Skip to main content
. 2022 Jan 21;12:658895. doi: 10.3389/fimmu.2021.658895

Figure 1.

Figure 1

Cholesterol-conjugated dsRNA40 induces a potent immunostimulation. (A) Wt Flt3L-DC were stimulated with 0.5 µM ssRNA40-chol or ssRNA40 complex to DOTAP. Medium, CpG2216 (1 µM) and DOTAP served as controls. Supernatants were taken after 20 h of incubation and cytokine production was analyzed by ELISA. Data were calculated from at least five individual experiments. Bars indicate mean + SEM. *p < 0.05, **p < 0.01, ****p < 0.0001; unpaired t-test. (B) Wt and TLR7-deficient Flt3L-DC were incubated with 0.5 µM dsRNA40-chol, dsRNA40 complexed to DOTAP and dsRNA40-chol-2’OM. Medium ssRNA40 (0.75 µM) complexed to DOTAP, CpG2216 (1 µM) and DOTAP served as controls. Cytokine production was analyzed by ELISA after 20 h of incubation. CpG induced >1000 U/ml IFN-α in wt and TLR7-deficient cells, 6.3 ng/ml IL-6 in wt, 6.9 ng/ml IL-6 in TLR7-deficient cells and 0.8 ng/ml TNF-α in wt and 1.0 ng/ml TNF-α in TLR7-deficient cells. Data were calculated from five to seven individual experiments. Bars indicate mean + SEM. *p < 0.05, **p < 0.01, ***p < 0.001; one-way ANOVA with Sidak’s post-hoc test. (C) Polyacrylamide gel electrophoresis of annealed dsRNA40-chol and corresponding sequences. Nucleic acids were stained with SybrGold. One representative gel out of three is shown. (D) Purified human pDC were stimulated with dsRNA40-chol, dsRNA40 complexed to DOTAP and dsRNA40-chol-2’OM at 2.5 µM, 0.5 µM and 0.1 µM. Medium, ssRNA40 (0.75 µM), CpG2216 (1 µM) and DOTAP served as controls. Supernatants were harvested after 20 h of incubation and IFN-α was analyzed by ELISA. Data were calculated from three independent experiments. Bars indicate mean + SEM. *p < 0.05, ***p < 0.001; one-way ANOVA with Sidak’s post-hoc test.