|
|
Blood, Vol. 112, Issue 4, 1184-1194, August 15, 2008
CD300a/c regulate type I interferon and TNF- secretion by human plasmacytoid dendritic cells stimulated with TLR7 and TLR9 ligands
Blood Ju et al.
112: 1184
Supplemental materials for: Ju et al
Files in this Data Supplement:
- Figure S1. CpG downregulates CD300a and CD300c transcripts by RT-PCR (JPG, 39.4 KB)
-
Purified pDC were cultured for 0, 2, 24, 48 hours with CpG or control ODN in the presence of IL-3. CD300a and CD300c mRNA level were analyzed by RT-PCR. Housekeeping gene β-actin was used as control.
- Figure S2. Surface molecule expression on two subpopulation of tonsil pDC (JPG, 103 KB)
-
Tonsil mononuclear cells were isolated by density gradient centrifugation of a cell suspension made by mechanical dissociation of tonsil tissues. Cells were stained with CMRF-35 mAb, Alexa Fluor 488-conjugated-F(ab′) goat anti-mouse IgG (H+L) and BDCA-4-APC, CMRF-35+BDCA-4+ and CMRF-35−BDCA-4+ cells were gated for analysing the expression of NKp44, CD56, HLA-DR, CD80, CD83, CD86 by flow cytometry. Data were representative of four tonsil samples.
- Figure S3. Surface molecule expression on CMRF35high and CMRF35low pDC (JPG, 79.4 KB)
-
pDC were isolated from peripheral blood and purified with BDCA-4 isolation kit. Cells were cultured with CpG ODN or control ODN in the present of IL-3 for 48 hours. Cells were stained with CMRF-35 mAb and Alexa Fluor 488-conjugated-F(ab′) goat anti-mouse IgG (H+L). CMRF35high and CMRF35low pDC were analysed by staining with HLA-DR-PerCP and CD86-PE with flow cytometry. Data were from one of the four experiments.
- Figure S4. CMRF-35 crosslinking of pDC on CD4+ T-cell proliferation (JPG, 49.7 KB)
-
(A) Tetanus toxoid recall analysis. pDC with or without CMRF-35 crosslinking or crosslinked with control antibody were activated with CpG in the presence of IL-3 for 48 hours, then cocultured with purified CD4+ T lymphocytes from the same donor in the presence of tetanus toxoid (TT). Following 5 days of culture, lymphocyte proliferation was assessed by the addition of 3H-thymidine (1 µCi/well) 16 hours before harvesting (n=3). (B) MLR analysis. pDC with or without CMRF-35 crosslinking or crosslinked with control antibody were activated with CpG for 48 hours, then cocultured with 105 allogeneic CD4+ T cells for 5 days. T cell proliferation was measured by 3H-thymidine uptake. Data are representative of three experiments.
|
|
