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CD40L+ CD4+ memory T cells migrate in a CD62P-dependent fashion into reactive lymph nodes and license dendritic cells for T cell priming.

Martín-Fontecha A, Baumjohann D, Guarda G, Reboldi A, Hons M, Lanzavecchia A, Sallusto F - J. Exp. Med. (2008)

Bottom Line: CD4(+) T(EM) cells were excluded from resting lymph nodes but migrated in a CD62P-dependent fashion into reactive lymph nodes that were induced to express CD62P, in a transient or sustained fashion, on high endothelial venules.Antibodies to CD62P, which blocked CD4(+) T(EM) cell migration into reactive lymph nodes, inhibited DC maturation, T cell priming, and induction of EAE.These results show that T(EM) cells can behave as endogenous adjuvants and suggest a mechanistic link between lymphocyte traffic in lymph nodes and induction of autoimmunity.

View Article: PubMed Central - PubMed

Affiliation: Institute for Research in Biomedicine, 6500 Bellinzona, Switzerland. alfonso.martin-fontecha@kcl.ac.uk

ABSTRACT
There is growing evidence that the maturation state of dendritic cells (DCs) is a critical parameter determining the balance between tolerance and immunity. We report that mouse CD4(+) effector memory T (T(EM)) cells, but not naive or central memory T cells, constitutively expressed CD40L at levels sufficient to induce DC maturation in vitro and in vivo in the absence of antigenic stimulation. CD4(+) T(EM) cells were excluded from resting lymph nodes but migrated in a CD62P-dependent fashion into reactive lymph nodes that were induced to express CD62P, in a transient or sustained fashion, on high endothelial venules. Trafficking of CD4(+) T(EM) cells into chronic reactive lymph nodes maintained resident DCs in a mature state and promoted naive T cell responses and experimental autoimmune encephalomyelitis (EAE) to antigens administered in the absence of adjuvants. Antibodies to CD62P, which blocked CD4(+) T(EM) cell migration into reactive lymph nodes, inhibited DC maturation, T cell priming, and induction of EAE. These results show that T(EM) cells can behave as endogenous adjuvants and suggest a mechanistic link between lymphocyte traffic in lymph nodes and induction of autoimmunity.

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Expression of CD62P on HEVs of reactive lymph nodes correlates with increased lymph node cellularity and selective increase of TEM cell recruitment. (a) Reactive lymph nodes were induced by one or two consecutive (2 d apart) injections of 106 LPS-matured DCs (DC 1× and DC 2×, respectively). After 2 or 30 d, mice received an i.v. injection of 100 μg FITC-labeled anti-CD62P antibody. After 30 min, lymph nodes were collected, snap frozen, and processed for immunohistochemistry. Shown is the expression of PNAd (red) and CD62P (green) in resting and reactive lymph nodes. Bar, 100 μm. (b) CD62P expression in reactive lymph nodes detected by in vivo staining 2 d after s.c. injection of the indicated adjuvants. Results are representative of three independent experiments. Bar, 100 μm. (c) Absolute cell number in resting and reactive draining lymph nodes at different time points after s.c. injection of PBS, as control or CFA, or one or two consecutive s.c. injections of mature DCs. The asterisk indicates the Student's t test on values: CFA versus PBS, P = 0.013; DC 2× vs. PBS, P = 0.001. (d) Fold increase of the indicated cell populations in day 30 reactive lymph nodes compared with resting lymph nodes (naive, CD44lo; memory, CD44hi; TCM, CD44hiCD62L+; TEM, CD44hiCD62L−; NK, CD3− NK1.1+; NKT, CD3+ NK1.1+; B, CD3− CD19+; pDC, B220+ CD11clo; dermal DC, B220− CD11clo; LC, B220− CD11chi). (e) Absolute number of endogenous TEM cells recovered in resting and reactive lymph nodes 30 d after injection of DCs or adjuvants, as indicated. Data are the means ± SD of two to three separate experiments, each performed with two or three mice per condition. p-values were obtained with the Student's t test.
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fig4: Expression of CD62P on HEVs of reactive lymph nodes correlates with increased lymph node cellularity and selective increase of TEM cell recruitment. (a) Reactive lymph nodes were induced by one or two consecutive (2 d apart) injections of 106 LPS-matured DCs (DC 1× and DC 2×, respectively). After 2 or 30 d, mice received an i.v. injection of 100 μg FITC-labeled anti-CD62P antibody. After 30 min, lymph nodes were collected, snap frozen, and processed for immunohistochemistry. Shown is the expression of PNAd (red) and CD62P (green) in resting and reactive lymph nodes. Bar, 100 μm. (b) CD62P expression in reactive lymph nodes detected by in vivo staining 2 d after s.c. injection of the indicated adjuvants. Results are representative of three independent experiments. Bar, 100 μm. (c) Absolute cell number in resting and reactive draining lymph nodes at different time points after s.c. injection of PBS, as control or CFA, or one or two consecutive s.c. injections of mature DCs. The asterisk indicates the Student's t test on values: CFA versus PBS, P = 0.013; DC 2× vs. PBS, P = 0.001. (d) Fold increase of the indicated cell populations in day 30 reactive lymph nodes compared with resting lymph nodes (naive, CD44lo; memory, CD44hi; TCM, CD44hiCD62L+; TEM, CD44hiCD62L−; NK, CD3− NK1.1+; NKT, CD3+ NK1.1+; B, CD3− CD19+; pDC, B220+ CD11clo; dermal DC, B220− CD11clo; LC, B220− CD11chi). (e) Absolute number of endogenous TEM cells recovered in resting and reactive lymph nodes 30 d after injection of DCs or adjuvants, as indicated. Data are the means ± SD of two to three separate experiments, each performed with two or three mice per condition. p-values were obtained with the Student's t test.

Mentions: We next investigated whether and under which conditions CD62P would be expressed on HEVs of lymph nodes. Reactive lymph nodes were induced by s.c. injection of either LPS-matured DCs or adjuvants. After 2 or 30 d, mice were injected i.v. with fluorescinated antibodies to CD62P to stain the luminal side of HEVs. Reactive and resting lymph nodes were collected and examined by immunohistology after counterstaining with PNAd antibodies to identify HEVs. As shown in Fig. 4 a, CD62P was not expressed on the HEVs of resting lymph nodes. After a single injection of mature DCs (DC 1×), CD62P was readily detected on day 2, persisted for ∼6 d, and was no longer detectable on day 30 (Fig. 4 a and not depicted). CD62P was also detected on HEVs of draining lymph nodes 2 d after injection of adjuvants such as CFA, CpG, LPS, IFA (Fig. 4 b), R848, and TNF (not depicted). Remarkably, however, two consecutive injections 2 d apart of DCs (DC 2×) or a single injection of CFA, but not one or two injections of CpG or LPS, led to a sustained expression of CD62P on HEVs for >30 d (Fig. 4 a and not depicted). The different staining patterns of CD62P and PNAd on HEVs may be caused by differences in cellular localization or by technical reasons, because antibodies to CD62P were injected i.v., whereas anti-PNAd antibodies were added on tissue sections.


CD40L+ CD4+ memory T cells migrate in a CD62P-dependent fashion into reactive lymph nodes and license dendritic cells for T cell priming.

Martín-Fontecha A, Baumjohann D, Guarda G, Reboldi A, Hons M, Lanzavecchia A, Sallusto F - J. Exp. Med. (2008)

Expression of CD62P on HEVs of reactive lymph nodes correlates with increased lymph node cellularity and selective increase of TEM cell recruitment. (a) Reactive lymph nodes were induced by one or two consecutive (2 d apart) injections of 106 LPS-matured DCs (DC 1× and DC 2×, respectively). After 2 or 30 d, mice received an i.v. injection of 100 μg FITC-labeled anti-CD62P antibody. After 30 min, lymph nodes were collected, snap frozen, and processed for immunohistochemistry. Shown is the expression of PNAd (red) and CD62P (green) in resting and reactive lymph nodes. Bar, 100 μm. (b) CD62P expression in reactive lymph nodes detected by in vivo staining 2 d after s.c. injection of the indicated adjuvants. Results are representative of three independent experiments. Bar, 100 μm. (c) Absolute cell number in resting and reactive draining lymph nodes at different time points after s.c. injection of PBS, as control or CFA, or one or two consecutive s.c. injections of mature DCs. The asterisk indicates the Student's t test on values: CFA versus PBS, P = 0.013; DC 2× vs. PBS, P = 0.001. (d) Fold increase of the indicated cell populations in day 30 reactive lymph nodes compared with resting lymph nodes (naive, CD44lo; memory, CD44hi; TCM, CD44hiCD62L+; TEM, CD44hiCD62L−; NK, CD3− NK1.1+; NKT, CD3+ NK1.1+; B, CD3− CD19+; pDC, B220+ CD11clo; dermal DC, B220− CD11clo; LC, B220− CD11chi). (e) Absolute number of endogenous TEM cells recovered in resting and reactive lymph nodes 30 d after injection of DCs or adjuvants, as indicated. Data are the means ± SD of two to three separate experiments, each performed with two or three mice per condition. p-values were obtained with the Student's t test.
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fig4: Expression of CD62P on HEVs of reactive lymph nodes correlates with increased lymph node cellularity and selective increase of TEM cell recruitment. (a) Reactive lymph nodes were induced by one or two consecutive (2 d apart) injections of 106 LPS-matured DCs (DC 1× and DC 2×, respectively). After 2 or 30 d, mice received an i.v. injection of 100 μg FITC-labeled anti-CD62P antibody. After 30 min, lymph nodes were collected, snap frozen, and processed for immunohistochemistry. Shown is the expression of PNAd (red) and CD62P (green) in resting and reactive lymph nodes. Bar, 100 μm. (b) CD62P expression in reactive lymph nodes detected by in vivo staining 2 d after s.c. injection of the indicated adjuvants. Results are representative of three independent experiments. Bar, 100 μm. (c) Absolute cell number in resting and reactive draining lymph nodes at different time points after s.c. injection of PBS, as control or CFA, or one or two consecutive s.c. injections of mature DCs. The asterisk indicates the Student's t test on values: CFA versus PBS, P = 0.013; DC 2× vs. PBS, P = 0.001. (d) Fold increase of the indicated cell populations in day 30 reactive lymph nodes compared with resting lymph nodes (naive, CD44lo; memory, CD44hi; TCM, CD44hiCD62L+; TEM, CD44hiCD62L−; NK, CD3− NK1.1+; NKT, CD3+ NK1.1+; B, CD3− CD19+; pDC, B220+ CD11clo; dermal DC, B220− CD11clo; LC, B220− CD11chi). (e) Absolute number of endogenous TEM cells recovered in resting and reactive lymph nodes 30 d after injection of DCs or adjuvants, as indicated. Data are the means ± SD of two to three separate experiments, each performed with two or three mice per condition. p-values were obtained with the Student's t test.
Mentions: We next investigated whether and under which conditions CD62P would be expressed on HEVs of lymph nodes. Reactive lymph nodes were induced by s.c. injection of either LPS-matured DCs or adjuvants. After 2 or 30 d, mice were injected i.v. with fluorescinated antibodies to CD62P to stain the luminal side of HEVs. Reactive and resting lymph nodes were collected and examined by immunohistology after counterstaining with PNAd antibodies to identify HEVs. As shown in Fig. 4 a, CD62P was not expressed on the HEVs of resting lymph nodes. After a single injection of mature DCs (DC 1×), CD62P was readily detected on day 2, persisted for ∼6 d, and was no longer detectable on day 30 (Fig. 4 a and not depicted). CD62P was also detected on HEVs of draining lymph nodes 2 d after injection of adjuvants such as CFA, CpG, LPS, IFA (Fig. 4 b), R848, and TNF (not depicted). Remarkably, however, two consecutive injections 2 d apart of DCs (DC 2×) or a single injection of CFA, but not one or two injections of CpG or LPS, led to a sustained expression of CD62P on HEVs for >30 d (Fig. 4 a and not depicted). The different staining patterns of CD62P and PNAd on HEVs may be caused by differences in cellular localization or by technical reasons, because antibodies to CD62P were injected i.v., whereas anti-PNAd antibodies were added on tissue sections.

Bottom Line: CD4(+) T(EM) cells were excluded from resting lymph nodes but migrated in a CD62P-dependent fashion into reactive lymph nodes that were induced to express CD62P, in a transient or sustained fashion, on high endothelial venules.Antibodies to CD62P, which blocked CD4(+) T(EM) cell migration into reactive lymph nodes, inhibited DC maturation, T cell priming, and induction of EAE.These results show that T(EM) cells can behave as endogenous adjuvants and suggest a mechanistic link between lymphocyte traffic in lymph nodes and induction of autoimmunity.

View Article: PubMed Central - PubMed

Affiliation: Institute for Research in Biomedicine, 6500 Bellinzona, Switzerland. alfonso.martin-fontecha@kcl.ac.uk

ABSTRACT
There is growing evidence that the maturation state of dendritic cells (DCs) is a critical parameter determining the balance between tolerance and immunity. We report that mouse CD4(+) effector memory T (T(EM)) cells, but not naive or central memory T cells, constitutively expressed CD40L at levels sufficient to induce DC maturation in vitro and in vivo in the absence of antigenic stimulation. CD4(+) T(EM) cells were excluded from resting lymph nodes but migrated in a CD62P-dependent fashion into reactive lymph nodes that were induced to express CD62P, in a transient or sustained fashion, on high endothelial venules. Trafficking of CD4(+) T(EM) cells into chronic reactive lymph nodes maintained resident DCs in a mature state and promoted naive T cell responses and experimental autoimmune encephalomyelitis (EAE) to antigens administered in the absence of adjuvants. Antibodies to CD62P, which blocked CD4(+) T(EM) cell migration into reactive lymph nodes, inhibited DC maturation, T cell priming, and induction of EAE. These results show that T(EM) cells can behave as endogenous adjuvants and suggest a mechanistic link between lymphocyte traffic in lymph nodes and induction of autoimmunity.

Show MeSH
Related in: MedlinePlus