The P2RX7/NLRP3/IL-18 pathway in immune cells is required for HEI3090’s antifibrotic effect
(A) Experimental design. p2rx7-/- mice were given 3.106 WT, nlrp3-/- or il18-/- splenocytes i.v. one day prior to BLM delivery (i.n. 2.5 U/kg). Mice were treated daily i.p. with 1.5 mg/kg HEI3090 or vehicle for 14 days. (B,E,H) Representative images of lung sections at day 14 after treatment stained with H&E and Sirius Red, scale bar= 100 µm. (C, F, I), Fibrosis score assessed by the Ashcroft method and (D, G, J), Collagen fiber intensity. (C) WT splenocytes, p-values were determined by Two-tailed Mann-Whitney test. (D) WT splenocytes, p-values were determined by Two-tailed unpaired t-test. (F, G) nlrp3-/- splenocytes, p-values were determined by Two-tailed Mann-Whitney test. (I, J) il18-/- splenocytes, p-values were determined by Two-tailed Mann-Whitney test. (K) IL-18 levels in sera of WT BLM-induced mice at day 14. P-values were determined by Two-tailed unpaired t-test. (L) Ratio of IFN-γ over IL-17A in lung T cells (CD3+NK1.1-) of WT mice neutralized for IL-18 or not (isotype control) every 3 days. P-values were determined by one-way ANOVA with tukey’s multiple comparisons test. For all analyses, the violin plot illustrates the distribution of Ashcroft scores across indicated groups. The width of the violin at each point represents the density of data, and the central line indicates the median expression level. Each point represents one biological replicate. *p < 0.05, **p < 0.01 WT: Wildtype, BLM: bleomycin, i.p.: intraperitoneal, i.n.: intranasal, i.v.: intravenous, H&E: hematoxylin & eosin.