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Stromal mesenteric lymph node cells are essential for the generation of gut-homing T cells in vivo.

Hammerschmidt SI, Ahrendt M, Bode U, Wahl B, Kremmer E, Förster R, Pabst O - J. Exp. Med. (2008)

Bottom Line: DC that fail to induce alpha4beta7-integrin and CCR9 in vitro readily induce these factors in vivo upon injection into mLN afferent lymphatics.Moreover, uniquely mesenteric but not pLN stroma cells express high levels of RA-producing enzymes and support induction of CCR9 on activated T cells in vitro.These results demonstrate a hitherto unrecognized contribution of stromal cell delivered signals, including RA, on the imprinting of tissue tropism in vivo.

View Article: PubMed Central - PubMed

Affiliation: Institute of Immunology, Hannover Medical School, 30625 Hannover, Germany.

ABSTRACT
T cells primed in the gut-draining mesenteric lymph nodes (mLN) are imprinted to express alpha4beta7-integrin and chemokine receptor CCR9, thereby enabling lymphocytes to migrate to the small intestine. In vitro activation by intestinal dendritic cells (DC) or addition of retinoic acid (RA) is sufficient to instruct expression of these gut-homing molecules. We report that in vivo stroma cells, but not DC, allow the mLN to induce the generation of gut tropism. Peripheral LN (pLN) transplanted into the gut mesenteries fail to support the generation of gut-homing T cells, even though gut-derived DC enter the transplants and prime T cells. DC that fail to induce alpha4beta7-integrin and CCR9 in vitro readily induce these factors in vivo upon injection into mLN afferent lymphatics. Moreover, uniquely mesenteric but not pLN stroma cells express high levels of RA-producing enzymes and support induction of CCR9 on activated T cells in vitro. These results demonstrate a hitherto unrecognized contribution of stromal cell delivered signals, including RA, on the imprinting of tissue tropism in vivo.

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BM-DC and spleen-derived DC generate gut-homing T cells in vivo but not in vitro. (A and B) DO11.10 cells were adoptively transferred into WT recipients. 1 d later, BM-DC were either injected i.l. or s.c., and proliferating DO11.10 T cells in the mLN and pLN were analyzed for expression of α4β7-integrin, CCR9, and E- and P-selectin ligand as indicated. Moreover, expression of these molecules was assessed after in vitro coculture of antigen-loaded BM-DC with DO11.10 T cells at a 1:10 ratio. BM-DC injected into mLN afferent lymphatics, but not injected s.c., or in vitro cocultures led to the up-regulation of α4β7-integrin and CCR9 on proliferating T cells. Conversely, s.c., but not i.l., injection of BM-DC induced expression of E- and P-selectin ligand. (C and D) DC were isolated from mLN and spleen, loaded in vitro with Ova peptide, and injected i.l. into recipient mice as described in Fig. 3. Spleen-derived DC that fail to up-regulate gut-homing factors after 3 d of coculture with DO11.10 cells in vitro (not depicted) readily induced expression of α4β7-integrin and modest expression of CCR9 after i.l. injection in vivo. All experiments were performed at least three times with at least two mice per group.
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fig4: BM-DC and spleen-derived DC generate gut-homing T cells in vivo but not in vitro. (A and B) DO11.10 cells were adoptively transferred into WT recipients. 1 d later, BM-DC were either injected i.l. or s.c., and proliferating DO11.10 T cells in the mLN and pLN were analyzed for expression of α4β7-integrin, CCR9, and E- and P-selectin ligand as indicated. Moreover, expression of these molecules was assessed after in vitro coculture of antigen-loaded BM-DC with DO11.10 T cells at a 1:10 ratio. BM-DC injected into mLN afferent lymphatics, but not injected s.c., or in vitro cocultures led to the up-regulation of α4β7-integrin and CCR9 on proliferating T cells. Conversely, s.c., but not i.l., injection of BM-DC induced expression of E- and P-selectin ligand. (C and D) DC were isolated from mLN and spleen, loaded in vitro with Ova peptide, and injected i.l. into recipient mice as described in Fig. 3. Spleen-derived DC that fail to up-regulate gut-homing factors after 3 d of coculture with DO11.10 cells in vitro (not depicted) readily induced expression of α4β7-integrin and modest expression of CCR9 after i.l. injection in vivo. All experiments were performed at least three times with at least two mice per group.

Mentions: We next explored the phenotype of T cells primed in the mLN upon i.l. injection of DC. As described in the previous paragraph, antigen-loaded DC were injected into mLN afferent lymphatics of WT recipients that previously received CFSE-labeled Ova-specific DO11.10 cells. 3 d later, cells were isolated from the mLN and analyzed for the expression of α4β7-integrin, CCR9, and E- and P-selectin ligand. For comparison, another group of mice received 106 DC-injected s.c., and cells isolated from the skin-draining pLN were analyzed. Strikingly, BM-DC that fail to induce CCR9 on T cells in vitro (Fig. 4 B) (9) efficiently induced CCR9 after i.l. injection in vivo (Fig. 4, A and B). Similarly, in vitro expression of α4β7-integrin is induced by BM-DC only after prolonged culture (18; for review see reference 3) but was rapidly induced in vivo (Fig. 4, A and B). Thus, T cells primed by BM-DC in vitro largely differ with respect to their homing properties from T cells primed by BM-DC in vivo. We next extended our experiments to primary DC. DC were isolated from spleen or mLN of WT mice and loaded with Ova peptide in vitro. Expectedly, only mLN-derived DC induced a gut-homing phenotype on T cells in vitro (unpublished data). In contrast, both mLN- and spleen-derived DC induced considerable up-regulation of CCR9 and α4β7-integrin upon i.l. injection (Fig. 4, C and D). In line with these observations, i.p. injection of BM-DC has also been reported to induce expression of α4β7-integrin on antigen-specific proliferating T cells in the mLN (18). This indicates that even though mLN-derived DC might be more efficient in inducing CCR9 (Fig. 4, C and D) compared with spleen-derived DC, the LN environment and not the origin of DC determines the generation of gut-homing T cells in vivo.


Stromal mesenteric lymph node cells are essential for the generation of gut-homing T cells in vivo.

Hammerschmidt SI, Ahrendt M, Bode U, Wahl B, Kremmer E, Förster R, Pabst O - J. Exp. Med. (2008)

BM-DC and spleen-derived DC generate gut-homing T cells in vivo but not in vitro. (A and B) DO11.10 cells were adoptively transferred into WT recipients. 1 d later, BM-DC were either injected i.l. or s.c., and proliferating DO11.10 T cells in the mLN and pLN were analyzed for expression of α4β7-integrin, CCR9, and E- and P-selectin ligand as indicated. Moreover, expression of these molecules was assessed after in vitro coculture of antigen-loaded BM-DC with DO11.10 T cells at a 1:10 ratio. BM-DC injected into mLN afferent lymphatics, but not injected s.c., or in vitro cocultures led to the up-regulation of α4β7-integrin and CCR9 on proliferating T cells. Conversely, s.c., but not i.l., injection of BM-DC induced expression of E- and P-selectin ligand. (C and D) DC were isolated from mLN and spleen, loaded in vitro with Ova peptide, and injected i.l. into recipient mice as described in Fig. 3. Spleen-derived DC that fail to up-regulate gut-homing factors after 3 d of coculture with DO11.10 cells in vitro (not depicted) readily induced expression of α4β7-integrin and modest expression of CCR9 after i.l. injection in vivo. All experiments were performed at least three times with at least two mice per group.
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fig4: BM-DC and spleen-derived DC generate gut-homing T cells in vivo but not in vitro. (A and B) DO11.10 cells were adoptively transferred into WT recipients. 1 d later, BM-DC were either injected i.l. or s.c., and proliferating DO11.10 T cells in the mLN and pLN were analyzed for expression of α4β7-integrin, CCR9, and E- and P-selectin ligand as indicated. Moreover, expression of these molecules was assessed after in vitro coculture of antigen-loaded BM-DC with DO11.10 T cells at a 1:10 ratio. BM-DC injected into mLN afferent lymphatics, but not injected s.c., or in vitro cocultures led to the up-regulation of α4β7-integrin and CCR9 on proliferating T cells. Conversely, s.c., but not i.l., injection of BM-DC induced expression of E- and P-selectin ligand. (C and D) DC were isolated from mLN and spleen, loaded in vitro with Ova peptide, and injected i.l. into recipient mice as described in Fig. 3. Spleen-derived DC that fail to up-regulate gut-homing factors after 3 d of coculture with DO11.10 cells in vitro (not depicted) readily induced expression of α4β7-integrin and modest expression of CCR9 after i.l. injection in vivo. All experiments were performed at least three times with at least two mice per group.
Mentions: We next explored the phenotype of T cells primed in the mLN upon i.l. injection of DC. As described in the previous paragraph, antigen-loaded DC were injected into mLN afferent lymphatics of WT recipients that previously received CFSE-labeled Ova-specific DO11.10 cells. 3 d later, cells were isolated from the mLN and analyzed for the expression of α4β7-integrin, CCR9, and E- and P-selectin ligand. For comparison, another group of mice received 106 DC-injected s.c., and cells isolated from the skin-draining pLN were analyzed. Strikingly, BM-DC that fail to induce CCR9 on T cells in vitro (Fig. 4 B) (9) efficiently induced CCR9 after i.l. injection in vivo (Fig. 4, A and B). Similarly, in vitro expression of α4β7-integrin is induced by BM-DC only after prolonged culture (18; for review see reference 3) but was rapidly induced in vivo (Fig. 4, A and B). Thus, T cells primed by BM-DC in vitro largely differ with respect to their homing properties from T cells primed by BM-DC in vivo. We next extended our experiments to primary DC. DC were isolated from spleen or mLN of WT mice and loaded with Ova peptide in vitro. Expectedly, only mLN-derived DC induced a gut-homing phenotype on T cells in vitro (unpublished data). In contrast, both mLN- and spleen-derived DC induced considerable up-regulation of CCR9 and α4β7-integrin upon i.l. injection (Fig. 4, C and D). In line with these observations, i.p. injection of BM-DC has also been reported to induce expression of α4β7-integrin on antigen-specific proliferating T cells in the mLN (18). This indicates that even though mLN-derived DC might be more efficient in inducing CCR9 (Fig. 4, C and D) compared with spleen-derived DC, the LN environment and not the origin of DC determines the generation of gut-homing T cells in vivo.

Bottom Line: DC that fail to induce alpha4beta7-integrin and CCR9 in vitro readily induce these factors in vivo upon injection into mLN afferent lymphatics.Moreover, uniquely mesenteric but not pLN stroma cells express high levels of RA-producing enzymes and support induction of CCR9 on activated T cells in vitro.These results demonstrate a hitherto unrecognized contribution of stromal cell delivered signals, including RA, on the imprinting of tissue tropism in vivo.

View Article: PubMed Central - PubMed

Affiliation: Institute of Immunology, Hannover Medical School, 30625 Hannover, Germany.

ABSTRACT
T cells primed in the gut-draining mesenteric lymph nodes (mLN) are imprinted to express alpha4beta7-integrin and chemokine receptor CCR9, thereby enabling lymphocytes to migrate to the small intestine. In vitro activation by intestinal dendritic cells (DC) or addition of retinoic acid (RA) is sufficient to instruct expression of these gut-homing molecules. We report that in vivo stroma cells, but not DC, allow the mLN to induce the generation of gut tropism. Peripheral LN (pLN) transplanted into the gut mesenteries fail to support the generation of gut-homing T cells, even though gut-derived DC enter the transplants and prime T cells. DC that fail to induce alpha4beta7-integrin and CCR9 in vitro readily induce these factors in vivo upon injection into mLN afferent lymphatics. Moreover, uniquely mesenteric but not pLN stroma cells express high levels of RA-producing enzymes and support induction of CCR9 on activated T cells in vitro. These results demonstrate a hitherto unrecognized contribution of stromal cell delivered signals, including RA, on the imprinting of tissue tropism in vivo.

Show MeSH