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The CD8alpha(+) dendritic cell is responsible for inducing peripheral self-tolerance to tissue-associated antigens.

Belz GT, Behrens GM, Smith CM, Miller JF, Jones C, Lejon K, Fathman CG, Mueller SN, Shortman K, Carbone FR, Heath WR - J. Exp. Med. (2002)

Bottom Line: Although tracking of YFP was inconclusive, the use of a highly sensitive gB-specific hybridoma that produced beta-galactosidase on encounter with antigen, enabled detection of antigen presentation by cells isolated from the pancreatic lymph node.This showed that a CD11c(+)CD8alpha(+) cell was responsible for cross-tolerance, the same DC subset as previously implicated in cross-priming.These data indicate that CD8alpha(+) DCs play a critical role in both tolerance and immunity to cell-associated antigens, providing a potential mechanism by which cytotoxic T lymphocyte can be immunized to viral antigens while maintaining tolerance to self.

View Article: PubMed Central - PubMed

Affiliation: Immunology Division, The Walter and Eliza Hall Institute of Medical Research, Victoria 3050, Australia. belz@wehi.edu.au

ABSTRACT
We previously described a mechanism for the maintenance of peripheral self-tolerance. This involves the cross-presentation of tissue-associated antigens by a bone marrow-derived cell type that stimulates the proliferation and ultimate deletion of self-reactive CD8 T cells. This process has been referred to as cross-tolerance. Here, we characterize the elusive cell type responsible for inducing cross-tolerance as a CD8alpha(+) dendritic cell (DC). To achieve this aim, transgenic mice were generated expressing yellow fluorescent protein (YFP) linked to CTL epitopes for ovalbumin and glycoprotein B (gB) of herpes simplex virus under the rat insulin promoter (RIP). Although tracking of YFP was inconclusive, the use of a highly sensitive gB-specific hybridoma that produced beta-galactosidase on encounter with antigen, enabled detection of antigen presentation by cells isolated from the pancreatic lymph node. This showed that a CD11c(+)CD8alpha(+) cell was responsible for cross-tolerance, the same DC subset as previously implicated in cross-priming. These data indicate that CD8alpha(+) DCs play a critical role in both tolerance and immunity to cell-associated antigens, providing a potential mechanism by which cytotoxic T lymphocyte can be immunized to viral antigens while maintaining tolerance to self.

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Related in: MedlinePlus

Isolation of the cell type responsible for cross-presentation of islet antigens. Pancreatic LNs were removed from RIP-YSS mice and digested with collagenase and DNase. The preparations were then enriched for DCs by magnetic bead depletion of other cells (Materials and Methods). Finally, DC-enriched populations were stained with anti-CD11c, anti-CD45RA, and anti-CD8α and sorted for CD11c+CD45RA− cells that were either CD8α+ or CD8α−. CD45RA was used to exclude plasmacytoid DCs. The CD8α+ and CD8α− DCs were titrated and used to stimulate the gB-specific hybridoma. CD8α+ DCs were recovered in small numbers that only allowed analysis of single wells per dilution, whereas CD8α− DCs were titrated in duplicate and the mean count shown. This experiment was performed twice with similar results.
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fig5: Isolation of the cell type responsible for cross-presentation of islet antigens. Pancreatic LNs were removed from RIP-YSS mice and digested with collagenase and DNase. The preparations were then enriched for DCs by magnetic bead depletion of other cells (Materials and Methods). Finally, DC-enriched populations were stained with anti-CD11c, anti-CD45RA, and anti-CD8α and sorted for CD11c+CD45RA− cells that were either CD8α+ or CD8α−. CD45RA was used to exclude plasmacytoid DCs. The CD8α+ and CD8α− DCs were titrated and used to stimulate the gB-specific hybridoma. CD8α+ DCs were recovered in small numbers that only allowed analysis of single wells per dilution, whereas CD8α− DCs were titrated in duplicate and the mean count shown. This experiment was performed twice with similar results.

Mentions: To confirm that CD8α+ DCs were responsible for cross-tolerance, we sorted CD11c+CD8α+ and CD11c+CD8α− DCs from the pancreatic LNs of RIP-YSS mice and examined their ability to stimulate the gB-specific hybridoma (Fig. 5) . This required 29 mice to recover 11,500 CD8α1 DCs, so we were only able to do a titration of DCs from 5,750 cells per well with a single well per dilution. In this case, however, CD8α+ DCs clearly stimulated gB-specific hybridoma cells in a dose-dependent manner, with no response generated by CD8α− DCs.


The CD8alpha(+) dendritic cell is responsible for inducing peripheral self-tolerance to tissue-associated antigens.

Belz GT, Behrens GM, Smith CM, Miller JF, Jones C, Lejon K, Fathman CG, Mueller SN, Shortman K, Carbone FR, Heath WR - J. Exp. Med. (2002)

Isolation of the cell type responsible for cross-presentation of islet antigens. Pancreatic LNs were removed from RIP-YSS mice and digested with collagenase and DNase. The preparations were then enriched for DCs by magnetic bead depletion of other cells (Materials and Methods). Finally, DC-enriched populations were stained with anti-CD11c, anti-CD45RA, and anti-CD8α and sorted for CD11c+CD45RA− cells that were either CD8α+ or CD8α−. CD45RA was used to exclude plasmacytoid DCs. The CD8α+ and CD8α− DCs were titrated and used to stimulate the gB-specific hybridoma. CD8α+ DCs were recovered in small numbers that only allowed analysis of single wells per dilution, whereas CD8α− DCs were titrated in duplicate and the mean count shown. This experiment was performed twice with similar results.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2194045&req=5

fig5: Isolation of the cell type responsible for cross-presentation of islet antigens. Pancreatic LNs were removed from RIP-YSS mice and digested with collagenase and DNase. The preparations were then enriched for DCs by magnetic bead depletion of other cells (Materials and Methods). Finally, DC-enriched populations were stained with anti-CD11c, anti-CD45RA, and anti-CD8α and sorted for CD11c+CD45RA− cells that were either CD8α+ or CD8α−. CD45RA was used to exclude plasmacytoid DCs. The CD8α+ and CD8α− DCs were titrated and used to stimulate the gB-specific hybridoma. CD8α+ DCs were recovered in small numbers that only allowed analysis of single wells per dilution, whereas CD8α− DCs were titrated in duplicate and the mean count shown. This experiment was performed twice with similar results.
Mentions: To confirm that CD8α+ DCs were responsible for cross-tolerance, we sorted CD11c+CD8α+ and CD11c+CD8α− DCs from the pancreatic LNs of RIP-YSS mice and examined their ability to stimulate the gB-specific hybridoma (Fig. 5) . This required 29 mice to recover 11,500 CD8α1 DCs, so we were only able to do a titration of DCs from 5,750 cells per well with a single well per dilution. In this case, however, CD8α+ DCs clearly stimulated gB-specific hybridoma cells in a dose-dependent manner, with no response generated by CD8α− DCs.

Bottom Line: Although tracking of YFP was inconclusive, the use of a highly sensitive gB-specific hybridoma that produced beta-galactosidase on encounter with antigen, enabled detection of antigen presentation by cells isolated from the pancreatic lymph node.This showed that a CD11c(+)CD8alpha(+) cell was responsible for cross-tolerance, the same DC subset as previously implicated in cross-priming.These data indicate that CD8alpha(+) DCs play a critical role in both tolerance and immunity to cell-associated antigens, providing a potential mechanism by which cytotoxic T lymphocyte can be immunized to viral antigens while maintaining tolerance to self.

View Article: PubMed Central - PubMed

Affiliation: Immunology Division, The Walter and Eliza Hall Institute of Medical Research, Victoria 3050, Australia. belz@wehi.edu.au

ABSTRACT
We previously described a mechanism for the maintenance of peripheral self-tolerance. This involves the cross-presentation of tissue-associated antigens by a bone marrow-derived cell type that stimulates the proliferation and ultimate deletion of self-reactive CD8 T cells. This process has been referred to as cross-tolerance. Here, we characterize the elusive cell type responsible for inducing cross-tolerance as a CD8alpha(+) dendritic cell (DC). To achieve this aim, transgenic mice were generated expressing yellow fluorescent protein (YFP) linked to CTL epitopes for ovalbumin and glycoprotein B (gB) of herpes simplex virus under the rat insulin promoter (RIP). Although tracking of YFP was inconclusive, the use of a highly sensitive gB-specific hybridoma that produced beta-galactosidase on encounter with antigen, enabled detection of antigen presentation by cells isolated from the pancreatic lymph node. This showed that a CD11c(+)CD8alpha(+) cell was responsible for cross-tolerance, the same DC subset as previously implicated in cross-priming. These data indicate that CD8alpha(+) DCs play a critical role in both tolerance and immunity to cell-associated antigens, providing a potential mechanism by which cytotoxic T lymphocyte can be immunized to viral antigens while maintaining tolerance to self.

Show MeSH
Related in: MedlinePlus