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Blood, Vol. 104, Issue 6, 1808-1815, September 15, 2004
Murine plasmacytoid dendritic cells induce effector/memory CD8+ T-cell responses in vivo after viral stimulation
Blood Schlecht et al.
104: 1808
Supplemental materials for: Schlecht et al, Vol 104, Issue 6, 1808-1815
Files in this Data Supplement:
- Figure S1. Virus-activated pDCs activate CD8+ transgenic T-cell responses in vivo (JPG file, 68 KB)
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We determined the ability of male immature, CpG- and virus-activated pDCs to induce HY Tg T-cell proliferation, activation, and differentiation into effector cells in vivo in Rag−/− c−/− hosts. As described in the "T-cell isolation and adoptive transfer" subsection of "Materials and methods," HY Tg T cells were isolated from the LNs of female HY mice, labeled with CFSE, and transferred (4×105 cells) with purified immature, CpG- or virus-activated pDCs or cDCs (5×104 cells) from male 129sv mice in Rag−/−c−/− hosts. Seven days later, splenocytes were recovered and either stained for FACS analysis or used in an ELISPOT assay. (A) CFSE profiles (upper panels) and CD44 markers (lower panels) on CD8+ HY Tg T cells were analyzed. Dot plots show representative results obtained in 1 mouse from each group tested in the experiment. (B) The frequency of HY-specific, IFN-–producing T cells in the spleens of injected mice was quantified by ELISPOT. Each bar corresponds to 1 individual mouse and represents the mean of duplicates ± SD. One representative mouse is depicted out of 3 in each case. As shown in the figure, while immature and CpG-activated pDCs failed to induce efficient proliferation and accumulation of HY T cells, both male cDCs and virus-activated pDCs stimulated most HY T cells to divide more than 7 times and to upregulate CD44 expression. Accordingly, HY-specific IFN-–secreting cells were induced by cDCs and virus-activated pDCs while no effector activity was detected in mice transferred with HY T cells and immature or CpG-activated pDCs.
- Figure S2. Immature pDCs do not exhibit inhibitory activity on activation of CD8+ transgenic T cells by cDCs in vivo (JPG file, 72 KB)
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We tested the inhibitory activity of male immature pDCs on HY Tg T-cell proliferation, activation, and differentiation into effector cells in vivo in Rag−/−c−/− hosts induced by male cDCs. As described in the "T-cell isolation and adoptive transfer" subsection of "Materials and methods," HY Tg T cells were isolated from the LNs of female HY mice, labeled with CFSE, and transferred (4×105 cells) with 8×104 male 129sv cDCs alone or with graded numbers of male 129sv immature pDCs (8×104, 4×104 and 2×104 cells) in Rag−/−c−/− hosts. Seven days later, splenocytes were recovered and either stained for FACS analysis or used in an ELISPOT assay. (A) CFSE profiles (upper panels) and CD44 markers (lower panels) on CD8+ HY Tg T cells were analyzed. Dot plots show representative results obtained in 1 mouse from each group tested in the experiment. Percentages indicated in the upper panels correspond to the percentages of HY T cells that have only slightly proliferated. Percentages indicated in the lower panels correspond to the percentages of HY T cells that have not up-regulated the activation marker CD44. (B) The frequency of HY-specific, IFN-–producing T cells in the spleens of injected mice was quantified by ELISPOT. Each bar corresponds to 1 individual mouse and represents the mean of duplicates ± SD. One representative mouse out of 2 is depicted in each case. As shown in the figure, similar proliferation and activation were obtained when cDCs were injected with or without male pDCs. Accordingly, similar numbers of HY-specific, IFN-–secreting cells were induced by cDCs when injected in the presence or absence of pDCs. These results confirm that immature pDCs have no inhibitory activity on induction of CD8+ T-cell responses.
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