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Lymph node stromal cells acquire peptide-MHCII complexes from dendritic cells and induce antigen-specific CD4⁺ T cell tolerance.

Dubrot J, Duraes FV, Potin L, Capotosti F, Brighouse D, Suter T, LeibundGut-Landmann S, Garbi N, Reith W, Swartz MA, Hugues S - J. Exp. Med. (2014)

Bottom Line: Although LNSCs express MHCII, it is unknown whether they can also impact CD4(+) T cell functions.We show that the promoter IV (pIV) of class II transactivator (CIITA), the master regulator of MHCII expression, controls endogenous MHCII expression by LNSCs.Our data reveals a novel, alternative mechanism where LN-resident stromal cells tolerize CD4(+) T cells through the presentation of self-antigens via transferred peptide-MHCII complexes of DC origin.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology and Immunology, University of Geneva Medical School, 1211 Geneva, Switzerland.

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DCs transfer pMHCII complexes to LEC and FRC in vitro. (A and B) WT (A) and CIITA−/− (B) LEC/FRC were co-cultured with BM-DCs preloaded with OVA323-339 (red histograms) or FITC-labeled OVA323-339 (green histograms). After 24 h, FITC signals was analyzed by FACS in LECs and FRCs and compared with isotype control (gray histogram). Graphs show FITC MFI and are representative of 3 independent experiments. Error bars depict mean ± SEM. ***, P < 0.001. (C) Exosomes were purified from supernatant of 5 × 106 BMDCs loaded with FITC-labeled (green) or unlabeled (red) OVA peptide (1 µM). DC-derived exosomes were coupled with latex beads and analyzed by FACS for FITC fluorescence (left) and for MHCII expression (compared with isotype control in gray) after gating on FITC+ exosomes (right). Data representative of 3 independent experiments.
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fig6: DCs transfer pMHCII complexes to LEC and FRC in vitro. (A and B) WT (A) and CIITA−/− (B) LEC/FRC were co-cultured with BM-DCs preloaded with OVA323-339 (red histograms) or FITC-labeled OVA323-339 (green histograms). After 24 h, FITC signals was analyzed by FACS in LECs and FRCs and compared with isotype control (gray histogram). Graphs show FITC MFI and are representative of 3 independent experiments. Error bars depict mean ± SEM. ***, P < 0.001. (C) Exosomes were purified from supernatant of 5 × 106 BMDCs loaded with FITC-labeled (green) or unlabeled (red) OVA peptide (1 µM). DC-derived exosomes were coupled with latex beads and analyzed by FACS for FITC fluorescence (left) and for MHCII expression (compared with isotype control in gray) after gating on FITC+ exosomes (right). Data representative of 3 independent experiments.

Mentions: Next, we asked whether peptide-loaded MHCII molecules (pMHCII) could also be transferred from DCs to LNSCs, and if so, whether these transferred complexes could be functionally presented to CD4+ T cells. To this end, we first loaded DCs with FITC-labeled OVA323-339 peptide and, after thorough washing, co-cultured them with LEC/FRCs. We found efficient transfer of FITC fluorescence to both LECs and FRCs (Fig. 6 A), suggesting that DCs can transfer pMHCII complexes to LNSCs in vitro. Importantly, similar results were obtained using CIITA−/− LEC/FRC cells, ruling out the possibility that endogenous MHCII molecules expressed by LECs and FRCs were loaded with free FITC+ OVA peptide (Fig. 6 B). Consistent with a possible role for exosomes in the transfer of pMHCII to LNSCs, exosomes derived from FITC-labeled OVA323-339-loaded DCs were FITC+ (Fig. 6 C). Importantly, all FITC+ exosomes were MHCII positive, reinforcing the idea that exosomes might transfer pMHCII complexes rather than free peptide (Fig. 6 C).


Lymph node stromal cells acquire peptide-MHCII complexes from dendritic cells and induce antigen-specific CD4⁺ T cell tolerance.

Dubrot J, Duraes FV, Potin L, Capotosti F, Brighouse D, Suter T, LeibundGut-Landmann S, Garbi N, Reith W, Swartz MA, Hugues S - J. Exp. Med. (2014)

DCs transfer pMHCII complexes to LEC and FRC in vitro. (A and B) WT (A) and CIITA−/− (B) LEC/FRC were co-cultured with BM-DCs preloaded with OVA323-339 (red histograms) or FITC-labeled OVA323-339 (green histograms). After 24 h, FITC signals was analyzed by FACS in LECs and FRCs and compared with isotype control (gray histogram). Graphs show FITC MFI and are representative of 3 independent experiments. Error bars depict mean ± SEM. ***, P < 0.001. (C) Exosomes were purified from supernatant of 5 × 106 BMDCs loaded with FITC-labeled (green) or unlabeled (red) OVA peptide (1 µM). DC-derived exosomes were coupled with latex beads and analyzed by FACS for FITC fluorescence (left) and for MHCII expression (compared with isotype control in gray) after gating on FITC+ exosomes (right). Data representative of 3 independent experiments.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4042642&req=5

fig6: DCs transfer pMHCII complexes to LEC and FRC in vitro. (A and B) WT (A) and CIITA−/− (B) LEC/FRC were co-cultured with BM-DCs preloaded with OVA323-339 (red histograms) or FITC-labeled OVA323-339 (green histograms). After 24 h, FITC signals was analyzed by FACS in LECs and FRCs and compared with isotype control (gray histogram). Graphs show FITC MFI and are representative of 3 independent experiments. Error bars depict mean ± SEM. ***, P < 0.001. (C) Exosomes were purified from supernatant of 5 × 106 BMDCs loaded with FITC-labeled (green) or unlabeled (red) OVA peptide (1 µM). DC-derived exosomes were coupled with latex beads and analyzed by FACS for FITC fluorescence (left) and for MHCII expression (compared with isotype control in gray) after gating on FITC+ exosomes (right). Data representative of 3 independent experiments.
Mentions: Next, we asked whether peptide-loaded MHCII molecules (pMHCII) could also be transferred from DCs to LNSCs, and if so, whether these transferred complexes could be functionally presented to CD4+ T cells. To this end, we first loaded DCs with FITC-labeled OVA323-339 peptide and, after thorough washing, co-cultured them with LEC/FRCs. We found efficient transfer of FITC fluorescence to both LECs and FRCs (Fig. 6 A), suggesting that DCs can transfer pMHCII complexes to LNSCs in vitro. Importantly, similar results were obtained using CIITA−/− LEC/FRC cells, ruling out the possibility that endogenous MHCII molecules expressed by LECs and FRCs were loaded with free FITC+ OVA peptide (Fig. 6 B). Consistent with a possible role for exosomes in the transfer of pMHCII to LNSCs, exosomes derived from FITC-labeled OVA323-339-loaded DCs were FITC+ (Fig. 6 C). Importantly, all FITC+ exosomes were MHCII positive, reinforcing the idea that exosomes might transfer pMHCII complexes rather than free peptide (Fig. 6 C).

Bottom Line: Although LNSCs express MHCII, it is unknown whether they can also impact CD4(+) T cell functions.We show that the promoter IV (pIV) of class II transactivator (CIITA), the master regulator of MHCII expression, controls endogenous MHCII expression by LNSCs.Our data reveals a novel, alternative mechanism where LN-resident stromal cells tolerize CD4(+) T cells through the presentation of self-antigens via transferred peptide-MHCII complexes of DC origin.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology and Immunology, University of Geneva Medical School, 1211 Geneva, Switzerland.

Show MeSH
Related in: MedlinePlus