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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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Injection of antigen-loaded BM-DC into mLN afferent lymphatics induces proliferation of antigen-specific T cells. Recipient mice received CFSE-labeled LN cells isolated from DO11.10, OT-I, or OT-II donors. 1 d later, these mice received oil by gavage 1 h before surgery to visualize mLN afferent lymphatics and to facilitate i.l. injection. (A) The small intestine was exposed by surgery and mLN afferent lymphatics were identified by their white color. (B) To demonstrate the method of i.l. injection, ∼0.3 μl of blue dye was injected into a single lymph vessel. After injection of 2× 1 μl into two separate vessels, as performed for routine injection of DC, the fluid spread throughout the LN sinus (not depicted). (C) 105 BM-DC was injected into two separate lymphatic vessels opening into the distal aspect of the mLN chain. Injection of Ova loaded DC (solid black line), but not heat-killed DC (red line), induced proliferation of Ova-specific DO11.10 T cells (DAPI−KJ16-26+CD4+). BM-DC derived from MHC class II–deficient mice induced only marginal proliferation of OT-II cells. Similarly, DC derived from BM of bm-1 mice failed to activate OT-I T cells. All experiments have been performed at least two times with three or more mice per group.
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fig3: Injection of antigen-loaded BM-DC into mLN afferent lymphatics induces proliferation of antigen-specific T cells. Recipient mice received CFSE-labeled LN cells isolated from DO11.10, OT-I, or OT-II donors. 1 d later, these mice received oil by gavage 1 h before surgery to visualize mLN afferent lymphatics and to facilitate i.l. injection. (A) The small intestine was exposed by surgery and mLN afferent lymphatics were identified by their white color. (B) To demonstrate the method of i.l. injection, ∼0.3 μl of blue dye was injected into a single lymph vessel. After injection of 2× 1 μl into two separate vessels, as performed for routine injection of DC, the fluid spread throughout the LN sinus (not depicted). (C) 105 BM-DC was injected into two separate lymphatic vessels opening into the distal aspect of the mLN chain. Injection of Ova loaded DC (solid black line), but not heat-killed DC (red line), induced proliferation of Ova-specific DO11.10 T cells (DAPI−KJ16-26+CD4+). BM-DC derived from MHC class II–deficient mice induced only marginal proliferation of OT-II cells. Similarly, DC derived from BM of bm-1 mice failed to activate OT-I T cells. All experiments have been performed at least two times with three or more mice per group.

Mentions: Analyzing the potential of distinct DC subsets to generate gut-homing T cells in vivo requires uncoupling the origin of the DC from their usual destination, i.e., the draining LN. To this aim, we established a new method to inject DC directly into the mLN afferent lymphatics (intralymphatic [i.l.] injection). After oil feeding, the small intestine was exposed by surgery and the mLN afferent lymphatics were located. 105 BM-DC was injected into two separate lymphatic vessels opening into the distal aspect of the mLN chain using a fine glass capillary (Fig. 3, A and B). Injection of Ova-loaded BM-DC readily induced proliferation of only antigen-specific adoptively transferred T cells, indicating that no overt unspecific T cell proliferation was induced. Moreover, DC defective in antigen presentation in the context of either MHC class I or class II failed to provoke T cell proliferation (Fig. 3 C). Thus, T cell proliferation upon i.l. injection of BM-DC reflects the direct priming by the injected DC and does not result from antigen passed on to LN resident DC.


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)

Injection of antigen-loaded BM-DC into mLN afferent lymphatics induces proliferation of antigen-specific T cells. Recipient mice received CFSE-labeled LN cells isolated from DO11.10, OT-I, or OT-II donors. 1 d later, these mice received oil by gavage 1 h before surgery to visualize mLN afferent lymphatics and to facilitate i.l. injection. (A) The small intestine was exposed by surgery and mLN afferent lymphatics were identified by their white color. (B) To demonstrate the method of i.l. injection, ∼0.3 μl of blue dye was injected into a single lymph vessel. After injection of 2× 1 μl into two separate vessels, as performed for routine injection of DC, the fluid spread throughout the LN sinus (not depicted). (C) 105 BM-DC was injected into two separate lymphatic vessels opening into the distal aspect of the mLN chain. Injection of Ova loaded DC (solid black line), but not heat-killed DC (red line), induced proliferation of Ova-specific DO11.10 T cells (DAPI−KJ16-26+CD4+). BM-DC derived from MHC class II–deficient mice induced only marginal proliferation of OT-II cells. Similarly, DC derived from BM of bm-1 mice failed to activate OT-I T cells. All experiments have been performed at least two times with three or more mice per group.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2571923&req=5

fig3: Injection of antigen-loaded BM-DC into mLN afferent lymphatics induces proliferation of antigen-specific T cells. Recipient mice received CFSE-labeled LN cells isolated from DO11.10, OT-I, or OT-II donors. 1 d later, these mice received oil by gavage 1 h before surgery to visualize mLN afferent lymphatics and to facilitate i.l. injection. (A) The small intestine was exposed by surgery and mLN afferent lymphatics were identified by their white color. (B) To demonstrate the method of i.l. injection, ∼0.3 μl of blue dye was injected into a single lymph vessel. After injection of 2× 1 μl into two separate vessels, as performed for routine injection of DC, the fluid spread throughout the LN sinus (not depicted). (C) 105 BM-DC was injected into two separate lymphatic vessels opening into the distal aspect of the mLN chain. Injection of Ova loaded DC (solid black line), but not heat-killed DC (red line), induced proliferation of Ova-specific DO11.10 T cells (DAPI−KJ16-26+CD4+). BM-DC derived from MHC class II–deficient mice induced only marginal proliferation of OT-II cells. Similarly, DC derived from BM of bm-1 mice failed to activate OT-I T cells. All experiments have been performed at least two times with three or more mice per group.
Mentions: Analyzing the potential of distinct DC subsets to generate gut-homing T cells in vivo requires uncoupling the origin of the DC from their usual destination, i.e., the draining LN. To this aim, we established a new method to inject DC directly into the mLN afferent lymphatics (intralymphatic [i.l.] injection). After oil feeding, the small intestine was exposed by surgery and the mLN afferent lymphatics were located. 105 BM-DC was injected into two separate lymphatic vessels opening into the distal aspect of the mLN chain using a fine glass capillary (Fig. 3, A and B). Injection of Ova-loaded BM-DC readily induced proliferation of only antigen-specific adoptively transferred T cells, indicating that no overt unspecific T cell proliferation was induced. Moreover, DC defective in antigen presentation in the context of either MHC class I or class II failed to provoke T cell proliferation (Fig. 3 C). Thus, T cell proliferation upon i.l. injection of BM-DC reflects the direct priming by the injected DC and does not result from antigen passed on to LN resident DC.

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