were classified as dendritic cells (DC) (E) (see also S4 Fig). The identity of these cells as DCs was confirmed using PBMC from healthy volunteers stained with monoclonal antibodies to BDCA-1, BDCA-2 and BDCA-3 (S4 Fig). Plots shown are from a representative volunteer. Gray histograms represent fluorescent minus one (FMO) samples.
Mentions: Monocytes (CD14+ CD16+ HLA-DR+ CD56- CD66b- CD3- CD19-) were identified in the non-lymphocytic region of the FSC/SSC bi-exponential plots (Fig 1). No significant differences were observed in the percentages of monocytes recorded before S. Typhi challenge between participants who developed TD and those who did not (NoTD) upon challenge with wt S. Typhi (Fig 2A). We next assessed changes in the percentage of monocytes after challenge. No changes were observed in TD volunteers until 2 days after typhoid diagnosis (TD-48h), when a small reduction in the percentage of these cells was noted (Fig 2B). The reductions in the percentages of monocytes appeared to persist for several days; however, the differences compared to pre-challenge were not statistically significant. The frequency of these cells began to return to baseline levels 18 days after diagnosis. In contrast, no changes in the frequency of monocytes following challenge were recorded in NoTD volunteers (Fig 2C). Interestingly, the number of circulating monocytes measured in whole blood (whole blood cell counts-WCC-) [11] did not show significant changes compared to pre-challenge levels in either TD or NoTD participants, although a small decrease was observed in TD participants starting 8 days after diagnosis (S1A and S1B Fig).

Bottom Line: Typhi bacteremia) endpoints.Typhi binding 48 h and 96 h post-TD, and only Erk1/2 phosphorylation.Typhi.

View Article: PubMed Central - PubMed

Affiliation: Center for Vaccine Development, University of Maryland School of Medicine, Baltimore, Maryland, United States of America; Department of Medicine, University of Maryland School of Medicine, Baltimore, Maryland, United States of America.

ABSTRACT
A new human oral challenge model with wild-type Salmonella Typhi (S. Typhi) was recently developed. In this model, ingestion of 104 CFU of Salmonella resulted in 65% of subjects developing typhoid fever (referred here as typhoid diagnosis -TD-) 5-10 days post-challenge. TD criteria included meeting clinical (oral temperature ≥38°C for ≥12 h) and/or microbiological (S. Typhi bacteremia) endpoints. One of the first lines of defense against pathogens are the cells of the innate immune system (e.g., monocytes, dendritic cells -DCs-). Various changes in circulating monocytes and DCs have been described in the murine S. Typhimurium model; however, whether similar changes are present in humans remains to be explored. To address these questions, a subset of volunteers (5 TD and 3 who did not develop typhoid despite oral challenge -NoTD-) were evaluated for changes in circulating monocytes and DCs. Expression of CD38 and CD40 were upregulated in monocytes and DCs in TD volunteers during the disease days (TD-0h to TD-96h). Moreover, integrin α4β7, a gut homing molecule, was upregulated on monocytes but not DCs. CD21 upregulation was only identified in DCs. These changes were not observed among NoTD volunteers despite the same oral challenge. Moreover, monocytes and DCs from NoTD volunteers showed increased binding to S. Typhi one day after challenge. These monocytes showed phosphorylation of p38MAPK, NFkB and Erk1/2 upon stimulation with S. Typhi-LPS-QDot micelles. In contrast, monocytes from TD volunteers showed only a moderate increase in S. Typhi binding 48 h and 96 h post-TD, and only Erk1/2 phosphorylation. This is the first study to describe different activation and migration profiles, as well as differential signaling patterns, in monocytes and DCs which relate directly to the clinical outcome following oral challenge with wild type S. Typhi.

No MeSH data available.


Related in: MedlinePlus