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. 2014 Feb 21;33(6):542–558. doi: 10.1002/embj.201386117

Figure 3.

Figure 3

R:D45 activates DCs and induce a systemic cytokine response in vivo in mice and ex vivo in human cells.
  1. Delivery of R:D45 complexed to Invivofectamine in vivo phenotypically activates DCs. C57BL/6 mice were injected intraperitoneally with 80 μg R:D45 or 80 μg R:D45 complexed to Invivofectamine and the activation of splenic DC populations was analysed 12 h later by flow cytometry. Left, representative histograms comparing cell surface expression of the indicated marker on DCs from mice treated with Invivofectamine alone (grey) and R:D45 complexed to Invivofectamine (black). Isotype control shaded grey. Right, MFI values for CD40 (P = 0.0000013), CD80 (P = 0.000106) and CD86 (P = 0.00000024). Data shown are from one experiment ± s.e.m. (n = 5 mice per group), representative of a total of three independent experiments. Large-scale R:D45 hybrid synthesis was performed for each experiment, with purity estimated by PAGE at 96%, 91%, and 98% hybrid, respectively.
  2. R:D45 complexed to Invivofectamine induces cytokine production in vivo. C57BL/6 mice were injected with R:D45 as described for (A). Serum levels of IL-6 (***P = 0.000526) and IFN-α (***P = 0.0000026) were determined 12 h post-injection. Data pooled from three independent experiments ± s.e.m. (10–15 mice total per condition).
  3. R:D45 induces cytokine production when transfected ex vivo in human PBMCs. Freshly isolated PBMCs were transfected with R:D45 complexed to Lipofectamine LTX. Supernatant cytokine levels were quantified 18 h later by ELISA. Data pooled from two independent experiments ± s.e.m., **P = 0.00166 (IL-6), ***P = 0.00011 (IFN-α) (7 donors in total).