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BCG Skin Infection Triggers IL-1R-MyD88-Dependent Migration of EpCAMlow CD11bhigh Skin Dendritic cells to Draining Lymph Node During CD4+ T-Cell Priming.

Bollampalli VP, Harumi Yamashiro L, Feng X, Bierschenk D, Gao Y, Blom H, Henriques-Normark B, Nylén S, Rothfuchs AG - PLoS Pathog. (2015)

Bottom Line: Migratory skin DCs distributed to the T-cell area of the LN, co-localized with BCG and were found in close apposition to antigen-specific CD4+ T cells.Consequently, blockade of skin DC traffic into DLN dramatically reduced mycobacterial entry into DLN and muted T-cell priming.In addition, we found using DC adoptive transfers that the requirement for MyD88 in BCG-triggered migration was not restricted to the migrating DC itself and that hematopoietic expression of MyD88 was needed in part for full-fledged migration.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden.

ABSTRACT
The transport of antigen from the periphery to the draining lymph node (DLN) is critical for T-cell priming but remains poorly studied during infection with Mycobacterium bovis Bacille Calmette-Guérin (BCG). To address this we employed a mouse model to track the traffic of Dendritic cells (DCs) and mycobacteria from the BCG inoculation site in the skin to the DLN. Detection of BCG in the DLN was concomitant with the priming of antigen-specific CD4+ T cells at that site. We found EpCAMlow CD11bhigh migratory skin DCs to be mobilized during the transport of BCG to the DLN. Migratory skin DCs distributed to the T-cell area of the LN, co-localized with BCG and were found in close apposition to antigen-specific CD4+ T cells. Consequently, blockade of skin DC traffic into DLN dramatically reduced mycobacterial entry into DLN and muted T-cell priming. Interestingly, DC and mycobacterial entry into the DLN was dependent on IL-1R-I, MyD88, TNFR-I and IL-12p40. In addition, we found using DC adoptive transfers that the requirement for MyD88 in BCG-triggered migration was not restricted to the migrating DC itself and that hematopoietic expression of MyD88 was needed in part for full-fledged migration. Our observations thus identify a population of DCs that contribute towards the priming of CD4+ T cells to BCG infection by transporting bacilli into the DLN in an IL-1R-MyD88-dependent manner and reveal both DC-intrinsic and -extrinsic requirements for MyD88 in DC migration.

No MeSH data available.


Related in: MedlinePlus

PTx mutes P25 TCRTg-cell priming by inhibiting skin DC migration and BCG transport to DLN.(A) Total number of CFSE+ MHC-IIhigh CD11c+/low skin DCs in BCG-draining pLN from animals treated in the footpad with PTx. WT mice received an injection of 1 μg PTx (Sigma) or PBS in the footpad. Animals were inoculated 4hrs later with BCG in the same footpad. CFSE was injected into the same footpads 24hrs before sacrifice. Three days after BCG, pLNs were isolated and frequency of migrating skin DCs analyzed by flow cytometry. Dashed line depicts average number of CFSE+ skin DCs in uninfected, PBS-injected controls. (B) CFUs of BCG in pLN at the indicated time points after infection were determined on 7H11 agar from WT mice treated with PTx or PBS as in (A). (C) Naïve P25 TCRTg cells were CFSE-labeled and transferred into congenic CD45.1+ recipients. Twenty-four hrs later, animals were treated with PTx as in (A) and inoculated with BCG in the same footpads. Expansion of P25 TCRTg cells was determined 3 days later by flow cytometry. Dashed line depicts average frequency of P25 TCRTg cells obtained from uninfected, PBS-injected controls. For each experiment, at least 5 mice were used for BCG-infected groups and 3 for PBS controls. Bars indicate standard error of the mean. One of two independent experiments shown. *Denotes statistically significant difference in BCG-infected, PTx-treated group compared to BCG-infected, PBS-treated animals.
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ppat.1005206.g005: PTx mutes P25 TCRTg-cell priming by inhibiting skin DC migration and BCG transport to DLN.(A) Total number of CFSE+ MHC-IIhigh CD11c+/low skin DCs in BCG-draining pLN from animals treated in the footpad with PTx. WT mice received an injection of 1 μg PTx (Sigma) or PBS in the footpad. Animals were inoculated 4hrs later with BCG in the same footpad. CFSE was injected into the same footpads 24hrs before sacrifice. Three days after BCG, pLNs were isolated and frequency of migrating skin DCs analyzed by flow cytometry. Dashed line depicts average number of CFSE+ skin DCs in uninfected, PBS-injected controls. (B) CFUs of BCG in pLN at the indicated time points after infection were determined on 7H11 agar from WT mice treated with PTx or PBS as in (A). (C) Naïve P25 TCRTg cells were CFSE-labeled and transferred into congenic CD45.1+ recipients. Twenty-four hrs later, animals were treated with PTx as in (A) and inoculated with BCG in the same footpads. Expansion of P25 TCRTg cells was determined 3 days later by flow cytometry. Dashed line depicts average frequency of P25 TCRTg cells obtained from uninfected, PBS-injected controls. For each experiment, at least 5 mice were used for BCG-infected groups and 3 for PBS controls. Bars indicate standard error of the mean. One of two independent experiments shown. *Denotes statistically significant difference in BCG-infected, PTx-treated group compared to BCG-infected, PBS-treated animals.

Mentions: To further study the contribution of migratory skin DCs in T-cell priming, Pertussis toxin (PTx) was injected in the footpad of mice prior to BCG infection as a means of blocking DC migration [20]. Such treatment with PTx lead to a total ablation of BCG-triggered skin DC mobilization to DLN (Fig 5A) and a dramatic reduction in the number of BCG Colony-forming units (CFUs) in the DLN (Fig 5B). Importantly, PTx treatment muted the early expansion of naïve P25 TCRTg cells in the DLN (Fig 5C). PTx did not seem to have a major, direct impact on P25 TCRTg cells as the activation profile of this population in PTx-treated, BCG-infected animals was similar to that of infected controls (S4A and S4B Fig). Similarly, injection of PTx 3 days after BCG infection, when skin DCs and mycobacteria had already reached the DLN, did not alter the activation of naïve P25 TCRTg cells transferred at the time of PTx treatment (S4C Fig). A direct effect of PTx on BCG growth was also ruled out since addition of PTx to BCG cultures did not affect mycobacterial growth in 7H11 agar (S4D Fig). Thus, upon localized injection in the footpad, PTx seems mainly to inhibit skin DC migration to DLN and consequently, the entry of BCG to the DLN. These observations support a role for migratory skin DCs in channeling BCG to the DLN and in doing so, the priming of CD4+ T cells is unleashed therein.


BCG Skin Infection Triggers IL-1R-MyD88-Dependent Migration of EpCAMlow CD11bhigh Skin Dendritic cells to Draining Lymph Node During CD4+ T-Cell Priming.

Bollampalli VP, Harumi Yamashiro L, Feng X, Bierschenk D, Gao Y, Blom H, Henriques-Normark B, Nylén S, Rothfuchs AG - PLoS Pathog. (2015)

PTx mutes P25 TCRTg-cell priming by inhibiting skin DC migration and BCG transport to DLN.(A) Total number of CFSE+ MHC-IIhigh CD11c+/low skin DCs in BCG-draining pLN from animals treated in the footpad with PTx. WT mice received an injection of 1 μg PTx (Sigma) or PBS in the footpad. Animals were inoculated 4hrs later with BCG in the same footpad. CFSE was injected into the same footpads 24hrs before sacrifice. Three days after BCG, pLNs were isolated and frequency of migrating skin DCs analyzed by flow cytometry. Dashed line depicts average number of CFSE+ skin DCs in uninfected, PBS-injected controls. (B) CFUs of BCG in pLN at the indicated time points after infection were determined on 7H11 agar from WT mice treated with PTx or PBS as in (A). (C) Naïve P25 TCRTg cells were CFSE-labeled and transferred into congenic CD45.1+ recipients. Twenty-four hrs later, animals were treated with PTx as in (A) and inoculated with BCG in the same footpads. Expansion of P25 TCRTg cells was determined 3 days later by flow cytometry. Dashed line depicts average frequency of P25 TCRTg cells obtained from uninfected, PBS-injected controls. For each experiment, at least 5 mice were used for BCG-infected groups and 3 for PBS controls. Bars indicate standard error of the mean. One of two independent experiments shown. *Denotes statistically significant difference in BCG-infected, PTx-treated group compared to BCG-infected, PBS-treated animals.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4594926&req=5

ppat.1005206.g005: PTx mutes P25 TCRTg-cell priming by inhibiting skin DC migration and BCG transport to DLN.(A) Total number of CFSE+ MHC-IIhigh CD11c+/low skin DCs in BCG-draining pLN from animals treated in the footpad with PTx. WT mice received an injection of 1 μg PTx (Sigma) or PBS in the footpad. Animals were inoculated 4hrs later with BCG in the same footpad. CFSE was injected into the same footpads 24hrs before sacrifice. Three days after BCG, pLNs were isolated and frequency of migrating skin DCs analyzed by flow cytometry. Dashed line depicts average number of CFSE+ skin DCs in uninfected, PBS-injected controls. (B) CFUs of BCG in pLN at the indicated time points after infection were determined on 7H11 agar from WT mice treated with PTx or PBS as in (A). (C) Naïve P25 TCRTg cells were CFSE-labeled and transferred into congenic CD45.1+ recipients. Twenty-four hrs later, animals were treated with PTx as in (A) and inoculated with BCG in the same footpads. Expansion of P25 TCRTg cells was determined 3 days later by flow cytometry. Dashed line depicts average frequency of P25 TCRTg cells obtained from uninfected, PBS-injected controls. For each experiment, at least 5 mice were used for BCG-infected groups and 3 for PBS controls. Bars indicate standard error of the mean. One of two independent experiments shown. *Denotes statistically significant difference in BCG-infected, PTx-treated group compared to BCG-infected, PBS-treated animals.
Mentions: To further study the contribution of migratory skin DCs in T-cell priming, Pertussis toxin (PTx) was injected in the footpad of mice prior to BCG infection as a means of blocking DC migration [20]. Such treatment with PTx lead to a total ablation of BCG-triggered skin DC mobilization to DLN (Fig 5A) and a dramatic reduction in the number of BCG Colony-forming units (CFUs) in the DLN (Fig 5B). Importantly, PTx treatment muted the early expansion of naïve P25 TCRTg cells in the DLN (Fig 5C). PTx did not seem to have a major, direct impact on P25 TCRTg cells as the activation profile of this population in PTx-treated, BCG-infected animals was similar to that of infected controls (S4A and S4B Fig). Similarly, injection of PTx 3 days after BCG infection, when skin DCs and mycobacteria had already reached the DLN, did not alter the activation of naïve P25 TCRTg cells transferred at the time of PTx treatment (S4C Fig). A direct effect of PTx on BCG growth was also ruled out since addition of PTx to BCG cultures did not affect mycobacterial growth in 7H11 agar (S4D Fig). Thus, upon localized injection in the footpad, PTx seems mainly to inhibit skin DC migration to DLN and consequently, the entry of BCG to the DLN. These observations support a role for migratory skin DCs in channeling BCG to the DLN and in doing so, the priming of CD4+ T cells is unleashed therein.

Bottom Line: Migratory skin DCs distributed to the T-cell area of the LN, co-localized with BCG and were found in close apposition to antigen-specific CD4+ T cells.Consequently, blockade of skin DC traffic into DLN dramatically reduced mycobacterial entry into DLN and muted T-cell priming.In addition, we found using DC adoptive transfers that the requirement for MyD88 in BCG-triggered migration was not restricted to the migrating DC itself and that hematopoietic expression of MyD88 was needed in part for full-fledged migration.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden.

ABSTRACT
The transport of antigen from the periphery to the draining lymph node (DLN) is critical for T-cell priming but remains poorly studied during infection with Mycobacterium bovis Bacille Calmette-Guérin (BCG). To address this we employed a mouse model to track the traffic of Dendritic cells (DCs) and mycobacteria from the BCG inoculation site in the skin to the DLN. Detection of BCG in the DLN was concomitant with the priming of antigen-specific CD4+ T cells at that site. We found EpCAMlow CD11bhigh migratory skin DCs to be mobilized during the transport of BCG to the DLN. Migratory skin DCs distributed to the T-cell area of the LN, co-localized with BCG and were found in close apposition to antigen-specific CD4+ T cells. Consequently, blockade of skin DC traffic into DLN dramatically reduced mycobacterial entry into DLN and muted T-cell priming. Interestingly, DC and mycobacterial entry into the DLN was dependent on IL-1R-I, MyD88, TNFR-I and IL-12p40. In addition, we found using DC adoptive transfers that the requirement for MyD88 in BCG-triggered migration was not restricted to the migrating DC itself and that hematopoietic expression of MyD88 was needed in part for full-fledged migration. Our observations thus identify a population of DCs that contribute towards the priming of CD4+ T cells to BCG infection by transporting bacilli into the DLN in an IL-1R-MyD88-dependent manner and reveal both DC-intrinsic and -extrinsic requirements for MyD88 in DC migration.

No MeSH data available.


Related in: MedlinePlus