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B cells enhance antigen-specific CD4 T cell priming and prevent bacteria dissemination following Chlamydia muridarum genital tract infection.

Li LX, McSorley SJ - PLoS Pathog. (2013)

Bottom Line: While most lymph node Chlamydia-specific CD4 T cells expressed T-bet, a small percentage co-expressed Foxp3, and RORγt-expressing T cells were enriched within the reproductive tract.Local Chlamydia-specific CD4 T cell priming was markedly reduced in mice lacking B cells, and bacteria were able to disseminate to the peritoneal cavity, initiating a cellular infiltrate and ascites.However, bacterial dissemination also coincided with elevated systemic Chlamydia-specific CD4 T cell responses and resolution of primary infection.

View Article: PubMed Central - PubMed

Affiliation: Center for Comparative Medicine, Department of Anatomy, Physiology and Cell Biology, School of Veterinary Medicine, University of California, Davis, Davis, California, United States of America.

ABSTRACT
B cells can contribute to acquired immunity against intracellular bacteria, but do not usually participate in primary clearance. Here, we examined the endogenous CD4 T cell response to genital infection with Chlamydia muridarum using MHC class-II tetramers. Chlamydia-specific CD4 T cells expanded rapidly and persisted as a stable memory pool for several months after infection. While most lymph node Chlamydia-specific CD4 T cells expressed T-bet, a small percentage co-expressed Foxp3, and RORγt-expressing T cells were enriched within the reproductive tract. Local Chlamydia-specific CD4 T cell priming was markedly reduced in mice lacking B cells, and bacteria were able to disseminate to the peritoneal cavity, initiating a cellular infiltrate and ascites. However, bacterial dissemination also coincided with elevated systemic Chlamydia-specific CD4 T cell responses and resolution of primary infection. Together, these data reveal heterogeneity in pathogen-specific CD4 T cell responses within the genital tract and an unexpected requirement for B cells in regulating local T cell activation and bacterial dissemination during genital infection.

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Development of Th1, Tregs and Th17 cell subsets after C. muridarum infections.(A) and (B) C57BL/6 mice were infected with 1×105C. muridarum i.v. or i.vag.. PmpG-1303–311:I-Ab+ cells were enriched from spleen, DLNs and NDLNs 7 and 14 days post i.v. and i.vag. infections, respectively. (A) Histograms showing expression levels of T-bet and GATA3 on CD4+PmpG-1303–311:I-Ab+ cells from naïve and infected mice. (B) FACS plots showing expression of Foxp3 on CD4+ cells. (C) RORγt-GFP reporter mice were infected i.vag. with 1×105C. muridarum. Cells from spleen, DLNs and genital tract were isolated and stained with pooled RplF51–59:I-Ab, Aasf24–32:I-Ab and PmpG-1303–311:I-Ab tetramers (R.A.P.:I- Ab). CD4+CD44hi R.A.P.:I- Ab+ cells were further gated for GFP expression. RORγt-GFP− mice were used as a negative control for setting the GFP+ gates in our analysis. (D) C57BL/6 mice were infected with 1×105C. muridarum intravaginally. IFNγ and IL-17A production by CD4 T cells from DLNs and genital tract were detected by flow cytometry after ex vivo stimulation with phorbol 12-myristate 13-acetate and ionomycin in the presence of brefeldin A.
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ppat-1003707-g003: Development of Th1, Tregs and Th17 cell subsets after C. muridarum infections.(A) and (B) C57BL/6 mice were infected with 1×105C. muridarum i.v. or i.vag.. PmpG-1303–311:I-Ab+ cells were enriched from spleen, DLNs and NDLNs 7 and 14 days post i.v. and i.vag. infections, respectively. (A) Histograms showing expression levels of T-bet and GATA3 on CD4+PmpG-1303–311:I-Ab+ cells from naïve and infected mice. (B) FACS plots showing expression of Foxp3 on CD4+ cells. (C) RORγt-GFP reporter mice were infected i.vag. with 1×105C. muridarum. Cells from spleen, DLNs and genital tract were isolated and stained with pooled RplF51–59:I-Ab, Aasf24–32:I-Ab and PmpG-1303–311:I-Ab tetramers (R.A.P.:I- Ab). CD4+CD44hi R.A.P.:I- Ab+ cells were further gated for GFP expression. RORγt-GFP− mice were used as a negative control for setting the GFP+ gates in our analysis. (D) C57BL/6 mice were infected with 1×105C. muridarum intravaginally. IFNγ and IL-17A production by CD4 T cells from DLNs and genital tract were detected by flow cytometry after ex vivo stimulation with phorbol 12-myristate 13-acetate and ionomycin in the presence of brefeldin A.

Mentions: To examine CD4 T helper differentiation, the expression of lineage-specific transcription factors was examined in expanded CD44hi PmpG-1303–311:I-Ab+ CD4 T cells. Following either systemic or intravaginal infection, almost all PmpG-1-specific CD4 T cells expressed T-bet while no GATA3 expression was detected (Fig. 3A). This is consistent with previous reports that Th1 CD4 T cells are the dominant helper subset following Chlamydia infection [10], [11]. However, a distinct population of PmpG-1-specific CD4 T cells that co-expressed Foxp3 and T-bet was also detected after i.v. and i.vag. infection (Fig. 3B), suggesting that induced Chlamydia-specific Treg cells are also contained within the expanded CD4 pool. We also utilized RORγt-GFP reporter mice and combined staining with all three Chlamydia tetramers to examine the potential development of Chlamydia-specific Th17 cells after vaginal infection. While GFP-positive cells were undetectable among expanded tetramer-positive cells in the spleen or draining lymph nodes of infected mice, approximately 7% of CD4+CD44hitetramer+ T cells in infected non-lymphoid tissues expressed GFP (Fig. 3C). Furthermore, stimulation of lymphocytes purified from the genital tract confirmed the presence of T cells producing both IL-17A and IFN-γ (Fig. 3D). Thus, Chlamydia infection of the reproductive tract induces a heterogenous T helper response that comprises expanded T-bet+ Th1 cells, T-bet+Foxp3+ Tregs, and Th17 cells that are enriched in infected non-lymphoid tissues.


B cells enhance antigen-specific CD4 T cell priming and prevent bacteria dissemination following Chlamydia muridarum genital tract infection.

Li LX, McSorley SJ - PLoS Pathog. (2013)

Development of Th1, Tregs and Th17 cell subsets after C. muridarum infections.(A) and (B) C57BL/6 mice were infected with 1×105C. muridarum i.v. or i.vag.. PmpG-1303–311:I-Ab+ cells were enriched from spleen, DLNs and NDLNs 7 and 14 days post i.v. and i.vag. infections, respectively. (A) Histograms showing expression levels of T-bet and GATA3 on CD4+PmpG-1303–311:I-Ab+ cells from naïve and infected mice. (B) FACS plots showing expression of Foxp3 on CD4+ cells. (C) RORγt-GFP reporter mice were infected i.vag. with 1×105C. muridarum. Cells from spleen, DLNs and genital tract were isolated and stained with pooled RplF51–59:I-Ab, Aasf24–32:I-Ab and PmpG-1303–311:I-Ab tetramers (R.A.P.:I- Ab). CD4+CD44hi R.A.P.:I- Ab+ cells were further gated for GFP expression. RORγt-GFP− mice were used as a negative control for setting the GFP+ gates in our analysis. (D) C57BL/6 mice were infected with 1×105C. muridarum intravaginally. IFNγ and IL-17A production by CD4 T cells from DLNs and genital tract were detected by flow cytometry after ex vivo stimulation with phorbol 12-myristate 13-acetate and ionomycin in the presence of brefeldin A.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3814678&req=5

ppat-1003707-g003: Development of Th1, Tregs and Th17 cell subsets after C. muridarum infections.(A) and (B) C57BL/6 mice were infected with 1×105C. muridarum i.v. or i.vag.. PmpG-1303–311:I-Ab+ cells were enriched from spleen, DLNs and NDLNs 7 and 14 days post i.v. and i.vag. infections, respectively. (A) Histograms showing expression levels of T-bet and GATA3 on CD4+PmpG-1303–311:I-Ab+ cells from naïve and infected mice. (B) FACS plots showing expression of Foxp3 on CD4+ cells. (C) RORγt-GFP reporter mice were infected i.vag. with 1×105C. muridarum. Cells from spleen, DLNs and genital tract were isolated and stained with pooled RplF51–59:I-Ab, Aasf24–32:I-Ab and PmpG-1303–311:I-Ab tetramers (R.A.P.:I- Ab). CD4+CD44hi R.A.P.:I- Ab+ cells were further gated for GFP expression. RORγt-GFP− mice were used as a negative control for setting the GFP+ gates in our analysis. (D) C57BL/6 mice were infected with 1×105C. muridarum intravaginally. IFNγ and IL-17A production by CD4 T cells from DLNs and genital tract were detected by flow cytometry after ex vivo stimulation with phorbol 12-myristate 13-acetate and ionomycin in the presence of brefeldin A.
Mentions: To examine CD4 T helper differentiation, the expression of lineage-specific transcription factors was examined in expanded CD44hi PmpG-1303–311:I-Ab+ CD4 T cells. Following either systemic or intravaginal infection, almost all PmpG-1-specific CD4 T cells expressed T-bet while no GATA3 expression was detected (Fig. 3A). This is consistent with previous reports that Th1 CD4 T cells are the dominant helper subset following Chlamydia infection [10], [11]. However, a distinct population of PmpG-1-specific CD4 T cells that co-expressed Foxp3 and T-bet was also detected after i.v. and i.vag. infection (Fig. 3B), suggesting that induced Chlamydia-specific Treg cells are also contained within the expanded CD4 pool. We also utilized RORγt-GFP reporter mice and combined staining with all three Chlamydia tetramers to examine the potential development of Chlamydia-specific Th17 cells after vaginal infection. While GFP-positive cells were undetectable among expanded tetramer-positive cells in the spleen or draining lymph nodes of infected mice, approximately 7% of CD4+CD44hitetramer+ T cells in infected non-lymphoid tissues expressed GFP (Fig. 3C). Furthermore, stimulation of lymphocytes purified from the genital tract confirmed the presence of T cells producing both IL-17A and IFN-γ (Fig. 3D). Thus, Chlamydia infection of the reproductive tract induces a heterogenous T helper response that comprises expanded T-bet+ Th1 cells, T-bet+Foxp3+ Tregs, and Th17 cells that are enriched in infected non-lymphoid tissues.

Bottom Line: While most lymph node Chlamydia-specific CD4 T cells expressed T-bet, a small percentage co-expressed Foxp3, and RORγt-expressing T cells were enriched within the reproductive tract.Local Chlamydia-specific CD4 T cell priming was markedly reduced in mice lacking B cells, and bacteria were able to disseminate to the peritoneal cavity, initiating a cellular infiltrate and ascites.However, bacterial dissemination also coincided with elevated systemic Chlamydia-specific CD4 T cell responses and resolution of primary infection.

View Article: PubMed Central - PubMed

Affiliation: Center for Comparative Medicine, Department of Anatomy, Physiology and Cell Biology, School of Veterinary Medicine, University of California, Davis, Davis, California, United States of America.

ABSTRACT
B cells can contribute to acquired immunity against intracellular bacteria, but do not usually participate in primary clearance. Here, we examined the endogenous CD4 T cell response to genital infection with Chlamydia muridarum using MHC class-II tetramers. Chlamydia-specific CD4 T cells expanded rapidly and persisted as a stable memory pool for several months after infection. While most lymph node Chlamydia-specific CD4 T cells expressed T-bet, a small percentage co-expressed Foxp3, and RORγt-expressing T cells were enriched within the reproductive tract. Local Chlamydia-specific CD4 T cell priming was markedly reduced in mice lacking B cells, and bacteria were able to disseminate to the peritoneal cavity, initiating a cellular infiltrate and ascites. However, bacterial dissemination also coincided with elevated systemic Chlamydia-specific CD4 T cell responses and resolution of primary infection. Together, these data reveal heterogeneity in pathogen-specific CD4 T cell responses within the genital tract and an unexpected requirement for B cells in regulating local T cell activation and bacterial dissemination during genital infection.

Show MeSH
Related in: MedlinePlus