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Blood, Vol. 114, Issue 3, 555-563, July 16, 2009
Indoleamine 2,3-dioxygenase–expressing mature human monocyte-derived dendritic cells expand potent autologous regulatory T cells
Blood Chung et al.
114: 555
Supplemental materials for: Chung et al
Files in this Data Supplement:
- Figure S1. Schema of suppressor T-cell generation by IDO-expressing, mature moDCs (JPG, 181 KB)
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Autologous or allogeneic descriptor was always relative to the initially primed, responder T-cell population. Mature moDCs were cultured with autologous T cells at a 1:10 ratio (autologous MLR #1). After 6 days, T cells were harvested and recultured with freshly thawed autologous, mature moDCs for another 6 days at the same ratio as in autologous MLR #1. T cells were then harvested and added to an allogeneic MLR composed of allogeneic T cells and autologous, mature moDCs, or in some experiments, autologous T cells and allogeneic, mature moDCs. A suppressive effect of T cells, primed by two rounds of autologous moDC stimulation, on the proliferative response of T cells in an allogeneic MLR was assessed by measuring 3HTdR incorporation. Initial priming autologous MLRs were performed with or without the IDO-inhibitor, 1-methyl-tryptophan (1MT), to assess the effect of inhibiting IDO on the generation of suppressor T cells.
- Figure S2. Inhibition of IFN-gamma secretion by CD4+CD25brightFoxp3+ Tregs (JPG, 69.6 KB)
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Autologous T cells stimulated by IDO-expressing, mature moDCs were sorted cytofluorographically to collect CD4+CD25neg, CD4+CD25int, and CD4+CD25bright T cells. These were then added 1:1 with new allogeneic responder T cells stimulated by freshly thawed, mature moDCs, autologous to the candidate Tregs. After 4 days’ culture, supernatant was collected from each culture condition, and IFN-gamma levels were compared between cultures containing no candidate Tregs (black bar), CD4+CD25neg T cells (white bar), CD4+CD25int T cells (gray bar), and CD4+CD25bright T cells (horizontally striped bar) (average of triplicate means ± SEM, n = 3 independent experiments; P values indicated on graph).
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