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Efficient targeting of protein antigen to the dendritic cell receptor DEC-205 in the steady state leads to antigen presentation on major histocompatibility complex class I products and peripheral CD8+ T cell tolerance.

Bonifaz L, Bonnyay D, Mahnke K, Rivera M, Nussenzweig MC, Steinman RM - J. Exp. Med. (2002)

Bottom Line: In vivo, the OVA protein was selectively presented by DCs to TCR transgenic CD8+ cells, again at least 400 times more effectively than soluble OVA and in a TAP-dependent fashion.The CD8+ T cells responding in the presence of agonistic alphaCD40 antibody produced large amounts of interleukin 2 and interferon gamma, acquired cytolytic function in vivo, emigrated in large numbers to the lung, and responded vigorously to OVA rechallenge.Therefore, DEC-205 provides an efficient receptor-based mechanism for DCs to process proteins for MHC class I presentation in vivo, leading to tolerance in the steady state and immunity after DC maturation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cellular Physiology and Immunology, The Rockefeller University, New York, NY 10021, USA.

ABSTRACT
To identify endocytic receptors that allow dendritic cells (DCs) to capture and present antigens on major histocompatibility complex (MHC) class I products in vivo, we evaluated DEC-205, which is abundant on DCs in lymphoid tissues. Ovalbumin (OVA) protein, when chemically coupled to monoclonal alphaDEC-205 antibody, was presented by CD11c+ lymph node DCs, but not by CD11c- cells, to OVA-specific, CD4+ and CD8+ T cells. Receptor-mediated presentation was at least 400 times more efficient than unconjugated OVA and, for MHC class I, the DCs had to express transporter of antigenic peptides (TAP) transporters. When alphaDEC-205:OVA was injected subcutaneously, OVA protein was identified over a 4-48 h period in DCs, primarily in the lymph nodes draining the injection site. In vivo, the OVA protein was selectively presented by DCs to TCR transgenic CD8+ cells, again at least 400 times more effectively than soluble OVA and in a TAP-dependent fashion. Targeting of alphaDEC-205:OVA to DCs in the steady state initially induced 4-7 cycles of T cell division, but the T cells were then deleted and the mice became specifically unresponsive to rechallenge with OVA in complete Freund's adjuvant. In contrast, simultaneous delivery of a DC maturation stimulus via CD40, together with alphaDEC-205:OVA, induced strong immunity. The CD8+ T cells responding in the presence of agonistic alphaCD40 antibody produced large amounts of interleukin 2 and interferon gamma, acquired cytolytic function in vivo, emigrated in large numbers to the lung, and responded vigorously to OVA rechallenge. Therefore, DEC-205 provides an efficient receptor-based mechanism for DCs to process proteins for MHC class I presentation in vivo, leading to tolerance in the steady state and immunity after DC maturation.

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Maturation of DCs in vivo by agonistic αCD40 but not by αDEC-205:OVA. (A) C57BL/6 mice were injected subcutaneously with PBS or 4.0 μg (1.0 μg/footpad) of αDEC-205:OVA conjugate with or without αCD40 (100 μg FGK45.5 subcutaneously), 1 and 3 d before sacrifice. CD11c+ cells were sorted by MACS® from lymph nodes and evaluated by flow cytometry for expression of CD80, CD86, and MHC class II. Prior to injection of the OVA conjugate and αCD40, the mice were given PBS (−) or OT-I (+) cells. The bold symbols are mean fluorescence indices of the CD11c+ cells in the presence of a maturation stimulus, while the gray-bold at day 3 indicate a significant increase, consistent with maturation. (B) Illustrative FACS® data showing the maturation of the DEC-205hi CD11c+ cells and DEC-205loCD11c+ cells, in mice treated 3 d before with PBS and αCD40 as in panel A.
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fig4: Maturation of DCs in vivo by agonistic αCD40 but not by αDEC-205:OVA. (A) C57BL/6 mice were injected subcutaneously with PBS or 4.0 μg (1.0 μg/footpad) of αDEC-205:OVA conjugate with or without αCD40 (100 μg FGK45.5 subcutaneously), 1 and 3 d before sacrifice. CD11c+ cells were sorted by MACS® from lymph nodes and evaluated by flow cytometry for expression of CD80, CD86, and MHC class II. Prior to injection of the OVA conjugate and αCD40, the mice were given PBS (−) or OT-I (+) cells. The bold symbols are mean fluorescence indices of the CD11c+ cells in the presence of a maturation stimulus, while the gray-bold at day 3 indicate a significant increase, consistent with maturation. (B) Illustrative FACS® data showing the maturation of the DEC-205hi CD11c+ cells and DEC-205loCD11c+ cells, in mice treated 3 d before with PBS and αCD40 as in panel A.

Mentions: To examine whether αDEC-205:OVA treatment results in DC maturation in the presence or absence of OVA-specific OT-I T cells, we did FACS® studies of DCs from mice injected with conjugates 1 or 3 d earlier under a variety of conditions. As illustrated in Fig. 4 A, surface expression of CD80, CD86, as well as MHC class II products were unchanged in αDEC-205:OVA-injected mice, whether or not they received OT-1 T cells. The number of DCs also did not change in mice given αDEC-205:OVA. However, coadministration of an agonistic αCD40 antibody (FGK-45.5) as an adjuvant activated the DCs in situ over a 3 d period and increased their numbers about twofold. The extent of maturation with αCD40 was similar in the absence or presence of antigen (αDEC-205:OVA) or OT-I T cells (Fig. 4 A). Maturation was detected in CD11c+ DCs that had low and high levels of DEC-205, but the levels of CD86 were higher in the DEC-205 high fraction (Fig. 4 B). In summary, although lymph node DCs in the steady state express molecules used in T cell activation like CD86, these DCs do not seem to differentiate further when exposed to αDEC-205:OVA but do differentiate in response to agonistic αCD40 antibody.


Efficient targeting of protein antigen to the dendritic cell receptor DEC-205 in the steady state leads to antigen presentation on major histocompatibility complex class I products and peripheral CD8+ T cell tolerance.

Bonifaz L, Bonnyay D, Mahnke K, Rivera M, Nussenzweig MC, Steinman RM - J. Exp. Med. (2002)

Maturation of DCs in vivo by agonistic αCD40 but not by αDEC-205:OVA. (A) C57BL/6 mice were injected subcutaneously with PBS or 4.0 μg (1.0 μg/footpad) of αDEC-205:OVA conjugate with or without αCD40 (100 μg FGK45.5 subcutaneously), 1 and 3 d before sacrifice. CD11c+ cells were sorted by MACS® from lymph nodes and evaluated by flow cytometry for expression of CD80, CD86, and MHC class II. Prior to injection of the OVA conjugate and αCD40, the mice were given PBS (−) or OT-I (+) cells. The bold symbols are mean fluorescence indices of the CD11c+ cells in the presence of a maturation stimulus, while the gray-bold at day 3 indicate a significant increase, consistent with maturation. (B) Illustrative FACS® data showing the maturation of the DEC-205hi CD11c+ cells and DEC-205loCD11c+ cells, in mice treated 3 d before with PBS and αCD40 as in panel A.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2196060&req=5

fig4: Maturation of DCs in vivo by agonistic αCD40 but not by αDEC-205:OVA. (A) C57BL/6 mice were injected subcutaneously with PBS or 4.0 μg (1.0 μg/footpad) of αDEC-205:OVA conjugate with or without αCD40 (100 μg FGK45.5 subcutaneously), 1 and 3 d before sacrifice. CD11c+ cells were sorted by MACS® from lymph nodes and evaluated by flow cytometry for expression of CD80, CD86, and MHC class II. Prior to injection of the OVA conjugate and αCD40, the mice were given PBS (−) or OT-I (+) cells. The bold symbols are mean fluorescence indices of the CD11c+ cells in the presence of a maturation stimulus, while the gray-bold at day 3 indicate a significant increase, consistent with maturation. (B) Illustrative FACS® data showing the maturation of the DEC-205hi CD11c+ cells and DEC-205loCD11c+ cells, in mice treated 3 d before with PBS and αCD40 as in panel A.
Mentions: To examine whether αDEC-205:OVA treatment results in DC maturation in the presence or absence of OVA-specific OT-I T cells, we did FACS® studies of DCs from mice injected with conjugates 1 or 3 d earlier under a variety of conditions. As illustrated in Fig. 4 A, surface expression of CD80, CD86, as well as MHC class II products were unchanged in αDEC-205:OVA-injected mice, whether or not they received OT-1 T cells. The number of DCs also did not change in mice given αDEC-205:OVA. However, coadministration of an agonistic αCD40 antibody (FGK-45.5) as an adjuvant activated the DCs in situ over a 3 d period and increased their numbers about twofold. The extent of maturation with αCD40 was similar in the absence or presence of antigen (αDEC-205:OVA) or OT-I T cells (Fig. 4 A). Maturation was detected in CD11c+ DCs that had low and high levels of DEC-205, but the levels of CD86 were higher in the DEC-205 high fraction (Fig. 4 B). In summary, although lymph node DCs in the steady state express molecules used in T cell activation like CD86, these DCs do not seem to differentiate further when exposed to αDEC-205:OVA but do differentiate in response to agonistic αCD40 antibody.

Bottom Line: In vivo, the OVA protein was selectively presented by DCs to TCR transgenic CD8+ cells, again at least 400 times more effectively than soluble OVA and in a TAP-dependent fashion.The CD8+ T cells responding in the presence of agonistic alphaCD40 antibody produced large amounts of interleukin 2 and interferon gamma, acquired cytolytic function in vivo, emigrated in large numbers to the lung, and responded vigorously to OVA rechallenge.Therefore, DEC-205 provides an efficient receptor-based mechanism for DCs to process proteins for MHC class I presentation in vivo, leading to tolerance in the steady state and immunity after DC maturation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cellular Physiology and Immunology, The Rockefeller University, New York, NY 10021, USA.

ABSTRACT
To identify endocytic receptors that allow dendritic cells (DCs) to capture and present antigens on major histocompatibility complex (MHC) class I products in vivo, we evaluated DEC-205, which is abundant on DCs in lymphoid tissues. Ovalbumin (OVA) protein, when chemically coupled to monoclonal alphaDEC-205 antibody, was presented by CD11c+ lymph node DCs, but not by CD11c- cells, to OVA-specific, CD4+ and CD8+ T cells. Receptor-mediated presentation was at least 400 times more efficient than unconjugated OVA and, for MHC class I, the DCs had to express transporter of antigenic peptides (TAP) transporters. When alphaDEC-205:OVA was injected subcutaneously, OVA protein was identified over a 4-48 h period in DCs, primarily in the lymph nodes draining the injection site. In vivo, the OVA protein was selectively presented by DCs to TCR transgenic CD8+ cells, again at least 400 times more effectively than soluble OVA and in a TAP-dependent fashion. Targeting of alphaDEC-205:OVA to DCs in the steady state initially induced 4-7 cycles of T cell division, but the T cells were then deleted and the mice became specifically unresponsive to rechallenge with OVA in complete Freund's adjuvant. In contrast, simultaneous delivery of a DC maturation stimulus via CD40, together with alphaDEC-205:OVA, induced strong immunity. The CD8+ T cells responding in the presence of agonistic alphaCD40 antibody produced large amounts of interleukin 2 and interferon gamma, acquired cytolytic function in vivo, emigrated in large numbers to the lung, and responded vigorously to OVA rechallenge. Therefore, DEC-205 provides an efficient receptor-based mechanism for DCs to process proteins for MHC class I presentation in vivo, leading to tolerance in the steady state and immunity after DC maturation.

Show MeSH
Related in: MedlinePlus