Fig. 4: RNA-LL37 effects on PMNs depend on TLR8 and are blocked by iODNs.

a Tnf release (ELISA, 5 h stimulation as indicated) by BM-PMNs from different mouse strains (n = 4–8 each). b, c Fluorescence microscopy of fixed and Hoechst and SYTO RNAselect-stained stimulated BM-PMNs from different mouse strains (n = 4 Tlr13^−/−, n = 5 Unc93b1^3d/3d, n = 8 WT; scale bar = 10 µm or n = 4, scale bar = 20 µm). d IL-8 and TNF release (ELISA, 18 h stimulation as indicated) by WT and TLR8 CRISPR-edited BLaER1 cells (n = 7). e NF-κB dual luciferase reporter assay in HEK293T cells, transfected with NF-κB firefly luciferase reporter, Renilla control reporter and TLR8 plasmid and subsequently stimulated with RNA without (arrow) or with IRS661, IRS954, IRS869 and IRS546 (0.15–3.5 µM, n = 2 each). MIP-1β (f, n = 4) release from stimulated PMNs with or without IRS661 (1 nM) or IRS954 (50 nM) pre-incubation (30 min) quantified by ELISA. g Fluorescence microscopy of fixed and Hoechst and anti-ΨU-stained RNA-LL37-stimulated PMNs (n = 3, scale bar = 10 µm). h Quantification of (g). Panels (a), (d), (e), (f), and (h) represent combined data (mean + SD) from ‘n’ biological replicates (each dot represents one mouse or donor). In (b), (c), (e), and (g) one representative of ‘n’ replicates is shown (mean + SD of technical triplicates). *p < 0.05 according to two-way ANOVA with Dunnett correction for multiple testing (e, comparison against ‘no inhibitor’ condition, arrow), one-way ANOVA with Sidak correction (a, d) Friedmann test with Dunn correction (f) or Mann–Whitney test (h). Source data is provided as a Source data file.